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Why would the stroma have a higher pH during photosynthesis?

Why would the stroma have a higher pH during photosynthesis?


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Here is a question form my test.

If you could measure the pH of the interior of thylakoids and the surrounding stroma during active photosynthesis what would be the relative pH of each?

The answer is C.

the stroma would have a higher pH

I understand that proton gradient is from stroma to thylakoid space, but that does not mean stroma would have less protons than thylakoid space, if stroma would have a higher pH, then the proton will never flow from stroma to thylakoid space, right?


Energy from an electron transport chain is used to actively transport protons from the stroma to the thylakoid lumen. Thus the lumen has a relatively higher proton concentration and therefore a lower pH than the stroma.

Higher [H+] means lower pH.

Lower [H+] means higher pH.

Keep in mind that the flow of actively transported protons from the stroma to the lumen is against their concentration gradient. There wouldn't be a net flow of protons this way without active transport.


Chapter 10 - Photosynthesis

  • Life on Earth is solar powered.
  • The chloroplasts of plants use a process called photosynthesis to capture light energy from the sun and convert it to chemical energy stored in sugars and other organic molecules.

Plants and other autotrophs are the producers of the biosphere.

  • Photosynthesis nourishes almost all the living world directly or indirectly.
    • All organisms use organic compounds for energy and for carbon skeletons.
    • Organisms obtain organic compounds by one of two major modes: autotrophic nutrition or heterotrophic nutrition.
    • Autotrophs are the ultimate sources of organic compounds for all heterotrophic organisms.
    • Autotrophs are the producers of the biosphere.
    • Photoautotrophs use light as a source of energy to synthesize organic compounds.
      • Photosynthesis occurs in plants, algae, some other protists, and some prokaryotes.
      • Chemoautotrophs harvest energy from oxidizing inorganic substances, such as sulfur and ammonia.
        • Chemoautotrophy is unique to prokaryotes.
        • These organisms are the consumers of the biosphere.
        • The most obvious type of heterotrophs feeds on other organisms.
          • Animals feed this way.
          • Most fungi and many prokaryotes get their nourishment this way.

          Concept 10.1 Photosynthesis converts light energy to the chemical energy of food

          • All green parts of a plant have chloroplasts.
          • However, the leaves are the major site of photosynthesis for most plants.
            • There are about half a million chloroplasts per square millimeter of leaf surface.
            • Chlorophyll plays an important role in the absorption of light energy during photosynthesis.
            • The interior of the thylakoids forms another compartment, the thylakoid space.
            • Thylakoids may be stacked into columns called grana.
            • Photosynthetic prokaryotes lack chloroplasts.
            • Their photosynthetic membranes arise from infolded regions of the plasma membranes, folded in a manner similar to the thylakoid membranes of chloroplasts.

            Evidence that chloroplasts split water molecules enabled researchers to track atoms through photosynthesis.

            • Powered by light, the green parts of plants produce organic compounds and O2 from CO2 and H2O.
            • The equation describing the process of photosynthesis is:
              • 6CO2 + 12H2O + light energy --> C6H12O6 + 6O2+ 6H2O
              • C6H12O6 is glucose.
              • 6CO2 + 6H2O + light energy --> C6H12O6 + 6O2
              • [CH2O] represents the general formula for a sugar.
              • Before the 1930s, the prevailing hypothesis was that photosynthesis split carbon dioxide and then added water to the carbon:
                • Step 1: CO2 --> C + O2
                • Step 2: C + H2O --> CH2O
                • CO2 + 2H2S --> [CH2O] + H2O + 2S
                • CO2 + 2H2O --> [CH2O] + H2O + O2
                • They used 18O, a heavy isotope, as a tracer.
                • They could label either C18O2 or H218O.
                • They found that the 18O label only appeared in the oxygen produced in photosynthesis when water was the source of the tracer.
                • It reverses the direction of electron flow in respiration.
                • Because the electrons increase in potential energy as they move from water to sugar, the process requires energy.
                • The energy boost is provided by light.

                Here is a preview of the two stages of photosynthesis.

                • Photosynthesis is two processes, each with multiple stages.
                • The light reactions (photo) convert solar energy to chemical energy.
                • The Calvin cycle (synthesis) uses energy from the light reactions to incorporate CO2 from the atmosphere into sugar.
                • In the light reactions, light energy absorbed by chlorophyll in the thylakoids drives the transfer of electrons and hydrogen from water to NADP+ (nicotinamide adenine dinucleotide phosphate), forming NADPH.
                  • NADPH, an electron acceptor, provides reducing power via energized electrons to the Calvin cycle.
                  • Water is split in the process, and O2 is released as a by-product.

                  Concept 10.2 The light reactions convert solar energy to the chemical energy of ATP and NADPH

                  • The thylakoids convert light energy into the chemical energy of ATP and NADPH.
                  • Light is a form of electromagnetic radiation.
                  • Like other forms of electromagnetic energy, light travels in rhythmic waves.
                  • The distance between crests of electromagnetic waves is called the wavelength.
                    • Wavelengths of electromagnetic radiation range from less than a nanometer (gamma rays) to more than a kilometer (radio waves).
                    • Photons are not tangible objects, but they do have fixed quantities of energy.
                    • Photons with shorter wavelengths pack more energy.
                    • Visible light is the radiation that drives photosynthesis.
                    • Different pigments absorb photons of different wavelengths, and the wavelengths that are absorbed disappear.
                    • A leaf looks green because chlorophyll, the dominant pigment, absorbs red and blue light, while transmitting and reflecting green light.
                    • It beams narrow wavelengths of light through a solution containing the pigment and measures the fraction of light transmitted at each wavelength.
                    • An absorption spectrum plots a pigment’s light absorption versus wavelength.
                    • Chlorophyll a, the dominant pigment, absorbs best in the red and violet-blue wavelengths and least in the green.
                    • Other pigments with different structures have different absorption spectra.
                    • An action spectrum measures changes in some measure of photosynthetic activity (for example, O2 release) as the wavelength is varied.
                    • In this experiment, different segments of a filamentous alga were exposed to different wavelengths of light.
                    • Areas receiving wavelengths favorable to photosynthesis produced excess O2.
                    • Engelmann used the abundance of aerobic bacteria that clustered along the alga at different segments as a measure of O2 production.
                    • Chlorophyll b, with a slightly different structure than chlorophyll a, has a slightly different absorption spectrum and funnels the energy from these wavelengths to chlorophyll a.
                    • Carotenoids can funnel the energy from other wavelengths to chlorophyll a and also participate in photoprotection against excessive light.
                    • These compounds absorb and dissipate excessive light energy that would otherwise damage chlorophyll.
                    • They also interact with oxygen to form reactive oxidative molecules that could damage the cell.
                    • The electron moves from its ground state to an excited state.
                    • The only photons that a molecule can absorb are those whose energy matches exactly the energy difference between the ground state and excited state of this electron.
                    • Because this energy difference varies among atoms and molecules, a particular compound absorbs only photons corresponding to specific wavelengths.
                    • Thus, each pigment has a unique absorption spectrum.
                    • If a solution of chlorophyll isolated from chloroplasts is illuminated, it will fluoresce and give off heat.
                    • The solar-powered transfer of an electron from a special chlorophyll a molecule to the primary electron acceptor is the first step of the light reactions.
                    • Photosystem I (PS I) has a reaction center chlorophyll a that has an absorption peak at 700 nm.
                    • Photosystem II (PS II) has a reaction center chlorophyll a that has an absorption peak at 680 nm.
                    • The differences between these reaction centers (and their absorption spectra) lie not in the chlorophyll molecules, but in the proteins associated with each reaction center.
                    • These two photosystems work together to use light energy to generate ATP and NADPH.
                    • Noncyclic electron flow, the predominant route, produces both ATP and NADPH.
                      1. Photosystem II absorbs a photon of light. One of the electrons of P680 is excited to a higher energy state.
                      2. This electron is captured by the primary electron acceptor, leaving the reaction center oxidized.
                      3. An enzyme extracts electrons from water and supplies them to the oxidized reaction center. This reaction splits water into two hydrogen ions and an oxygen atom that combines with another oxygen atom to form O2.
                      4. Photoexcited electrons pass along an electron transport chain before ending up at an oxidized photosystem I reaction center.
                      5. As these electrons “fall” to a lower energy level, their energy is harnessed to produce ATP.
                      6. Meanwhile, light energy has excited an electron of PS I’s P700 reaction center. The photoexcited electron was captured by PS I’s primary electron acceptor, creating an electron “hole” in P700. This hole is filled by an electron that reaches the bottom of the electron transport chain from PS II.
                      7. Photoexcited electrons are passed from PS I’s primary electron acceptor down a second electron transport chain through the protein ferredoxin (Fd).
                      8. The enzyme NADP+ reductase transfers electrons from Fd to NADP+. Two electrons are required for NADP+’s reduction to NADPH. NADPH will carry the reducing power of these high-energy electrons to the Calvin cycle.
                    • Excited electrons cycle from their reaction center to a primary acceptor, along an electron transport chain, and return to the oxidized P700 chlorophyll.
                    • As electrons flow along the electron transport chain, they generate ATP by cyclic photophosphorylation.
                    • There is no production of NADPH and no release of oxygen.
                    • In both organelles, an electron transport chain pumps protons across a membrane as electrons are passed along a series of increasingly electronegative carriers.
                    • This transforms redox energy to a proton-motive force in the form of an H+ gradient across the membrane.
                    • ATP synthase molecules harness the proton-motive force to generate ATP as H+ diffuses back across the membrane.
                    • When chloroplasts are illuminated, the pH in the thylakoid space drops to about 5 and the pH in the stroma increases to about 8, a thousandfold different in H+ concentration.
                    • This process also produces ATP and oxygen as a by-product.

                    Concept 10.3 The Calvin cycle uses ATP and NADPH to convert CO2 to sugar

                    • The Calvin cycle regenerates its starting material after molecules enter and leave the cycle.
                    • The Calvin cycle is anabolic, using energy to build sugar from smaller molecules.
                    • Carbon enters the cycle as CO2 and leaves as sugar.
                    • The cycle spends the energy of ATP and the reducing power of electrons carried by NADPH to make sugar.
                    • The actual sugar product of the Calvin cycle is not glucose, but a three-carbon sugar, glyceraldehyde-3-phosphate (G3P).
                    • Each turn of the Calvin cycle fixes one carbon.
                    • For the net synthesis of one G3P molecule, the cycle must take place three times, fixing three molecules of CO2.
                    • To make one glucose molecule requires six cycles and the fixation of six CO2 molecules.
                    • The Calvin cycle has three phases.

                    Phase 1: Carbon fixation

                    • In the carbon fixation phase, each CO2 molecule is attached to a five-carbon sugar, ribulose bisphosphate (RuBP).
                      • This is catalyzed by RuBP carboxylase or rubisco.
                      • Rubisco is the most abundant protein in chloroplasts and probably the most abundant protein on Earth.
                      • The six-carbon intermediate is unstable and splits in half to form two molecules of 3-phosphoglycerate for each CO2.

                      Phase 2: Reduction

                      • During reduction, each 3-phosphoglycerate receives another phosphate group from ATP to form 1,3-bisphosphoglycerate.
                      • A pair of electrons from NADPH reduces each 1,3-bisphosphoglycerate to G3P.
                        • The electrons reduce a carboxyl group to the aldehyde group of G3P, which stores more potential energy.
                        • One of these six G3P (3C) is a net gain of carbohydrate.
                          • This molecule can exit the cycle and be used by the plant cell.

                          Phase 3: Regeneration

                          • The other five G3P (15C) remain in the cycle to regenerate three RuBP. In a complex series of reactions, the carbon skeletons of five molecules of G3P are rearranged by the last steps of the Calvin cycle to regenerate three molecules of RuBP.
                          • For the net synthesis of one G3P molecule, the Calvin cycle consumes nine ATP and six NADPH.
                          • The light reactions regenerate ATP and NADPH.
                          • The G3P from the Calvin cycle is the starting material for metabolic pathways that synthesize other organic compounds, including glucose and other carbohydrates.

                          Concept 10.4 Alternative mechanisms of carbon fixation have evolved in hot, arid climates

                          • One of the major problems facing terrestrial plants is dehydration.
                          • At times, solutions to this problem require tradeoffs with other metabolic processes, especially photosynthesis.
                          • The stomata are not only the major route for gas exchange (CO2 in and O2 out), but also for the evaporative loss of water.
                          • On hot, dry days, plants close their stomata to conserve water. This causes problems for photosynthesis.
                          • In most plants (C3 plants), initial fixation of CO2 occurs via rubisco, forming a three-carbon compound, 3-phosphoglycerate.
                            • C3 plants include rice, wheat, and soybeans.
                            • The two-carbon fragment is exported from the chloroplast and degraded to CO2 by mitochondria and peroxisomes.
                            • Unlike normal respiration, this process produces no ATP.
                              • In fact, photorespiration consumes ATP.
                              • In fact, photorespiration decreases photosynthetic output by siphoning organic material from the Calvin cycle.
                              • The inability of the active site of rubisco to exclude O2 would have made little difference.
                              • Photorespiration can drain away as much as 50% of the carbon fixed by the Calvin cycle on a hot, dry day.
                              • Several thousand plants, including sugarcane and corn, use this pathway.
                              • Bundle-sheath cells are arranged into tightly packed sheaths around the veins of the leaf.
                              • Mesophyll cells are more loosely arranged cells located between the bundle sheath and the leaf surface.
                              • PEP carboxylase has a very high affinity for CO2 and can fix CO2 efficiently when rubisco cannot (i.e., on hot, dry days when the stomata are closed).
                              • The bundle-sheath cells strip a carbon from the four-carbon compound as CO2, and return the three-carbon remainder to the mesophyll cells.
                              • The bundle-sheath cells then use rubisco to start the Calvin cycle with an abundant supply of CO2.
                              • These plants are known as CAM plants for crassulacean acid metabolism.
                              • They open their stomata during the night and close them during the day.
                                • Temperatures are typically lower at night, and humidity is higher.
                                • In C4 plants, carbon fixation and the Calvin cycle are spatially separated.
                                • In CAM plants, carbon fixation and the Calvin cycle are temporally separated.

                                Here is a review of the importance of photosynthesis.

                                • In photosynthesis, the energy that enters the chloroplasts as sunlight becomes stored as chemical energy in organic compounds.
                                • Sugar made in the chloroplasts supplies the entire plant with chemical energy and carbon skeletons to synthesize all the major organic molecules of cells.
                                  • About 50% of the organic material is consumed as fuel for cellular respiration in plant mitochondria.
                                  • Carbohydrate in the form of the disaccharide sucrose travels via the veins to nonphotosynthetic cells.
                                    • There, it provides fuel for respiration and the raw materials for anabolic pathways, including synthesis of proteins and lipids and formation of the extracellular polysaccharide cellulose.
                                    • Cellulose, the main ingredient of cell walls, is the most abundant organic molecule in the plant, and probably on the surface of the planet.
                                    • Starch is stored in chloroplasts and in storage cells in roots, tubers, seeds, and fruits.
                                    • It is responsible for the presence of oxygen in our atmosphere.
                                    • Each year, photosynthesis synthesizes 160 billion metric tons of carbohydrate.

                                    Lecture Outline for Campbell/Reece Biology, 7th Edition, © Pearson Education, Inc. 10-1


                                    1st PUC Biology Photosynthesis in Higher Plants NCERT Text Book Questions and Answers

                                    Question 1.
                                    By looking at a plant externally can you tell whether a plant is C3 or C4? Why and how?
                                    Answer:
                                    C4 plants are adapted to the xerophytic climatic conditions they can grow well in high temperature. It cannot be said conclusively that the plant is a C3 or C4 by looking at external appearance, some guess can be made by looking at fleshy leaf structure of C4 plants.

                                    Question 2.
                                    By looking at which internal structure of a plant can you tell whether a plant is C3 or C4? Explain.
                                    Answer:
                                    The particularly large cells around the vascular bundles of the C4 pathway plants are called bundle sheath cells, and the leaves which have such anatomy are said to have ‘Kranz1 anatomy. ‘Kranz’ means ‘wreath’ and is a reflection of the arrangement of cells. The bundle sheath cells may form several layers around the vascular bundles, they are characterized by having a large number of chloroplasts, thick walls impervious to gaseous exchange and no intercellular spaces.

                                    Question 3.
                                    Even though very few cells In a C4 plant carry out the biosynthetic – Calvin pathway, yet they are highly productive. Can you discuss why?
                                    Answer:

                                    1. In C4 plants, the biosynthetic Calvin cycle occurs only in bundle sheaths. Despite few cells performing the Calvin cycle in C4 plants, they are highly productive due to minimum photorespiration losses.
                                    2. They are adopted to diverse climatic conditions as C4 plants can synthesize at very low CO2 concentration while for C3 plants CO2 concentration is the limiting factor.
                                    3. C4 plants can synthesize at high temperatures while C3 plants cannot.
                                    4. Rapid withdrawal of photosynthates from the bundle sheath cells as they lie over the vascular bundles.
                                    5. Photosynthesis continues even when stomata are closed due to the fixation of CO2 released through respiration.

                                    Question 4.
                                    RuBisCO is an enzyme that acts both as a carboxylase and oxygenase. Why do you think RuBIsCO carries out more carboxylation in C4 plants?
                                    Answer:
                                    RuBisCo has a much greater affinity for CO2 than for O2. It is the relative concentration of O2 and CO2 that determines which of the two will bind to the enzyme. In C3 plants some O2 does bind to RuBisCo and hence CO2 fixation is decreased.

                                    In C4 plants photorespiration does not occur. This is because they have a mechanism that increases the concentration of CO2 at the enzyme site. This takes place when the C4 acid from the mesophyll is broken down in the bundle cells to release CO2. This results in increasing the intracellular concentration of CO2. In turn, this ensures that the RuBisCo functions as a carboxylase minimizing the oxygenase activity.

                                    Question 5.
                                    Suppose there were plants that had a high concentration of Chlorophyll b but lacked chlorophyll a, would It carry out photosynthesis? Then why do plants have chlorophyll b and other accessory pigments?
                                    Answer:
                                    No, photosynthesis occurs in plants having a high concentration of chlorophyll ‘b’ but lacks chlorophyll ‘a’ because chlorophyll ‘a’ molecule forms reaction center in both photosystems I and II which converts light energy into electrical energy and excites the electrons for photolysis of water.

                                    Maximum photosynthesis occurs at the wavelengths which is absorbed by chlorophyll ‘a’ molecule i.e. blue and red regions.

                                    Though chlorophyll a is the major pigment responsible for trapping light, other thylakoid pigments like chlorophyll b, xanthophylls, and carotenoids, which are called accessory pigments, also absorb light and transfer the energy to chlorophyll a.

                                    Indeed they not only enable a wider range of wavelengths of incoming light to be utilized for photosynthesis but also protect chlorophyll from photo-oxidation.

                                    Question 6.
                                    Why is the colour of a leaf kept In the dark frequently yellow, or pale green? Which pigment do you think Is more stable?
                                    Answer:
                                    We can look for an answer to these questions by trying to separate the leaf pigments of any green plant through paper chromatography. Chromatographic separation of the leaf pigments shows that the color that we can see in leaves is not due to a single pigment but due to four pigments: Chlorophyll (a) (bright or blue-green in the chromatogram), Chlorophyll (b) (yellow-green), xanthophylls (yellow) and carotenoids (yellow to yellow-orange). Chlorophyll (a) is the chief pigment associated with photosynthesis.

                                    Question 7.
                                    Look at leaves of the same plant on the shady side and compare it with the leaves on the sunny side. Or, compare the potted plants kept in the sunlight with those in the shade. Which of them has leaves that are darker green? Why?
                                    Answer:
                                    In sunny plant colour of leaves is darker green because in sunny plant photosynthesis takes place while in shady plant rate of photosynthesis is low.

                                    Question 8.
                                    The figure shows the effect of light on the rate of photosynthesis. Based on the graph, answer the following questions

                                    (a) At which point/s (A, B, or C) in the curve is light a limiting factor?
                                    Answer:
                                    Points B-C of the curve, the rate did not increase with an increase in its concentration because under these conditions, the light becomes a limiting factor.

                                    (b) What could be the limiting factor/s in region A?
                                    Answer:
                                    The rate of photosynthesis shows a proportionate increase up to a certain CO2 concentration (In region A of the curve), beyond which the rate again becomes constant, not showing any increase by increasing CO2 concentration.

                                    (c) What do C and D represent on the curve?
                                    Answer:
                                    If the light intensity is doubled, i.e., the plants are exposed to 2 Units of light, CO2 concentration again becomes a limiting factor beyond this concentration (Points C and D represent on the curve.

                                    Question 9.
                                    Give a comparison between the following:

                                    1. C3 and C4 Pathways
                                    2. Cyclic and non-cyclic photophosphorylation
                                    3. Anatomy of leaf in C3 and C4 plants.

                                    Answer:
                                    (1) Difference between C3 and C4 pathways:

                                    1. The primary acceptor of CO2 is RUBP, a 5 carbon compound
                                    2. The first stable product is a 3-PGA a 3 carbon compound.
                                    3. It operates under a low concentration of CO2 in mesophyll cells.
                                    4. CO2 once fixed is not released back.
                                    5. There is a net grin of one molecule of glucose with consumption of 6 CO2, molecules.
                                    6. Fixation of one molecule of CO2 needs 3ATP and 2NADPH2, molecules. Thus C3, pathway requires 18 ATP for the synthesis of one molecule of glucose.
                                    7. C3 cycle operates in all categories of plants.
                                    8. Rate of CO2 fixation in slow
                                    9. The optimum temperature for the operation of the Calvin cycle is 10-25° C.

                                    C4 Pathway or Hatch & Slack:

                                    1. The primary acceptor of CO2 is a PEP, a 3 carbon compound
                                    2. The first stable product is oxaloacetic acid, a 4 carbon compound.
                                    3. It can operate under very low CP2, concentration in mesophyll cells.
                                    4. CO2 once fixed is released back in bundle sheath cells which are finally fixed and reduced by the Calvin cycle.
                                    5. There is no net gain. It rather involves additional consumption of 12 ATP molecules per glucose molecule synthesized.
                                    6. Fixation of one molecule of CO2 needs 2ATP molecules in addition to that required in the C3 cycle. The C4 pathway requires 30 ATP for the synthesis of one molecule of glucose.
                                    7. C4 cycle operates only in C4 plants.
                                    8. The rate of CO2, fixation is faster.
                                    9. The optimum temperature for the operation of C4, the cycle is 30-45° C

                                    (2) Differences between Cyclic and Non-cyclic photophosphorylation:

                                    1. It is performed by photosystem I independently in stromal or thylakoids.
                                    2. In this, an external source of electrons is not required because the same electrons get recycled.
                                    3. It is not connected with the photolysis of water and thus no oxygen is evolved.
                                    4. In this an electron expelled by the exciting photo center P700 is returned to it after passing through a series of electron carriers in an ETS, hence it is called cyclic photophosphorylation.
                                    5. It is activated by light of 700nm wavelength.
                                    6. It generates ATP only. There is no formation of NADPH,
                                    7. Chlorophyll does not receive any electrons from water.
                                    8. It operates under the low intensity of light, anaerobic conditions, or when CO2, availability is poor.
                                    9. The system does not take part in photosynthesis except in certain bacteria.
                                    1. It is carried out collectively both by PSI & PSII photosystems in the grana thylakoids.
                                    2. This process required an external electron donor.
                                    3. It is connected with the photolysis of water and thus oxygen is evolved in it.
                                    4. In this an electron expelled by the exciting photo center P680 is not returned to it after passing through a series of electron carriers, but reaches NADP hence it is called non-cyclic photophosphorylation. In this water is the ultimate source of electrons and NADP + is the final acceptor.
                                    5. In this, the photo centers absorb the light of 680 nm as well as 700nm wavelength.
                                    6. It produces both ATP as well as NADPH2
                                    7. The ultimate source of electrons is the photolysis of water.
                                    8. It operates under optimum light, aerobic conditions, and in the presence of CO2
                                    9. This system is connected with CO2 fixation and is dominant in green plants.

                                    (3) Differences between the anatomy of leaf in C3 and C4 plants:

                                    C3, leal anatomy:
                                    The rate of photosynthesis is influenced by leaf anatomy as it greatly affects the availability of sunlight, the rate of diffusion of CO2, into the mesophyll cells, and the translocation of end products of photosynthesis. The important anatomical features that influence the rate of photosynthesis are the thickness of cuticle, number and distribution of stomata, degree of opening of stomata, the size and number of chloroplasts, size and distribution of intercellular spaces, and number and distribution of vascular strands.

                                    C4 Kranz anatomy:
                                    In some tropical grasses, the cells of bundle sheath around the vascular strand in the leaves are large green, and barrel-shaped. They are surrounded by one or more concentric layers of mesophyll cells. The mesophyll and bundle sheath cells are connected by plasmodesmata. The chloroplasts in the bundle sheath cells are large but do not have well-defined grana.

                                    1st PUC Biology Photosynthesis in Higher Plants Additional Questions and Answers

                                    1st PUC Biology Photosynthesis in Higher Plants One Mark Questions

                                    Question 1.
                                    What is the name of the green plastid?
                                    Answer:
                                    Chloroplast (Oct. 83)

                                    Question 2.
                                    Where does oxygen liberated during photosynthesis come from? (Oct. 90)
                                    Answer:
                                    Water

                                    Question 3.
                                    Which one is the most important limiting factor in photosynthesis?
                                    Answer:
                                    Carbon dioxide.

                                    Question 4.
                                    Who proposed the Law of Limiting Factors?
                                    Answer:
                                    F.F. Blackman (Apr. 1991,1999)

                                    Question 5.
                                    In which reaction of photosynthesis oxygen is released? (Oct. 1994)
                                    Answer:
                                    Photolysis of water

                                    Question 6.
                                    Mention the two ways in which Ca++ is involved in cell division in plants. Where are the photosynthetic pigments located in a chloroplast?
                                    Answer:
                                    In the thylakoid membrane.

                                    Question 7.
                                    What are Quantasomes? (Apr. 1997)
                                    Answer:
                                    Quantasomes is a functional unit (photosynthetic units) made of a group of pigment molecules required for carrying out a photochemical reaction.

                                    Question 8.
                                    Define Photophosphorylation. (Oct. 1997, Apr. 2000)
                                    Answer:
                                    The synthesis of ATP in the presence of light is called Photophosphorylation.

                                    Question 9.
                                    What is a C3 plant? (M.Q.P.)
                                    Answer:
                                    A C3 plant is one in which the first stable compound obtained during the dark reaction is a three-carbon compound.

                                    Question 10.
                                    Define photolysis of water. (Oct. 1998, 2000, M.Q.P.)
                                    Answer:
                                    Photolysis of photoionization is the splitting of water into protons, electrons, and oxygen in presence of light.

                                    Question 11.
                                    What does the variegated leaf experiment of photosynthesis prove?
                                    Answer:
                                    It proves that chlorophyll is necessary for photosynthesis.

                                    Question 12.
                                    Name the first stable product of the Calvin cycle. (Oct. 2003, July 2007, 2009)
                                    Answer:
                                    PGA – Phosphoglyceric Acid.

                                    Question 13.
                                    What is the CAM pathway?
                                    Answer:
                                    The fixation of CO2 obtained from organic acid like malic acid in members of the family Crassulaceae is called the CAM pathway.

                                    Question 14.
                                    What are C4 plants?
                                    Answer:
                                    The plants produce 4 – carbon compounds as the first stable substances during the dark reaction are called C4 plants.

                                    Question 15.
                                    Expand NADP. (April 2004)
                                    Answer:
                                    Nicotinamide Adenine Dinucleotide Phosphate.

                                    Question 16.
                                    CAM plants close their stomata during day time. Give reason. (April 2007)
                                    Answer:
                                    These plants are mainly xerophytes which open their stomata at night when temperatures are low and close stomata during daytime when temperatures are high as a mechanism to conserve water.

                                    Question 17.
                                    Who discovered the C4 cycle?
                                    Answer:
                                    Hatch and Slack.

                                    Question 18.
                                    Give reason: (March 2008)
                                    Very high temperature decreases the rate of photosynthesis.
                                    Answer:
                                    Chlorophyll undergoes photo-oxidation/ solarization at high temperature reducing photosynthesis.

                                    Question 19.
                                    Give reason: (July 2008, April 2009)
                                    Carotenoid and Xanthophyli are called accessory photosynthetic pigments.
                                    Answer:
                                    Carotenoid and Xanthophyli transfer the absorbed light to chlorophyll ‘a1, hence are called accessory pigments. They cannot release electrons on their own and require chlorophyll.

                                    1st PUC Biology Photosynthesis in Higher Plants Two Marks Questions

                                    Question 1.
                                    Write any two differences between Light and Dark Reactions of Photosynthesis. (Mar. 1988)
                                    Answer:
                                    (i) Light reaction – (a) Dark reaction

                                    • Takes place in presence of light.
                                      (a) Independent of Light.
                                    • Mainly Photolysis and ATP synthesis with O4 evolution.
                                      (b) Mainly concerned with carbon fixation.

                                    Question 2.
                                    Why does chlorophyll appear red in reflected light and green in transmitted light? Explain the significance of these phenomena in terms of photosynthesis.
                                    Answer:
                                    In reflected light, the chlorophyll appears red because of fluorescence. The light absorbed by chlorophyll molecules loses its energy and emits light of wavelengths corresponds to red colour. In transmitted light, chlorophyll appears green because it absorbs only light of wavelengths corresponds to green colour.

                                    Question 3.
                                    Mention the stages of light reaction. (M.Q.P.)
                                    Answer:
                                    The stages of light reaction are

                                    • Photoexcitation of chlorophyll
                                    • Photolysis of water
                                    • Photophosphorylation
                                    • Reduction of NADP

                                    Question 4.
                                    How does temperature influence the biosynthetic phase of photosynthesis?
                                    Answer:
                                    Influences of temperature on the biosynthetic phase of photosynthesis:

                                    • At higher temperature enzymes become inactive as it gets denatured.
                                    • At low temperatures also enzyme becomes inactive.
                                    • (Affinity of the enzymes for C02 decreases with increasing temperature.

                                    Question 5.
                                    Mention any two differences between Photosystem I and Photosystem II. (Oct. 1999)
                                    Answer:
                                    (i) Photosystem I – (a) Photosystem II

                                    1. The reaction centre is P 700 and absorbs red light of 700 nm efficiently.
                                      (a) Reaction centre is P680 and absorbs light of 680 nm efficiently.
                                    2. Located on the unstacked stroma thylakoids and regions of grana facing the stroma.
                                      (b) Located mostly on the stacked membranes of the thylakoids

                                    Question 6.
                                    Define Blackman’s law of limiting factors. (Apr. 2000, Oct. 2002)
                                    Answer:
                                    When “a process is conditioned as to its rapidity by a number of separate factors, the rate of the process is limited by the pace of the slowest factor”.

                                    Question 7.
                                    Why do scientists expect faster growth and more yield by C3 plants, if the atmospheric CO2 increases?
                                    Answer:
                                    If the concentration of CO2 in the atmosphere increases, the rate of photosynthesis by C3 plants will increase for the following two reasons.
                                    (a) High availability of substrate (CO2) for carboxylation.
                                    (b) Photorespiration is reduced due to more availability of CO2 as the enzyme will function only as a carboxylase.

                                    Question 8.
                                    List out any two differences between photo-phosphorylation and oxidative phosphorylation. (Apr. 2003)
                                    Answer:

                                    1. Photophosphorylation takes place in the presence of light, whereas Oxidative phosphorylation takes place in the presence of oxygen.
                                    2. Photophosphorylation occurs in the chloroplast during photosynthesis, whereas Oxidative occurs in the mitochondria during respiration.

                                    Question 9.
                                    Expand PEP. Where is it produced in C4 plants? What is its role in the biosynthetic process?
                                    Answer:
                                    PEP – phosphoenolpyruvate. It is produced in the mesophyll cells of the leaves of C4 plants. It is the primary acceptor of carbon dioxide and is converted into oxaloacetic acid (OAA). Thus it helps in carbon fixation in these plants. By this pathway, the carbon dioxide concentration in the bundle sheath increases, and photorespiration is prevented from occurring.

                                    Question 10.
                                    Draw a neat labelled diagram of the ultrastructure of T.S. of the chloroplast. (March 2008)
                                    Answer:


                                    Chloroplast is lens-shaped cell organelles located in the mesophyll cells. The chloroplast is bounded by a double membrane separated by the intermembrane space. The centre encloses a homogeneous gel-like fluid called stoma or matrix. It has 50% of the chloroplast proteins, Ribosomes, DNA, and enzymes of dark reaction.

                                    The matrix has a system of lamellae differentiated as grana lamellae made of flattened sacs called thylakoids arranged as a stack of coins. The thylakoid membrane has an outer stromal surface and an inner luminal surface in contact with the thylakoid lumen. Some of the lamellae are unstacked and connect the grana called stroma lamellae or inter grana lamellae. The grana lamellae form the site for light reaction and stroma from the site for a dark reaction.

                                    1st PUC Biology Photosynthesis in Higher Plants Five Mark Questions

                                    Question 1.
                                    Define Photosynthesis and explain the light reaction of Photosynthesis.
                                    (Apr. 1983, 1987, 1993, 1995, 1997, 1999, 2006, Oct. 1985, 1991, 1992, M.Q.P.)
                                    Answer:
                                    Photosynthesis is defined as the process in which carbohydrates are synthesised from CO2 and H2O by green plants using radiant energy of the sun, O2 being a byproduct.

                                    Light Reaction: Also called Photochemical reaction and is light-dependent.
                                    The light reaction can be summarized in 4 steps

                                    • Photoexcitation of chlorophyll
                                    • Photolysis of water
                                    • Photophosphorylation
                                    • Photoreduction of NADP

                                    Photoexcitation of chlorophyll: When a chlorophyll molecule is exposed to light it absorbs radiant energy and the electron in its structure picks up energy and becomes a high energy electron. The energy moves rapidly through the light-harvesting pigment molecules to reach the trie reaction centre.

                                    It causes an electron to acquire a large amount of energy and escapes from the reaction centre leaving the chlorophyll with a net positive charge.

                                    Photolysis or Photoionisation of water:
                                    The source of Oxygen was earlier thought to be CO2 but by the work of Van Neil, Ruben, and Kamen using O 18 it was proved that it comes from H2O.

                                    Water splits up in the Manganese containing Oxygen evolving complex under the influence of light to produce protons, electrons, oxygen and water. The overall equation is represented as

                                    Theoretically, 8 quanta of light are required to produce one molecule of oxygen from water.

                                    Photophosphorylation: It is the process by which ATP is synthesised in the presence of light. It was first discovered by Arnon. The process is further differentiated into cyclic and non-cyclic depending on the path taken by the electron. In cyclic the path traversed is cyclic and in non-cyclic, the path traversed is non-cyclic.

                                    Non-cyclic Photophosphorylation:
                                    Non-cyclic Photophosphorylation is initiated by the absorption of a photon of light by PS 11 whose reaction centre is P680 and results in the ejection of an electron creating a hole. The ejected electron is trapped by pheophytin, passes to plastoquinone (PQ) and then takes a downhill path along electron carriers cyt b6 cyt f and plastocyanin and reaches PSI with reaction centre P700.

                                    Absorption of light by PSI now causes the ejection of the electron released from PSII which is trapped by FRS moves to Ferredoxin and using the protons released by photolysis NADP is reduced to NADPH2.

                                    The hole created in PSII due to loss of the electron is filled by electrons produced during photolysis. Photolysis is aided by the Mn-containing oxygen-evolving complex. The path of electrons is from water to PSII, PSII to PSI and PSI to NADP in a zig-zag manner called a pathway.

                                    In non-cyclic photophosphorylation, the ejected electron does not come back to PSII hence the path is non-cyclic, ATP is synthesised between cyt b6 and cyt f, photolysis takes place, and photoreduction of NADP to NADPH2 takes place. (July 2006)

                                    Cyclic Photophosphorylation:
                                    Cyclic Photophosphorylation occurs when additional ATP molecules are required and is a supplementary process. Only PSI with reaction centre P700 is activated. The ejected electron of PSI is captured by Ferredoxin and cycled back through a series of electron carriers cyt b6, cyt f, and plastocyanin. During the downhill jour

                                    A schematic diagram of the Non cyclic photophosphorylation and Non cyclic ETS. Abbrevations: Ph = Pheophytin, Q = Quinone, PQ = Plastoquinone, PQH2 = Plastohydro quinone, cytb6 = Cytochrome b6, cytb6= Cytochrome f, PC = Plastocyanin, A1, A2, A3 = electron acceptors of PSI, Fd.= Ferridoxin, NADP = Nicotinamide adenine dinucleotide phosphate.

                                    ney of the electron free energy is utilised for ATP synthesis at 2 places i.e., between ferredoxin and cyt b6 and cyt b6 and cyt f. The process does not require photolysis of water and NADPH2 formation.

                                    Photoreduction of NADP:
                                    This takes place during non-cyclic Photoohosphorylation and NADPH2 is an excellent reducing agent. ATP and NADPH2 are called the assimilatory powers which are utilized during the dark reaction.

                                    Question 2.
                                    Draw labelled diagrams and describe the structure and functions of the chloroplast.
                                    (Apr. 1983, Oct. 2001, April 2004, 2006, March 2011)
                                    Answer:

                                    The chloroplast is lens-shaped cell organelles located in the mesophyll cells. The chloroplast is bounded by a double membrane separated by the intermembrane space. The centre encloses a homogeneous gel-like fluid called stoma or matrix. It has 50% of the chloroplast proteins, Ribosomes, DNA, and enzymes of dark reaction.

                                    The matrix has a system of lamellae differentiated as grana lamellae made of flattened sacs called thylakoids arranged as a stack of coins. The thylakoid membrane has an outer stromal surface and an inner luminal surface in contact with the thylakoid lumen. Some of the lamellae are unstacked and connect the grana called stroma lamellae or inter grana lamellae. The grana lamellae form the site for light reaction and stroma from the site for a dark reaction.

                                    Question 3.
                                    Explain the Calvin cycle of photosynthesis.
                                    OR
                                    Describe the essential steps of the dark reaction of photosynthesis. (April 84, 94, 96, 98, 03, Oct. 84, 90, 2001)
                                    OR
                                    Give the schematic representation of the Calvin cycle. (March 2009, 2010)
                                    Write the schematic representation of the C3-pathway. (July 2011)
                                    Answer:
                                    Calvin cycle or dark reaction or C3 cycle or thermochemical reaction forms the second step of photosynthesis which takes place independent of light. The steps involved here studied by Calvin using C14.

                                    Dark reaction mainly includes the fixation of carbon to produce the carbohydrate. This occurs in the stroma and utilises the assimilatory powers produced during the light reaction.
                                    The steps involved may be summarised as follows.

                                    1. First Phosphorylation: The starting compound RuMP(Ribulose Mono Phosphate) is converted to RuBP(Ribulose biphosphate) utilizing 6 molecules of ATP from the light reaction. 6 molecules of RuBP are obtained from 6 molecules of RuMP.
                                    2. Carbon fixation by RUBP: The CO2 is accepted by RuBP the primary acceptor to give 12 molecules of PGA(phosphoglycerate) a 3-carbon compound The 6-carbon compound is highly unstable and dissociates to form the 3-carbon compound.
                                    3. Second Phosphorylation: The 12 molecules of PGA utilize 12 ATP molecules to yield 12 Di PGA. The ATP used is produced during the light reaction.
                                    4. Reduction: 12 Di PGA combine with NADPH2 of the light reaction and are reduced to 12 PGAL(phosphoglyceraldehyde) a-3-carbon compound. The other products are 12iP, 12NADP, and H2O.

                                    Out of the 12 PGAL molecules, only 2 are transported to the cytoplasm and used for the formation of a sugar molecule (hexose) and the remaining two molecules are used to regenerate RuBP.

                                    Synthesis of sugar involves condensation of 2PGAL to give fructose 1-6-diphosphate which by dephosphorylation forms Fructose 6-Phosphate and by isomerization forms, glucose and finally is stored as sucrose.

                                    Regeneration of RuBP involves the formation of intermediates like Erythrose monophosphate, xylulose monophosphate, Sedoheptulose monophosphate, Ribose monophosphate, and finally Ribulose monophosphate-RuMP which gives RuBP.

                                    The overall equation of dark reaction is

                                    Question 4.
                                    Give the importance of C4 plants.
                                    Answer:
                                    The importance of C4 plants are:

                                    1. C4 plants have little photorespiration.
                                    2. C4 plants are more efficient in picking up CO2 even when it is found in low concentration because of the high affinity of phosphoenolpyruvate (i.e. PEP).
                                    3. The concentric arrangement of mesophyll cells produces a smaller area in relation to volume for better utilization of available water and reduce the intensity of solar radiation.
                                    4. They are adapted to high temperatures and intense radiation.
                                    5. It prevents photorespiration.

                                    Question 5.
                                    Schematically explain non-cyclic photophosphorylation. (Apr. 2002,2007)
                                    Answer:
                                    Non-cyclic photophosphorylation is the ATP synthesis in the presence of light following a non-cyclic pathway. It requires both PSI and PSII+ work in sequence and is linked by electron carriers of the electron transport system. The path of electrons is from H2O to PSII, PSII to PSI, and PSI to NADP.

                                    When a photon of light is absorbed by PSII its electron is boosted to a high energy level P680* and transferred to pheophytin which acts as the primary electron acceptor. The electron is then transferred to quinone on the stromal surface of the thylakoid membrane which by accepting a second electron and 2H+ from stroma forms H2. The H2 dissociates releasing 2e – on the luminal side to cyt b complex and 2H+ to the lumen to add to the proton pool. Electrons released by cyt b/f complex move to PC which diffuses through the lumen to P700 of PSI.

                                    The PSI is now activated and the electron from PSII is boosted to P700* and is passed along A0, A1, Fx, Fab and is transferred to Fd on the stromal surface and gets reduced. Reduced ferredoxin transfers its electrons to NADP+ which absorbs 2e- and H+ from stroma to produce NADPH and 1. excellent reducing agent by photoreduction. The electron gap formed in P680* is filled by electrons produced by photolysis through OEC. The electron flow is non-cyclic, H2 is used to produce NADPH, ATP is formed from ADP, O2 from H2O, and water.

                                    A schematic diagram of the Non cyclic photophosphorylation and Non cyclic ETS. Abbrevations : Ph = Pheophytin, Q = Quinone, PQ = Plastoquinone, PQH2 = Plastohydro quinone, cytb6 = Cytochrome b6, cyt f = Cytochrome f, PC = Plastocyanin, A1, A2, A3 = electron acceptors of PSI, Fd = Ferridoxin, NADP = Nicotinamide adenine dinucleotide phosphate.

                                    Question 6.
                                    Explain Photoexcitation and photolysis.
                                    Answer:
                                    The absorption of light energy and ejection of electrons by the chlorophyll is called photoexcitation. The chlorophyll and accessory pigments of LHC of both PSI and PSII absorb radiant energy in the form of photons and transfer their absorbed energy to the reaction centre. The reaction centre of PSI and PSII gets excited and ejects its outer valance electron and the chlorophyll becomes oxidized. The electron given out by the chlorophyll is accepted by the electron carrier molecule of each photosystem. So by giving out an electron, chlorophyll becomes oxidized and the electron carrier molecule is reduced. The electron that escaped from the excited chlorophyll is a high-energy electron, so that it carries a lot of energy.

                                    Photolysis of water (Photooxidation of H2O) Lysis of water into a proton (H + ), electrons (e ), and molecular O2 in the presence of light is called photolysis of water. Water by donating electron becomes oxidized.

                                    When light falls on PSII, its reaction centre (Chla680) gets excited by ejecting an electron. Water splits in the Mn 2+ associated Z – protein-containing oxygen-evolving complex under the influence of the light entering photosystem II. The protons (H + ) released into the thylakoid lumen, electrons (e – ) are used to fill a gap created in P680 and molecular O2 is released.

                                    Question 7.
                                    Write a note on photosynthetic pigments.
                                    Answer:
                                    The pigments which they absorb light during photosynthesis are called photosynthetic pigments. They are,
                                    (1) Chlorophyll: Chlorophyll pigments are of various types. Chlorophyll a, b, c, d, e, bacteriochlorophyll etc.

                                    Chlorophyll:
                                    It is the primary photosynthetic pigment. The chemical formula of chlorophyll is C55H72O5N4Mg. It contains a methyl group (CH3). Chlorophyll is called primary photosynthetic pigment because it not only absorbs light but it converts absorbed light energy into chemical energy. Chi absorbs red light and blue light.

                                    (2) Accessory pigments: The pigments absorb certain wavelengths of light and they pass that absorbed energy to chi a (primary photosynthetic pigment). They do not convert absorbed energy into chemical energy. The transfer of absorbed energy to chi a is called resonance transfer. Chlorophyll b and carotenoids are accessory pigments.

                                    (3) Chlorophyll: It is an accessory pigment and its formula is C55H70O6N4Mg. It differs from chi a in having an aldehyde group (CHO). It absorbs red light of 644nm and blue light of 455nm. It transfers absorbed light energy to chla.

                                    (4) Carotenoids:
                                    They are accessory pigments, they are of two types – carotenes (orange) and Xanthophylls (yellow) carotenes are insoluble in water and soluble in organic solvents. Carotenoids are always found associated with chlorophylls and in thylakoids, they are present as chromoproteins. Carotenes absorb blue and green lights and transmit red and yellow lights. Carotenoids absorb light of visible spectrum between 450 – 500nm. The formula of carotene is C40H56 (Carotene is a hydrocarbon compound). The formula of Xanthophyll is C40H56O2N4

                                    Question 8.
                                    Describe Mohl’s half leaf experiment. (April 2006)
                                    Answer:
                                    Aim: To show that CO2 is necessary for photosynthesis.
                                    Procedure: A potted plant is placed in the dark for 2 days to research it completely. To one of its leaf, a wide-mouthed bottle with a split cork is fixed in such a way that half the leaf is inside the bottle and the other half outside the bottle. The bottle contains KOH solution. The set up is placed under sunlight for a few hours, the leaf detached and tested for starch.
                                    Result: The leaf shows a lower portion dark blue in colour and the upper part light in colour.

                                    Inference: CO2 is one of the raw materials of photosynthesis. It is needed to produce carbohydrates. The portion of the leaf inside the bottle does not receive CO2 hence gives a negative test for starch, but the lower portion which is outside the bottle receives CO2 hence gives a positive test. The equation of photosynthesis is

                                    Question 9.
                                    Explain the fixation of carbon dioxide during the C3 pathway.
                                    Answer:
                                    The photosynthetic reaction which does not require light (light-independent) is called a dark reaction. A dark reaction occurs in the stroma portion of chloroplast so it is called a stroma reaction. It is also called Blackman’s reaction. Dark reaction is influenced by temperature. The different chemical reactions of dark reactions are enzymatic. Enzymes of stroma catalyze these reactions. Enzymatic reactions of dark reactions occur in the form of a Cycle and they were studied by Melvin Calvin, Benson, and Bassham in 1949.

                                    So dark reaction is also called Calvin cycle or Calvin Benson cycle. They used the tracer technique by using C 14 O2 in unicellular alga Chlorella and found a path of CO2 in each step of a chemical reaction. The ATP and NADPH of the light reaction are used for the fixation of CO2 during the dark reaction. The main event of dark reaction is the fixation of CO2. So dark reaction is also called the CO2 fixation cycle.

                                    The end product of the Calvin cycle or dark reaction is hexose sugar (glucose).
                                    CO2 acceptor compound: CO2 diffuses into the mesophyll tissue through stomata and then into the stroma of the chloroplast. CO2 is first accepted by a 5-carbon compound called Ribulose 1, 5 biphosphates (RUBP). So RUBP is a CO2 acceptor compound during dark reaction. RUBP is first available in the form of RUMP (Ribulose monophosphate).

                                    First stable intermediate compound of dark reaction: During the dark reaction, the first stable intermediate compound formed is phosphoric- eric acid and it is 3 – carbon compound. So dark reaction studied by Calvin (Calvin cycle) is called C3 cycle. The plants which produce 3 carbon compound (phosphoglyceric acid – PGA) as their first stable product are called C3 plants.

                                    The various steps of the C3 cycle are:
                                    (1) Phosphorylation: 6 molecules of RUMP (Ribulose monophosphate) reacts with 6 molecules of ATP and produce 6 molecules of RUBP (Ribulose 1,5 biphosphate) which is a CO2 acceptor compound.

                                    (2) Carboxylatlon: Six molecules of CO2 react with 6 molecules of RUBP in the presence of an enzyme RUBP carboxylase oxygenase or “Rubisco”. RUBP is converted into 12 molecules unstable 6 carbon compounds.

                                    (3) Cleavage (Splitting): 12 mole of 6 carbon unstable compounds splits into 12 molecules of 3 carbon compounds called phosphoglyceric acid (PGA). Since PGA (Phosphoglyceric acid) is a 3 carbon compound and it is the first stable intermediate product of the Calvin cycle. So it is the C3 cycle.

                                    (4) Phosphorylation: 12 molecules of PGA are phosphorylated in presence of 12 ATP molecules to 12 molecules of 1,3 ‘ diphosphogly ceric acid (12diPGA)

                                    (5) Reduction: 12 molecules of 1,3 diphosphoglyceric acid is reduced to form 12 molecules of 3 – phosphoglyceraldehyde (PG AL) by 12 molecules of NADPH, produced in the light reaction.
                                    12 mols, 1,3-di PGA+ 12 NADPH

                                    (6) Utilization of PGALD for the synthesis of sugar and the regeneration of RUMP.
                                    (A) Out of 12 PGAL, 5 PGAL are isomerized to 5 molecules of dihydroxyacetone phosphate (DHAP).

                                    (B) 3 PGAL undergo condensation with 3 DHAP to form 3 molecules of 6 carbon compound 3 Fructose 1,6 diphosphate (3 F.1,6 diphosphate)

                                    Question 2.
                                    Explain the structure of ATP and write three types of ATP synthesis.
                                    Answer:

                                    Adenosine triphosphate has three components
                                    (a) Adenine (a nitrogen base)
                                    (b) ribose sugar c) three inorganic phosphate.
                                    Adenine + ribose → Adenosine
                                    Adenosine + 3 Pi → Adenosine + triphosphate
                                    ATP can be written as A – (P)

                                    (P).
                                    The first phosphate which is linked by an ester linkage to ribose. The two-terminal bonds are energy-rich and the first bond is energy poor. The energy-rich phosphate bonds are represented by a curly line (

                                    ) and the energy-poor bond by a straight line (-).
                                    ATP is synthesized by the addition of inorganic phosphate to ADP. This process is called phosphorylation.
                                    n ADP + n Pi → n ATP.
                                    There are three different types of phosphorylations.


                                    NEET UG BIOLOGY NOTES: PHOTOSYNTHESIS

                                     Photosynthesis is simply defined as “ formation of carbohydrates from CO2 and H2O by illuminated green cells of plants, O2 and H2O by being the byproducts”. In other words, capture of photons of light by green plant cells and conversion of their radiant energy into chemical form of energy is called photosynthesis.

                                    IMPORTANCE OF PHOTOSYNTHESIS

                                    (i) Synthesis of organic food.

                                    (ii) Non-photosynthetic or heterotrophic organisms depend upon for organic food. Plants are, therefore called produces. Other are called consumers.

                                    (iii) It converts radiant or solar energy into chemical energy.

                                    (iv) Fossil fuels are products of photosynthetic activity of past plants.

                                    (v) All plant products of photosynthetic activity of past plants.

                                    (vi) It absorbs CO2 from atmosphere which tends to increase due to respiration of organisms and combustion.

                                    (vii) It evolves oxygen which is consumed in respiration and combustion of respiratory substrate and formation of ozone in stratosphere for filtering out harmful radiations.

                                    (viii) Productivity of crop depends upon rate of photosynthesis.

                                    LANDMARKS IN PHOTOSYNTHESIS

                                    1. STEPHAN HALES (1727) : Father of plant physiology pointed out that green plants require sunlight to obtain nutrition from air.

                                    2. JOSEPH PRIESTLY (1771): An English clergyman and chemist, showed that the plants purify air which becomes foul by the burning of candles and respiration by mice.

                                    3. INGENHOUSZ: A Dutch physician in 1779 demonstrated that light is necessary for purification of air by plants.

                                    4. JEAN SENEBIER (1782): He showed that the presence of noxious gas produced by animals and by plants in darkness (CO2) stimulated production of “purified air” (O2) in light.

                                    5. NICHOLAS THEODORE de SUSSURE (1804): He showed that the total weight of the organic matter produced and oxygen evolved by the green plants in presence of sunlight was greater than the weight of fixed air (CO2), consumed by them during this process. He concluded that besides fixed air ( CO2), water must constitute the raw material for this process.

                                    6. PALLETIER AND CAVENTION (1818): They discovered and named green colour of leaf as chlorophyll which could be separated from leaf by boiling in alcohol.

                                    7. JULIUS ROBERT MAYER (1845): He observed that the green plants utilize light energy and convert it into chemical energy of organic matter.

                                    8. JULIUS VON SACHS (1854): Showed that the process of photosynthesis takes place in chloroplasts and results in the synthesis of starch. He also showed that chlorophyll is confined to chloroplast.

                                    9. GG STOCKS (1864): Obtained pure fraction of chlorophyll –a and b and detected the presence of chlorophyll – c.

                                    10.ENGELMANN (1888): Plotted the action spectrum of photosynthesis.

                                    11. FF BLACKMAN (1905): Noted that photosynthesis is a two step process. A dark reaction also occurs along with photochemical reaction. He also proposed the law of limiting factor.

                                    12.WILLSTATTER AND STOLL (1913, 1918): Showed detailed account of chemical composition and functioning of chlorophyll.

                                    13.WARBERG (1920): Flash light experiment with chlorella as useful material for photosynthesis experiments.

                                    14.VAN NEIL (1931): Showed that the photosynthetic bacterial fixed CO2 in the presence of H2S. He postulated that the plants evolve O2 by splitting H2O not CO2.

                                    15.EMERSON AND ARNOLD (1932): Recognised light reaction consists of two distinct photochemical process. They showed that about 2500 chlorophyll molecules are required to fix one molecule of CO2 in photosynthesis.

                                    16.ROBIN HILL (1937): Isolated chloroplast suspended in water in presence of suitable hydrogen acceptor which evolve oxygen in presence of light. He demonstrated that the source of O2 evolved during photosynthesis is water and not CO2.

                                    17.RUBEN AND KAMEN(1941): Used radioactive oxygen O18 and proved that oxygen evolved was part of water.

                                    18.ARNON, ALLEN AND WHATLEY(1954): Demonstrated that fixation of CO2 by chloroplast using C14O2.

                                    19.MELVIN CALVIN(1954): Traced the path of carbon in photosynthesis using unicellular algae chorella. Melvin calvin gave C3-cycle and was awarded Nobel Prize in 1960 for the discovery

                                    20.PARK AND BIGGINS(1961): Discovered quantosome 100 Angstrom thick and stated that it contains about 230 chlorophyll molecules.

                                    21.HATCH AND SLACK(1967): Discovered C4 pathway for fixation of CO2 22.HUBER, MICHEL AND DISSENHOFER (1985): Crystallised photosynthesis reaction centre of bacterium Rhodobacter and got Nobel Prize in 1988.

                                    RAW MATERIALS FOR PHOTOSYNTHESIS

                                     In green plants including algae, photosynthesis takes place in chloroplasts of the cells. During this process, solar energy is trapped and synthesis of carbohydrates takes place from carbon dioxide and water. This sunlight, carbon dioxide, water, chloroplast are important components necessary for plants to derive the process of photosynthesis.

                                    SUNLIGHT

                                     Photosynthesis is a light dependent process. The literal meaning of world “Photosynthesis” is “ the synthesis, with the help of light”. To drive photosynthesis in plants, sunlight provides solar energy. Only 0.2% of the light energy, incident on earth is actually used by photo autotrophs.

                                     Light is the visible radiation which represents a very small portion of the total electromagnetic spectrum of radiation, emitted by the sun. Visible light (approx. between 400nm to 700 nm) causes the physiological sensation of vision of man. Visible light is actually a combination of several colours of different colours viz. Violet (400nm to 425 nm), blue (425nm to 490nm), green (490 – 550 nm), yellow (550 -585 nm), organge (585 – 640 nm) and red (640 – 700 nm)

                                     The most effective regions of visible light spectrum responsible for maximum photosynthesis in plants are blue and red regions of which red light is most effective. On the other hand, green light is least effective. Photosynthesis cannot take place beyond the range of visible spectrum.

                                    CARBON DIOXIDE

                                     In land plants, carbon dioxide is obtained from the atmosphere through the stomata. Small quantities of carbonates are also absorbed from soil through the roots. Hydrophytes get their carbon dioxide supply from the aquatic environment as bicarbonates. The latter are absorbed by hydrophytes through their general surface.

                                    WATER

                                     In the process of photosynthesis, the source of liberated oxygen in water. Photosynthetic land plants absorb a large amount of water from the soil through the root hairs-present on their roots. But relatively very small amount of this absorbed water is used in the process of photosynthesis. Aquatic photosynthesis plants absorb water through their body surface.

                                     As mentioned earlier, Van Niel (1931) hypothesized that the pototosynthetic organisms require a source of hydrogen. He proposed that oxygenic photosynthesis is an oxidation reduction reaction where hydrogen of water reacts with carbon dioxide to form organic compounds

                                     1937, Robin Hill demonstrated that in absence of carbon dioxide, isolated chloroplasts of stellaria media produced oxygen when they were illuminated in presence of hydrogen acceptor. Here ferricyanide is reduced to ferrocyanide by photolysis of water. This is Hill reaction and can be represented as

                                     The hydrogen acceptor is often called as Hill oxidant or Hill reagent. In plants, NADP+ ( Nicotinamide adenine dinucleotide phosphate) acts as a hydrogen acceptor.

                                     In 1941, by using non-radioactive heavy isotope of oxygen (O18), Ruben and Kamen proved that during photosynthesis, oxygen comes from the water.

                                    CHLOROPLASTS

                                     Chloroplasts ( Chloros = green, plastos = moulded) are the green plastids which occur in all the green parts of the plants.

                                     They are the actual sites of photosynthesis.

                                     The chloroplasts contain chlorophyll and carotenoid pigments which are responsible for trapping light energy essential for photosynthesis.

                                     Majority of the chloroplasts of the green plants are formed in the mesophyll cells of the leaves.

                                     They are lens shaped, oval, spherical, discoid or even ribbon like organelles having variable length ( 5- 10 mm) and width (2 -4 mm).

                                     The chloroplasts are double membrane bound, each membrane are 9-10 mm in thickness. The space limited by the inner membrane of the chloroplast is called the stroma. It is the site of dark reaction.

                                     A number of organized flattened membranous sac called the thylakoids are arranged in stacks like piles of coins called grana. Thylakoids lying outside the grana are called stroma, thylakoids or the intergrana thylakoids

                                     Each granum may contain 20 to 50 thylakoid discs. There may be 40 – 60 grana per chloroplasts.

                                     The major function of thylakoids is to perform photosynthetic light reaction ( photochemical reaction)

                                     The pigments and other factors of light reaction are usually locataed in thylakoid membranes.

                                     Cyanobacteria and other photosynthetic bacteria do not possess chloroplasts. However, the photosynthetic pigments which lie freely in the cytoplasm. There photosynthetic pigments are also different from those of eukaryotes.

                                     Thylakoids possess four types of major complexes photosystem I, photosystem II, cy b6 – f comples and coupling factor (ATP synthetase)

                                     Photosystem II is thought to mostly occur in the appressed or partition to mostly occur in the appressed or partition regions of granal thylakoids while photosystem I lies in the nonappressed parts as well as stroma thylakoids.

                                    PHOTOSYNTHETIC PIGMENTS

                                    (i) Chlorophylls It is a green pigment which traps solar radiation and convert light energy to the chemical energy. Generally, it is of two types.

                                    (a) Chlorophyll –a (C55H72O5N4Mg): It participates directly in the light reactions of photosynthesis has a head called a porphyrin ring with a magnesium atom at its centre. Attached to the porphyrin is a hydrocarbon tail, which interacts with hydrophobic regions of proteins in the thylakoid membrane.

                                    (b) Chlorophyll-b (C55H70O6N4Mg): It differs from chlorophyll-a only in one of the functional group bonded to porphyrin. This diagram simplifies by placing chlorophyll at the surface of the membrane most of the molecules are actually immersed in the hydrophobic core of the membrane.

                                    (ii) Carotenoids These are yellow, brown and orange pigments, which absorb light strongly in blue-violet range. These are called shield pigments, because they protect chlorophyll from photo ocidation by light intensity and also from oxygen produced during photosynthesis. Along with chlorophyll-b, the cartenoids are also called as accessory pigments, because they absorb energy and give it to chlorophyll-a. carotenoids are two types:

                                    (a) Carotenes: Carotenes consists of an open chain conjugated double bond system ending on both the sides with ionone rings. They are hydrocarbons with molecular formula C40H56 carotenes are orange in colour. The red colour of tomato and chillies is, because of carotene call lycopene. The common carotene is β-carotene which is converted to vitamine-A by animals and humans

                                    (b) Xanthophylls: Also known as carotenols. These are similar to carbon, but differ in having two oxygen atoms is the form of hydroxyl, carboxyl group attached to the ionone rings. Their molecular formula is C40H56O2. The yellow colour of autumn leaves is due to lutein and a characteristics xanthophylls of brown algae is fucoxanthin

                                    (iii) Phycobilins Phycobilins consist of four pyrrol rings and lack Mg and phytol tail. The phycobilin pigments are of two types.

                                    (a) Blue – Phycocyanin, allophycocyanin

                                    (b) Red – phycoerythein These pigments are useful in chromatic adaptations. Phycoerytherin transfer energy to phycocyanin which in turn transfer energy to carotenoids which is ultimately received by chlorophyll –a.

                                     The chlorophylls, carotenoids and phycobilins together form a complex of pigment in thylakoid membrane. These complexes work for the absorption of light and its transfer to a reaction center. These complexes are called photosynthetic unit or photosystem or pigment system. These system show clear division of labour. Some pigments called as accessory pigments such as carotenoids act to receive the light. They basically harvest the light molecules towards a reaction center thus, also called as Light Harvesting complexes (LHC). Chlorophyll-a act as reaction center and perform further reaction of photosynthesis.

                                    ABSORPTION SPECTRUM AND ACTION SPECTRUM

                                     The graphic representation of curve depicting the various wavelength of light absorbed by a substance is known as absorption spectrum. Chlorophyll mostly absorb light radiations in blue (more) and red parts of light spectrum ( 430 to 662 nm for chlorophyll a, 455 and 604 nm for chlorophyll b)

                                     Action Spectrum: It is a graphical representation of curve depicting the rate of photosynthesis in various wavelengths of light.

                                     Fluorescence: It is property of almost immediate emission of long wave radiation by substances after attaining excited state on receipt of light energy e.g. Chlorophyll

                                     Phosphorescence: the delayed emission of long-wave radiations from an activated molecule is called phosphorescence. It continues for some time after removal of irradiation source.

                                    PHOTOSYNTHETIC UNIT

                                     It is the smallest group of photosynthetic pigment molecules which can pick up light energy and convert it into chemical form. A photosynthetic unit has 250-400 pigment molecules. It has a photocentre of chlorophyll a molecules surrounded by harvesting molecules differentiated into core molecules and antenna molecules

                                     Antenna molecules are meant for absorbing radiation energy of different wavelengths. On absorbing a photon of light, the pigment molecule enters excited state. In this state the electrons move into outer orbital. The excited state lasts for 10-9 seconds. In this period the excited antenna pigment molecule transfer its energy to a core molecule through resonance. If this does not happen, the energy is lost as fluorescence. The core molecules pass over their energy to trap centre or photocentre. The frequency of excitation is very high. It is met by collaboration of core and antenna molecules. Each time the trap centre or photocentre gets excited, it expels an electron and becomes oxidized. An electron is required to convert it to normal state.

                                    PHOTOSYSTEM I (PS I)

                                     It is a photosynthetic pigment system along with some electron carriers that is located on both the nonappressed part of grana thylakoids as well as stroma thylakoids.

                                     PS-I has more of chlorophyll a

                                     Chlorophyll b and carotenoids are comparatively less.

                                     Photosystem I has a reducing agent X which is special chlorophyll P700 molecule, FeS centre B or ferredoxin, plastoquinone, cytochrome complex and plastocyanin.

                                     It takes part in both cyclic and non-cyclic photophosphorylation.

                                     PS-I can carry on cyclic phosphorylation independently.  Normally it drives an electron from photosystem II to NADP+

                                    PHOTOSYSTEM II (PS II)

                                     It is a photosynthetic pigment system alongwith some electron carriers that is located in the appressed part of grana thylakoids.

                                     PS –II has chlorophyll a,b and carotenoids.

                                     Chl a and Chl b contents are equal.

                                     Carotenoid content is higher as compared to that of PS I

                                     The photocentre is a special chlorophyll a molecule called P680  It is surrounded by other chlorophyll a molecules, chlorophyll b and carotenoid molecules

                                     PS II also contains Mn2+, Cl- , quencher molecules Q, plastoquinon (PQ), cytochrome complex and plastocyanin.  It picks up electron released during photolysis of water.

                                     The same is extruded on absorption of light energy.

                                     As the extruded electron passes over cytochrome complex, sufficient energy is released to take part in the synthesis of ATP from ADP and inorganic phosphate.

                                     This photophosphorylation is non-cyclic.

                                     PS II can operate only in conjugation with PS I

                                    NON – CYCLIC PHOTOPHOSPHORYLATION

                                     It is the normal process of photophosphorylation in which the electron expelled by the excited photocentre does not return to it.

                                     Non-cyclic photophosphorylation is carried out in collaboration of both photosystem I and II.

                                     Electron released during photolysis of water is picked up by photocentre of PS II called P680.

                                     The same is extruded out when the photocetre absorbs light energy.

                                     The extruded electron has an energy equivalent to 23 kCl / mole

                                     It passes through a series of electron carriers phaeophytin, PQ. Cytochrome b6 – f complex and plastocyanin.

                                     While passing over cytochrome complex, the electron loses sufficient energy for the synthesis of ATP.

                                     The electron is handed over to photocentre P700 of PS I by plastocyanin. P700 extrudes the electron after absorbing light energy. The extruded electron passes through special chlorophyll P680 molecules, Fe-S, ferrodix, to finally reach NADP+

                                     The latter then combines with H+ with the help of NADP – reductase to form NADPH.

                                     This is called Z scheme due to its characteristics zig-zag shaped based on redox potential of different electron carriers.

                                     Non-cyclic photophosphorylation or Z-scheme is inhibited by CMU and DCMU.

                                     DCMU ( Dichlorophenyldimethyl urea) is a herbicide which kills the weed by inhibiting CO2 fixation as it is strong inhibitor of PS II

                                    CYCLIC PHOTOPHOSHORYLATION

                                     It is a process of photophosphorylation in which an electron expelled by the excited photocentre is returned to it after passing through a series of electron carriers.

                                     It occurs under conditions of low light intensity, wavelength longer than 680nm and when CO2 fixation is inhibited.

                                     Absence of CO2 fixation results in non-requirement of electrons for formation of NADPH  Cyclic photophosphorylation is performed by photosystem I only.

                                     Its photocentre P700 extrudes an electron with gain of 23 kcal/mol of energy after absorbing a photon of light.  After losing the electron the photocentre becomes oxidized.

                                     The expelled electron passes through a series of carriers including P700 chlorophyll molecules, plastoquinone (PQ), FeS complex, ferrodix (Fd) cyt b6 – f and plastocyanin before returning to photocentre.

                                     Over the cytochrome complex (cyt b6-f), the electron creates a proton gradient for synthesis of ATP from ADP and inorganic phosphate.

                                     Halobacteria or halophile bacteria also perform photophosphorylation but ATP thus produced is not used in synthesis of food. These bacteria possess purple pigment bacteriorhodopsin attached to plasmamembranes. As light falls on the pigment, it creates a proton pump which is used in ATP synthesis.

                                     Cyclic photophosphorylation is the most effective anaerobic phosphorylation mechanism.

                                    CHEMIOSMOTIC HYPOTHESIS OF ATP FORMATION

                                     The view was propounded by Peter Mitchell in U.K. in 1961 in the case of mitochondria and chloroplast.

                                     Mitchell’s chemiosmotic theory was confirmed by G.Hind and Andre jagendorf at cornell university in 1963.

                                     According to this view, electron transport, both in respiration and photosynthesis produces a proton gradient (pH gradient)

                                     The gradient develops in the outer chamber or inter-membrane space of mitochondria and inside the thylakoid lumen in chloroplasts.

                                     Lumens of thylakoid becomes enriched with H+ ion due to photolytic splitting of water.

                                     Primary acceptor of electron is located on the outer side of thylakoid membrane.

                                     It transfer its electrons to a H-carrier. The carrier removes a proton from matrix while transporting electron to the inner side of membrane.

                                     The proton is released into the lumen while the electron passes to the next carrier.

                                     NADP reductase is situated on the outside of thylakoid membrane.

                                     It obtains electron from PS I and protons from matrix to reduce NADP+ to NADP + H+ state.

                                     The consequences of the three events is that concentration of proton decreases in matrix or stroma region while their concentration in thylakoid lumen rises resulting in decrease in pH.

                                     A proton gradient develops across the thylakoid.

                                     The proton gradient is broken down due to movement of protons through transmembrane channels, cFo of ATPase (cFo – F1 particle).

                                     The rest of the membrane is impermeable to H+ , cF0 provides facilitated diffusion of H+ or protons.

                                     As the protons move to the other side of ATP, they bring about conformational changes in cF1 particle of ATPase or coupling factor.

                                     The transient cF1 particles of ATPase enzyme from ATP from ADP and inorganic phosphate.

                                     Therefore, ATP synthesis through chemiosmosis requires a membrane, a proton pump, a proton gradient and cF0 – cF1 particle or ATP-ase

                                     One molecule of ATP is formed when 3H+ used by the ATP synthase.

                                    LIGHT REACTION ( Photochemical phase).

                                     It occurs inside the thylakoids, especially those of grana regions.

                                     Photochemical step is dependent upon light. The function of this phase is to produce assimilatory power consisting of reduced co-enzyme NADPH and energy rich ATP molecules.

                                     Photochemical phase involves photolysis of water and production of assimilatory power.

                                     The phenomenon of breaking up of water into hydrogen and oxygen in the illuminated chloroplast is called photolysis of water.

                                     Light energy, an oxygen evolving complex (OEC) and an electron carrier are required.

                                     Oxygen evolving complex was formly called Z-enzyme  It is attached to the inner surface of thylakoid membrane.

                                     The enzyme has four Mn ions. Light energized changes in Mn ( Mn2+, Mn3+, Mn4+) removes electrons from OHcomponent of water forming oxygen.

                                     Liberation of O2 requires two other ions Ca2+ and Cl- .

                                     Electron carrier transfer the released electrons to P680 4H2O ⇌ 4H+ + 4OH-

                                     The electron released during photolysis of water are picked up by P680 photocentre of photolystem II.

                                     On receiving a photon of light energy the photo-centre expels an electron with a gain of energy ( 23 kcal/mole).

                                     It is the primary reaction of photosynthesis which involves the conversion of light energy into chemical form.

                                     The phenomenon is also known as quantum conversion.

                                     The electron extruded by the photocentre of photosystem II is picked up by the quencher phaeophytin.

                                     From here the electron passes over a series of carriers in a downhill journey losing its energy at every step.

                                     The major carriers are plastoquinone (PQ) cytochrome b-f complex and plastocyanine (PC).

                                     While passing over cytochrome complex, the electron loses sufficient energy for the creation of proton gradient and synthesis of ATP from ADP and inorganic phosphate by the process of photophosphorylation.

                                     From plastocyanin the electron is picked up by the trap centre P700 of photosystem I.

                                     On absorbing a photon of light energy, P700 pushes out the electron with a gain of energy.

                                     The electron passes over carriers, FeS, feredoxine and NADP-reductase.

                                     The latter gives electron to NADP+ for combining with H+ ions to produce NADPH.

                                     NADPH is a strong reducing agent. It constitutes the reducing power which is also contains a large amount of chemical energy.

                                    DARK REACTION ( Biosynthetic phase)

                                     Dark reaction of phorosynthesis occurs in presence of or absence of light i.e. independent of light.

                                     Dark reaction occurs in stroma fraction of the chloroplast.

                                     Dark reaction is purely enzymatic reaction and is slower than light reaction of photosynthesis.

                                     Dark reaction was first of all established in detail by Dr. Calvin, Benson and J.Bassham and for this work they were given Nobel prize (1961).

                                     The techniques used for studying different steps were radioactive tracer technique using 14C chromatography and autoradiography and the material used were chlorella and scenedesmus. These are microscopic, unicellular algae and can be easily maintained in laboratory.

                                     Dark reaction is also named as Blackman’s reaction.

                                    C3- PATHWAY OR CALVIN CYCLE

                                     The details of the step involved in the dark reaction were discovered by Professor M. Calvin and hence the dark reaction known to be called as Calvin cycle.

                                     This is the major pathway for the fixation of carbon dioxide in green plants. It represents phase II i.e. dark reaction. It takes place in the stroma of the chloroplasts.

                                     The reactions are enzyme. Controlled and temperature dependent. After the fixation of carbon dioxide, the first stable compound formed is 3-carbon phosphoglyceric acid ( PGA). Hence, it is also called the C3 – pathway.

                                     Calvin cycle can described under three stages:

                                    (a) Carboxylation of RUBP:

                                    – In this process there is fixation of atmospheric CO2 into a stable organic compound with the help of enzyme RuBP, Carboxylase-oxygenase or RuBisCO

                                    (b) Reduction of CO2:

                                    – The 3-C PGA then undergoes reduction with the help of the assimilatory power to form 3-c phosphoglycerladehyde (PGAL). NADPH2 provides the hydrogen and ATP supplies energy for the reduction. Enzyme trisephosphate dehydrogenase catalyses the reaction.

                                    – Some molecules of PGAL are converted into another triosephosphate called Dihydroxy Acetone Phosphate (DHAP) in presence of enzyme phosphor triose isomerase.

                                    – The formation of sugars ( end products of photosynthesis), the 3-C triose phosphates ( PGAL 3-C and PHAP 3-C) to form 6-C hexose sugar fructose 1,6-biphosphate in the presence of enzyme aldolase.

                                    – Fructose biphosphate is the diphosphorylated first to fructose monophosphate and then to fructose ( 6-C) in the presence of enzyme phosphotase. Some fructose monophosphate molecules may be isomerised into glucose monophosphate by the enzyme isomerase and then into glucose ( 6-C). The hexose sugar may be further converted to sucrose (C12H22O11) or to starch (C6H10O5)n and are stored in storage cells.

                                    (c) Regeneration of RuBP

                                    – The 5-C RuBP is constantly required for the fixation of CO2 in the calvin cycle. It is regenerated through another chain of reactions.

                                    – Some molecules of triosephosphate and fructose monophosphates are used from the calvin cycle for the formation of RuBP to be used again to combine with CO2

                                    – The net reaction of calvin cycle can be represented by 6RuBP + 6CO2 +18ATP +12NADPH  6RuBP + C6H12O6 + 18ADP +12 NADPH+ + 18 Pi

                                    Balnce sheet of calvin cycle

                                    C4 – PATHWAY OR HATCH AND SLACK PATHWAY

                                     In some plants, the first stable product, after the fixation of CO2, is 4-C dicarboxylic acid called oxaloacetic acid (OAA), such plants are called C4 plants and path of carbon ) dark reaction) is called C4 – pathway.

                                     It was first noticed by Kortschak (1964) in the photosynthesis of sugarcane leaves. However details of the C4 – pathway, were worked out by Hatch and Slack ( 1966). Therefore, it is called Hatch and slack pathway.

                                    ANATOMICAL PECULIARITIES OF C4 – PLANTS

                                    (a) The leaf mesophyll consists of compactly arranged cells.

                                    (b) It is not differentiated into palisade and spongy mesophyll as in C3 plants

                                    (c) The vascular bundles (veins) in the leaves are surrounded by a distinct bundle sheath of radially enlarged parenchyma cells.

                                    (d) The chloroplast in leaf cells are dimorphic i.e. granal and agranal chloroplast

                                    – Chloroplasts in mesophyll cells are smaller and possess grana.

                                    – Chloroplasts in the bundle sheath cells are larger and without grana. This type of leaf anatomy in C4-plants is called as Kranz anatomy

                                    IMPORTANT STEPS IN C4 – PATHWAY

                                    (a) First part reactions are completed in the stroma of the chloroplasts in mesophyll cells.

                                    (b) Second part, reactions are completed in the stroma of the chloroplasts in bundle sheath cells.

                                    Part I ( in mesophyll cells)

                                    – First CO2 fixation: In this pathway, the first CO2 acceptor is 3-C phosphoenol Pyruvate (PEP), CO2 first combines with 3-C PEP to form 4-C OAA ( oxaloacetic acid). As DAA is a dicarboxylic acid pathway.

                                    – 4-C OAA is converted into 4-C malic acid or 4-C aspartic acid and transported to bundle sheath cells.

                                    Part II ( in bundle sheath cells)

                                    – In the chloroplasts of bundle sheath cells, 4-C malic acid undergoes decarboyylation to form CO2 and 3-C pyruvic acid.

                                    – Second CO2 fixation: The CO2 released in decarboxylation of malic acid combines with 5-C RuBP ( Ribulose 1,5-biphosphate) to form 2 molecules of 3-C PGA. Further, the conversion of PGA to sugar is the same as in the calvin cycle.

                                    – The pyruvic acid produced in decarboxylation of malic acid is transported back to the mesophyll cells. Here, it is converted to phosphoenol pyruvic acid (PEPA) and again made available for the C4-pathway.

                                     In C4 pathway when carbon dioxide fixation take place, an additional 2 molecules of ATP per molecule of CO2 fixed are also required to convert pyruvic acid to phosphoenol pyruvic acid. Thus in C4 cycle in all 30ATPs are required for fixing 6 molecules of carbon dioxide.

                                    CAM ( Crassulacean Acid Metabolism) PATHWAY

                                     In the member of crassulaceae, cactaceae, agavaceal, orchidaceae, CO2 fixation occurs during night only.

                                     In succulents belonging to the above families the stomata remain closed during day time in order to reduce transpiration and the stomata open during night.

                                     In CAM plants OAA is formed due to carboxylation as in C4 plants.

                                     Like C4 plants, OAA is reduced to make malic acid in CAM plants and is accumulated in the vacuole.

                                     Absorption of CO2 during night and its storage as organic acid (malic acid) is called acidification.

                                     During day time malic acid undergoes oxidative decarboxylation nad CO2 is released.

                                     Liberation of CO2 from an organic acid during day time is called deacidification.

                                     The diurnal acidification and deacidification during the night and day time respectively is called CAM

                                     In C4 plants, initial carboxylation and final carboxylation is separated by space but in CAM plants, they are separated by time.

                                     All reactions of CAM occurs in mesophyll cells.

                                     Chloroplasts are absent in bundle sheath cells of CAM plants

                                     CAM pathway is important for the survival of succulents

                                    NUMBER OF ATP AND NADPH REQUIRED FOR 1CO2 FIXATION

                                    PHOTORESPIRATION

                                     It was first observed by Otto Warburg (1920) that presence of high O2 concnetration and high temperature decreases the rate of photosynthesis. Later it was demonstrated by Dicker and Tijo (1959) in tobacco.

                                     RuBisCO is most abundant enzyme and it has affinity to both CO2 and O2. In C3 – plants, when there is higher O2 concentration and temperature, O2 binds with RuBisCO instead of CO2 and form one molecule of phosphoglycerate and phosphoglycolate in pathway called photorespiration, so there is neither synthesis of sugars, nor of ATP. Instead it results in the release of CO2 with the utilization of ATP. In photorespiratory pathway there is no synthesis of ATP and NADPH.

                                     The process can be understood in the following steps.

                                    1. Oxygen binds with RuBP oxygenase to form phosphoglycolate in chloroplast which gets converted to glycolate and transported to peroxisomes.

                                    2. In peroxisome it forms glyoxylate and then glycine.

                                    3. Glycine then enters mitochondria and looses NH4 and CO2 in a reaction and it form serine.

                                    4. Serine is transported to perioxisomes and in a series of reaction it form glycerate which gets converted to PGA and then RuBP is the chloroplast.

                                    5. So, here we can see, there is no fixing of CO2 instead CO2 is given off along with NH4. Thus it reduces the rate of photostnthesis in C3 plants

                                    PRINCIPLE OF LIMITING FACTORS ( Blackman, 1905 )

                                     When a process is conditioned as to its rapidity by number of separate factors, the rate of process is limited by the pace of the slowest factor. In other words, at one time only one factor limits the rate of the process. It is called limiting factor. A limiting factor is that factor which is deficient to such a extent that increase in its value directly increases the rate of the process.

                                    FACTORS AFFECTING PHOTOSYNTHESIS

                                     The light reaction totally depends on the availability of light, water, pigments etc and the dark reaction depends on the temperature and available

                                    CO2 EXTERNAL FACTORS – Light: In photosynthesis light is converted to chemical energy in the food formed.

                                    (i) Light intensity – Light intensity required to get the optimum value differs with different species. Usually with increase in light intensity increase in rate is noticed. The value of light saturation at which further increase in photosynthetic rate is not accompanied by an increase in CO2 uptake is called light saturation point.

                                    (ii) Light quality- Blue and red light of the spectrums is said to be the best for the photosynthesis. The maximum photosynthesis is shown to occur in the red part of the spectrum with the next peak in blue part. The green light has inhibitory effect.

                                    (iii) Light duration – Generally photosynthesis is independent of light duration. It is more in intermittent light than continuous light.

                                    – Carbon dioxide: Carbon dioxide is present in low concentration and form about 0.03% of total atmosphere CO2 is natural limiting factor of photosynthesis. It the concentration of CO2 is increased from 0.03% to 1%, the rate of photosynthesis increases, If concentration of CO2 exceeds 1% rate of photosynthesis decreases due to closer of stomata.

                                    – Water: Water deficiency may decrease the rate. Less availability of water may further check the rate by closing the stomata there by affecting the entry of CO2.

                                    – Temperature: The optimum temperature for photosynthesis is 15OC to 35OC. if the temperature is increased too high, the rate of photosynthesis is reduced due to denaturation of enzymes involved in the process. Photosynthesis occurs in conifers at high altitude at 35OC. Some algal in host springs can undergo photosynthesis even at 75OC. When other factors are not limiting rate of photosynthesis gets doubled for every 10OC rise in temperature untile an optimum is reached.

                                    – Oxygen: Excess of O2 may become inhibitory for the process. Enhanced supply of O2 increase the rate of respiration simultaneously decreasing the rate of photosynthesis. An increase in oxygen concentration decreases photosynthesis and the phenomenon is called Warbrug effect.

                                    – Mineral elements: Some mineral elements like Fe, Mg, Cu, Mn, Cl etc are associated with synthesis of chlorophyll and important reactions in photosynthesis like photolysis of water. So, absence of these elements decreases the rate of photosynthesis.

                                    INTERNAL FACTORS

                                    – Chlorophyll: Chlorophyll is an important internal factor for photosynthesis since it absorbs the radiant energy of light. Light initiates the mechanism of photosynthesis by transferring its electrons and getting excited. Emerson (1929) found direct relationship between the chlorophyll content and the rate of photosynthesis. The chlorophyll deficient mutants are albinos. They can’t synthesize carbohydrates by photosynthesis, so they cannot survives.

                                    – Leaf anatomy: Photosynthesis also depends upon the anatomy of leaf. If the assimilatory surface by palisade parenchyma is extensive there will be increased photosynthesis.

                                    – Leaf age: In immature leaf the rate of photosynthesis is at minimum level. A mature leaf shows phosynthetic rate at maximum. When leaf becomes old, the rate decreases.

                                    – End products: The end products of photosynthesis are carbohydrates. Accumulation of carbohydrates decreases the rate of photosynthesis. If the carbohydrates are translocated rapidly the rate of photosynthesis increases.

                                    – Protoplasmic factors: These factors include the hydration of protoplasm and also the enzymatic activity. If there is an appreciable decrease in the hydration of the protoplasm the process of photosynthesis is inhibited because the enzymes gets denatured.

                                    BACTERIAL PHOTOYNTHESIS

                                     It is an oxygenic ( without evolution of O2) because water is not employed as hydrogen donor. Instead H2 H2S and other compounds are employed. Trap centre is usually B890 of bacterio-chlorophyll a. It absorbs radiations between 870-890 nm of infra-red range. Though both cycles and non-cyclic photophosphorylations occur there is only one photosystem. Assimilatory power consists of ATP and NADH.

                                    CHEMOSYNTHESIS

                                     It is the manufacture of organic food from inorganic raw materials like carbon dioxide and a hydrogen donor with the help of energy obtained from exergonic reactions.

                                    Chemosynthesis is performed by certain bacteria. They are able to manufacture food in the absence of light.

                                     The organism carrying out chemosynthesis are called chemoautotrophs. Many of the chemoautotrops aare also able to obtain nourishment as saprotrophs and are thus actually facultative chemoautotrophs. They oxidize the inorganic substances present in their substrate. The energy is trapped and used in synthesis o organic compounds from inorganic raw materials. Chemoautotrops do not have a light trapping mechanism. They, however perform Calvin cycle reactions of carbon assimilation.

                                    Some common chemoautotrophs are nitrifying bacteria, sulphur bacteria, iron bacteria, methane bacteria, hydrogen bacteria and carboxy bacteria.

                                    TRANSLOCATION OF ORGANIC NUTRIENTS

                                     It is the movement of organic nutrients from the region of source or supply to the region of sink or utilisation. Phloem ( sieve tubes / sieve cells) is the pathway for this translocation as found out by

                                    (iii) Sieve tube puncturing

                                     Important theories about the mechanism of translocation of organic nutrients are:

                                    (a) Cytoplasmic / Protoplasmic Streaming Hypothesis Ina sieve tube element, organic solutes pass to all parts by cytoplasmic streaming while they pass from one element to another through diffusion.

                                    (b) Transcellular streaming hypothesis Sieve tubes possesses tubular transcellular strands which shows persistalis and hence take part in translocation of organic nutrients.

                                    (c) Mass flow hypothesis Organic region of high osmotic concentration to the region of low concentration in a mass flow due to occurrence of pressure gradient. It is most widely accepted theory.


                                    Results and discussion

                                    We used our merged model of photosynthesis and carbon fixation to perform a systematic supply–demand analysis of the coupled system. First, we have integrated the system for various constant light intensities until it reached steady state. Examples are provided in Fig. S1 in Appendix S1. We observed reasonable stationary values of intermediates and fluxes for most of the light intensities. However, under very low light intensities (below 5 μmol m −2 s −1 ), the phosphorylated CBB cycle intermediates dropped to zero, and ATP reached the maximal concentration equalling the total pool of adenosine phosphates. Depending on the initial conditions, either a non-functioning state, characterized by zero carbon fixation rate, or a functioning state, characterized by a positive stationary flux, was reached. This observation of bistability constituted the starting point of our analysis of the tight supply–demand relationship.

                                    In order to analyze this behavior in more detail, we performed time course simulations, in which the light was dynamically switched from constant sufficient light (between 20 and 200 μmol m −2 s −1 ), to a ‘dark phase’ of 200 s duration with a light intensity of 5 μmol m −2 s −1 , back to high light, and observed the dynamics of the model variables. In Fig. 2 we display the dynamics of the internal orthophosphate concentration, the sum of all three triose phosphate transporter (TPT) export rates and the RuBisCO rate (from top to bottom, respectively) during such light–dark–light simulations.

                                    In agreement with the steady-state simulations, higher light intensities result in a higher overall flux during the initial light phase. Higher carbon fixation and export fluxes are accompanied by lower orthophsophate concentrations, which reflect higher levels of CBB cycle intermediates. In the dark phase, the non-functional state with zero carbon flux is approached. While rates decrease, orthophosphate increases, reflecting a depletion of the CBB intermediate pools. In the second light phase, only the simulated transitions to light intensities of 150 and 200 μmol m −2 s −1 could recover a functional state under the chosen conditions. For lower light intensities, apparently the CBB intermediate pool was depleted to a level, at which re-illumination fails to recover the CBB cycle activity. Obviously, this behavior disagrees with everyday observations in nature (plant leaves recover from dark periods also under low light intensities). Nevertheless, the model is useful to generate novel insights. First, it illustrates that a critical threshold of intermediate concentrations exists. If levels drop below this threshold, the cycle cannot be re-activated. Second, it explains the mechanisms leading to intermediate depletion. Under low light conditions, insufficient energy supply results in reduced activity of ATP and NADPH dependent reactions in the carbon fixation cycle, leading to a reduced regeneration rate of ribulose 1,5-bisphosphate from ribulose-5-phosphate (Ru5P). Simultaneously, the reversible (ATP independent) reactions remain active. As triose phosphates are products of reversible reactions, these continue to be exchanged via the TPT export reactions with free phosphate, which leads to a depletion of the CBB cycle intermediates and a concomitant increase of the orthophosphate pool. This further illustrates that even deactivating key light-regulated CBB enzymes in the dark will not prevent the collapse of the cycle, because the continued activity of the reversible reactions and the triose phosphate translocator will still lead to depleted cycle intermediates (Fig. S2 in Appendix S1).

                                    Clearly, the model is missing important mechanisms that prevent such a functional failure. In particular, we are interested in how a stand-by mode can be realized, in which intermediate levels are maintained above the critical threshold, while at the same time the resources required to do so, are minimized. A possible strategy to prevent the collapse of the carbon fixation cycle is to resupply important intermediates. One biochemical process in plants that is known to produce Ru5P is the oxidative phase of the PPP, in which one glucose-6-phosphate molecule is oxidized and decarboxylated to Ru5P, while producing NADPH and CO2 (Kruger and Von Schaewen 2003 ). In order to estimate critical intermediate levels required to prevent the collapse of the carbon fixation cycle, we performed simulations under sufficient light (500 μmol m −2 s −1 ), with different initial conditions: the initial concentrations of all carbon fixation intermediates are set to zero, except for Ru5P. The simulated Ru5P concentration, depicted in Fig. 3, displays a characteristic dynamic. In the first seconds, the CBB cycle intermediates are equilibrated by the fast reversible reactions. If the equilibrated Ru5P concentration remains above the critical threshold of approximately 2.5 μM, the cycle reaches a functional state, if it falls below, it will collapse. Interestingly, the threshold concentration is rather independent of the light intensity (Fig. S3 in Appendix S1).

                                    To simulate a simple mechanism implementing a stand-by mode, which maintains sufficient CBB cycle intermediate levels, we introduced a trivial conceptual reaction, exchanging inorganic phosphate with Ru5P. Fig. 4 displays simulated steady state values of the relative stromal ATP concentrations, Ru5P concentrations and lumenal pH in insufficient light conditions (5 μmol m −2 s −1 ) as a function of the Ru5P influx. Again, a clear threshold behavior can be observed. If the Ru5P influx exceeds approximately 4 μM s −1 , not only CBB intermediates assume non-zero concentrations, but also the lumenal pH reaches realistic and non-lethal levels.

                                    As expected, increased Ru5P influx results in increased stationary Ru5P concentrations, which is accompanied by an increased flux through RuBisCO and the TPT exporter (Fig. S4 in Appendix S1), indicating a higher stand-by flux, and therefore, a higher requirement of resources to maintain this mode.

                                    These results suggest that a constant flux providing Ru5P in the dark with a rate just above the critical threshold of 4 μM s −1 should maintain intermediate CBB levels sufficiently high, while at the same time minimize the required investment. Indeed, with a constant supply of Ru5P with 5 μM s −1 , the system can be restarted and reaches a functional stationary state after a prolonged dark period (Fig. S5 in Appendix S1). Per carbon, this rate translates to 25–30 μM carbon/s, depending whether the pentoses are directly imported or derived from hexoses. Comparing this to stationary carbon fixation in the light of 0.1–1 mM s −1 (for light intensities between 20 and 200 μmol m −2 s −1 , Fig. 2 and Fig. S1 in Appendix S1) shows that resupply under these conditions would consume a considerable fraction of the previously fixed carbon. This calculation demonstrates the importance of down-regulating the CBB cycle in dark conditions for a positive carbon fixation balance over a day/night cycle. Indeed, key enzymes in the carbon fixation cycle are known to be regulated by the pH and the redox state of the chloroplast stroma. For example, RuBisCO activity is controlled by proton levels and magnesium ions (Tapia et al. 2000 , Andersson 2008 ). Fructose-1,6-bisphosphatase, seduheptulose-1,7-bisphosphatase (SBPase) and Phosphoribulokinase are all controlled by the redox state through the thioredoxin-ferredoxin system, and also by pH (Chiadmi et al. 1999 , Raines et al. 2000 , Raines 2003 ). Furthermore, Hendriks et al. showed the light dependency of the ADP-glucose pyrophosphorylase (Hendriks et al. 2003 ), which is part of the lumped reaction vStarch in our model. All these mechanisms will lead to a considerable reduction of the required stand-by flux of the CBB cycle, but are not yet included in our simple merged model.

                                    In the original formulation of our model without constant Ru5P supply or light-dependent regulation of CBB enzymes, low light intensities lead to a rapid collapse of the cycle. However, in sufficient light ATP levels are very high and carbon fixation rates are already saturated in moderate light conditions (Fig. 2 and Fig. S1 in Appendix S1). These findings indicate that the sets of parameters for the carbon fixation enzymes and the light reactions, derived from the respective original publications, might not be suitably adapted when employed in a merged, cooperating, system. This is not surprising considering that they originate from completely different systems and conditions. However, we wish to highlight here that systems biology models are known to include a ‘sloppy’ spectrum of parameter sensitivities, and yet still provide robust predictions (Gutenkunst et al. 2007 ).

                                    In order to systematically investigate the supply–demand behavior of the coupled system in different light conditions, we introduce a ‘regulation factor’ fCBB of the CBB cycle, by which all Vmax-values of the light-regulated enzymes (see above) are multiplied. This allows for a systematic variation of the energy demand by simulating accelerated or decelerated carbon fixation activity. Performing this variation under different light conditions gives insight into the synchronization of ATP and NADPH production and consumption rates, and thus enables a more profound analysis of the supply–demand regulation of photosynthesis (Brandes et al. 1996 , Chiadmi et al. 1999 , Raines et al. 2000 ). For the following steady-state analysis, the conceptual Ru5P influx reaction is not included.

                                    Fig. 5 displays stationary values of key model variables for different light intensities and regulation factors. In agreement with the observations presented above, that very low light intensities lead to a collapse of the cycle, ATP concentrations (Fig. 5A) are maximal (zero ADP), triose phosphate export (Fig. 5B) and starch production (Fig. 5C) are zero, and the lumenal pH (Fig. 5D) is very low (around 4). The latter is readily explained by the fact that the pH gradient built up by the low light cannot be reduced by the ATPase, which lacks the substrate ADP. Further, it becomes clear that the regulation factor of fCBB = 1, corresponding to the original parameters, is far from optimal. The ATP:ADP ratio remains very high, and TPT export and starch production rates are well below their optimum, regardless of the light intensities. The stationary lumenal pH further illustrates that parameters are not ideally adjusted. Not only for very low light, but also for moderate to high light conditions (above 300 μmol m −2 s −1 ) the lumen is dramatically acidic, indicating a mismatch in production and consumption processes. Increasing the regulation factor to values fCBB ≈ 4 leads to a dramatic improvement of the performance of the system. The ATP:ADP ratio assumes realistic and healthy values around one, triose phosphate export approximately doubles, and starch production increases by one order of magnitude compared with the original parameter values. Concomitantly, the lumenal pH remains moderate (pH 5.8, as suggested by Kramer et al. 1999 ). An advantage of mathematical modelling is that one can also predict the behavior of system variables, which are not easy to obtain experimentally. In Fig. S6 in Appendix S1, we exemplarily depict oxidized ferredoxin, oxidized PQ, relative NADP + and violaxanthin levels.

                                    (1) (2) where denotes the normalized control coefficient of reaction k on the steady-state carbon fixation rate. Fig. 6 displays the normalized overall control of demand reactions CDemand/(CDemand + CSupply), in dependence on different light intensities and carbon fixation regulation factors. Low light intensities and fast carbon fixation reactions shift the overall flux control to the supply reactions. This can readily be explained because under these conditions (low light and fast CBB enzymes) energy and redox provision by the light reactions are the limiting factor. Interestingly, PSII and PSI contribute strongest to the overall flux control on the supply side (Fig. S7 in Appendix S1). Conversely, high light intensities and slow carbon fixation reactions shift the overall flux control to the demand side, because under these conditions, the system is energetically saturated, and the bottleneck is in the CBB cycle consuming the energy and redox equivalents. Noteworthy, it is the SBPase reaction that exhibits the highest overall flux control (Fig. S8 in Appendix S1), while RuBisCO has only minor control.


                                    Why is photosynthesis important?

                                    Most organisms rely on photosynthesis for nutrients, either directly (plants) or indirectly (by relying on plants as an energy source). Organisms that undergo photosynthesis are called photoautotrophs. Photoautotrophs sustain themselves by synthesising their own food from carbon dioxide and other raw materials, and are referred to as the biosphere’s “producers”. In contrast, heterotrophs sustain themselves by relying on the material produced by other organisms and are referred to as the “consumers”. Humans are an example of heterotrophs.

                                    Photosynthesis is key for introducing energy into the ecosystem. It also removes lots of carbon dioxide from the atmosphere and produces oxygen for cellular respiration.

                                    Chloroplasts – the site of photosynthesis

                                    The site of photosynthesis in green plants are the leaves. Cells are able to carry out photosynthesis because of structures termed chloroplasts, which are the centre of the reactions. Chloroplasts are specialised membrane-bound organelles found only in plants and some types of algae. Each cell may have up to 200 chloroplasts.

                                    Chloroplasts are usually spherical or disc-shaped organelles in higher plants and are found in the cytosol of the cell. Chloroplasts contain a pigment called chlorophyll, which absorbs the light energy driving photosynthesis. It is chlorophyll that gives chloroplasts their green colour.

                                    Chloroplast structure

                                    Chloroplasts have an inner and an outer membrane often referred to as the chloroplast envelope. Inside the chloroplasts, there is a protein-rich substance called the stroma. Within the stroma, there is a second membrane system called the thylakoid membrane, which consists of disc-shaped sub-structures, arranged into stacks called grana. A single granum comprises around 20-60 thylakoids. The thylakoids contain light-harvesting complexes and photosynthetic pigments (i.e. chlorophyll), and it is here that the initial reactions of photosynthesis take place.


                                    4 DISCUSSION

                                    4.1 Regulation of ZEP activity

                                    Zeaxanthin epoxidase activity is known to be down-regulated upon increasing photoinhibition of PSII activity and hence photo-oxidative stress in chloroplasts (Reinhold et al., 2008 ). Accordingly, ZEP activity was nearly completely inhibited in tobacco or strongly down-regulated in the other species after 8 hr Hl treatment at 4°C, in parallel with PSII inactivation (Figures 2 and 3). Changes in ZEP protein amounts in response to HL stress have not been assessed so far. The HL-induced degradation of ZEP shown in the present work thus indicates an irreversible HL-induced damage of ZEP (analogous to D1). The HL-induced down-regulation of ZEP activity ensures that high levels of Zx are retained in response to prolonged HL stress to allow for efficient reactivation (or retention) of energy dissipation and thus photoprotection after intermediate LL phases. Hence, the inactivation of ZEP is understood as long-term memory of photo-oxidative stress (Jahns & Holzwarth, 2012 ). This regulatory principle also applies to in vivo conditions, since winter acclimation of evergreen plants was shown to be accompanied by retention of high Zx levels along with inactivation of PSII efficiency (Adams & Demmig-Adams, 1995 Adams, Demmig-Adams, Rosenstiel, Brightwell, & Ebbert, 2002 Adams, Demmig-Adams, & Verhoeven, 1995 Öquist & Huner, 2003 ). These characteristics support an essential photoprotective function of Zx during photoinhibition of PSII. Notably, such a memory function of Zx also applies to short-term down-regulation of PSII efficiency in context with the pH-regulated qE mechanism (Horton, Wentworth, & Ruban, 2005 ), which is also modulated by Zx. As the de-activation of qE by the lumen pH is much faster than the reconversion of Zx to Vx, Zx is retained also in the short-term, which ensures rapid reactivation of maximal qE capacity under rapidly fluctuating HL conditions.

                                    The regulation of ZEP activity is thus central for photoprotection and operates at different time scales. However, the molecular basis of ZEP regulation is not fully understood. Recent work showed that ZEP activity is regulated in the short-term by Trx (Da et al., 2017 ) and NTRC (Naranjo et al., 2016 ), implying an essential function of redox-sensitive sulfhydryl groups of ZEP protein in regulation. On basis of the proposed redox regulation, it can be assumed that ZEP has low or no activity in the dark, and is fully activated under illumination through Trx mediated reduction of specific sulfhydryl groups. In contrast, the molecular basis of HL-induced down-regulation of ZEP activity is unclear. Related to the proposed redox regulation of ZEP activity, however, it is tempting to speculate that ZEP is inactivated by reactive oxygen species (ROS), which possibly irreversibly oxidize redox-sensitive cysteine residues.

                                    4.2 Degradation of ZEP protein

                                    The observed HL-induced degradation of ZEP protein indicates that it is irreversibly damaged upon high photo-oxidative stress, possibly due to oxidation by ROS. Restoration of ZEP activity might thus require the degradation of inactive protein and import of newly synthesized protein. These characteristics resemble the general features of the well-known HL-induced D1 turnover. The very close correlation of the down-regulation of PSII and ZEP activity and the parallel degradation of D1 and ZEP suggests a coordinated regulation of ZEP activity/degradation related to PSII inactivation/D1 degradation, which might involve an interaction of PSII and ZEP. ZEP is localized at the stroma side of the membrane and should have access to the stroma exposed regions of the membrane only, but not to the inner part of the grana region, where functional PSII is located. Since damaged PSII centers are supposed to migrate to the stroma exposed regions of the membrane, it is conceivable that ZEP can interact particularly with damaged PSII. However, on basis of non-denaturing blue native gel electrophoresis, no co-migration of ZEP and PSII was detectable in thylakoids from HL treated plants (Schwarz et al., 2015 ), suggesting that either no interaction or only a weak interaction of the two proteins exists. Nevertheless, an interaction of damaged PSII and ZEP might explain, why inactivation and degradation of ZEP protein is enhanced in the presence of SM. Since ZEP is encoded in the nucleus, ZEP synthesis should not be influenced directly by SM, suggesting that accelerated ZEP degradation in the presence of SM is rather related to the inhibited D1 turnover and thus PSII repair. Consequently, ZEP degradation might be triggered by the accumulation of non-functional PSII, and hence under conditions when PSII repair cannot keep pace with PSII damage, either due to enhanced rates of PSII inhibition or due to reduced rates of PSII repair. The sensing of accumulation of non-functional PSII by ZEP does not necessarily require an interaction of damaged PSII and ZEP but could also be related to a, so far unknown, factor involved in PSII repair or reassembly. Interestingly, ZEP activity was found to be affected by SM even under moderate HL stress, independent of an additional inhibitory effect on PSII activity. Since SM had no direct effect on ZEP activity, this supports the view, that the accumulation of a putative signaling factor can also be triggered independent of pronounced inactivation of PSII in the presence of SM. Consequently, inactivation (and likely also degradation) of ZEP seems to be not directly related to inactivation of PSII but rather to a functional PSII repair mechanism.

                                    Irrespective of such a putative trigger factor, the down-regulation/degradation of ZEP upon accumulation of damaged PSII implies an essential function of Zx for photoprotection during the PSII repair cycle. Such a function is supported by the characteristics of the Arabidopsis xanthophyll cycle mutants npq1 and npq2 (Niyogi, Grossman, & Björkman, 1998 ), which revealed an increased HL sensitivity of Zx deficient npq1 (Havaux & Niyogi, 1999 Kalituho, Rech, & Jahns, 2007 Sarvikas, Hakala, Pätsikkä, Tyystjärvi, & Tyystjärvi, 2006 ) and a less pronounced HL sensitivity of Zx accumulating npq2 (Dall'Osto et al., 2005 Kalituho et al., 2007 ) compared to wild-type plants. Zx is known to be present in the thylakoid membrane either in association with antenna proteins or as non-protein bound molecule in the lipid phase of the membrane. Since no binding of the Zx to the PSII reaction center has been reported so far, it is thus likely that non-protein bound Zx serves as photoprotective xanthophyll during PSII repair. Such a function might be the basis for the shown qE-independent function of Zx (Havaux & Niyogi, 1999 ). This view is further supported by well-known increase of the VAZ pool size during long-term HL acclimation of plants (Bailey, Horton, & Walters, 2004 Demmig-Adams, Cohu, Muller, & Adams, 2012 Mishra et al., 2012 Schumann et al., 2017 ). Since long-term acclimation to HL also involves the reduction of the PSII antenna size and thus of xanthophyll binding sites, it is very likely that a significant fraction of the additionally accumulated VAZ pigments is not bound to antenna proteins. Recent work challenged the view that formed Zx rebinds to the Vx binding sites of the PSII antenna proteins (Xu et al., 2015 ). Thus it can be speculated, that Zx is generally located at the surface of antenna proteins and by that may contribute to protection of the PSII reaction center.

                                    4.3 Species-specific differences in HL sensitivity

                                    Plants of Arabidopsis and tobacco turned out to be more sensitive to HL than of pea and spinach. To determine putative specific physiological and morphological parameters of HL sensitivity, a comparative analysis of two more HL sensitive and two less HL sensitive species has been carried out. Based on our results, we suggest the following parameters to be crucial for the determination of HL sensitivity: (a) The presence of stroma-localized ZEP, (b) Leaf morphology and thylakoid membrane dynamics, (c) The pigment composition (VAZ pool size and Chl content per leaf area).

                                    The function of the stroma-localized ZEP protein determined in Arabidopsis (Schwarz et al., 2015 ) (Figure 1) is unclear. Interestingly, a stroma-localized fraction of ZEP was also found in tobacco chloroplasts, but not in the two less HL sensitive species, pea and spinach (Figure 1). Since the total amount of ZEP protein was found to be similar in all species (Figure 1), we conclude that the stroma-localized fraction does not represent an additional pool of ZEP protein, but that binding of a fraction of ZEP to the thylakoid membrane is restricted. The ZEP protein binds to the stroma exposed region of thylakoid membrane and interacts with the membrane through hydrophobic interactions (Schaller, Wilhelm, Strzałka, & Goss, 2012 Schwarz et al., 2015 ). Species-specific differences in the binding efficiency of ZEP protein to the thylakoid membrane might be due to differences in the properties of either the thylakoid membrane or the ZEP protein. Since the predicted amino acid sequences of the ZEP proteins in the different species show a high degree of identity, it seems unlikely that specific properties of the ZEP protein are responsible for the binding efficiency to the thylakoid membrane. It is unknown, however, which part of the protein is involved in binding to the membrane. It is further unclear, whether binding of ZEP requires a specific interaction with other proteins. In such a case, limited number of interactions partners present in the membrane might restrict ZEP binding to the membrane. Alternatively, different membrane properties could limit ZEP binding. For steric reasons, ZEP protein has only access to the stroma exposed regions of the membrane but not to the grana partitions. Thus, the relative portion of stroma exposed membranes might restrict ZEP binding as well. On basis of the EM analysis of thylakoid membrane organization (Figure 5), however, no obvious differences in thylakoid membrane organization were detectable among the different species under non-stressed conditions. It is thus most likely, that other membrane characteristics are responsible for the reduced binding of ZEP protein and hence the occurrence of a stroma-localized fraction of ZEP in Arabidopsis and tobacco.

                                    At the level of leaf morphology, particularly the parenchyma cell size and the number of layers of parenchyma cells correlated with the HL sensitivity. The two species with higher HL resistance, pea and spinach, were particularly characterized by about 50% smaller parenchyma cells (about 50 µm in pea and spinach vs. 100 µm in Arabidopsis and tobacco) leading to more layers of parenchyma cells. The resulting light gradient within the leaf due to shading and light scattering might reduce light absorption in chloroplasts of inner layers of cells and hence induce reduced inactivation of PSII relative to the total number of chloroplasts. Earlier work further showed that a lower Chl content per leaf area is accompanied by higher susceptibility to photoinhibition (Patsikka, Kairavuo, Šeršen, Aro, & Tyystjärvi, 2002 ). Therefore, the lower Chl content per leaf area found here for Arabidopsis and tobacco plants (Table 2) could contribute to the higher HL sensitivity of these two species. Moreover, recent work showed that structural rearrangement of the thylakoid membrane is accompanied with light-induced activation of qE (Schumann et al., 2017 ) and further regulates the balance between linear and cyclic electron transfer (Wood et al., 2018 ). Therefore, structural reorganization of membrane stacking is likely important for the activation of photoprotective mechanisms. The less pronounced thylakoid membrane reorganization in Arabidopsis and tobacco chloroplast in response to HL exposure compared to pea and spinach (Figure 5) might thus further determine the more pronounced HL sensitivity in these two species.

                                    One of the most striking species-specific feature of the two more HL resistant species, pea and spinach, is the strongly increased VAZ pool size (Table 2) compared to Arabidopsis and tobacco. The increased VAZ pool size has important consequences related to photoprotective properties. The large VAZ pool size provides a large fraction of non-protein bound Zx in the lipid phase of the thylakoid membrane, which might contribute significantly to the protection of damaged PSII monomers during the PSII repair cycle in non-appressed regions of the membrane, either due to ROS de-activation in the lipid phase of the membrane or by promoting energy dissipation upon interaction with PSII reaction centers. ROS de-activation might be of particular importance, because earlier work showed that D1 protein synthesis and hence repair of PSII is vulnerable to ROS (Kojima et al., 2007 Nishiyama et al., 2001 ), while contribution to energy dissipation would require an interaction of Zx with PSII, either at specific binding sites or at the surface of PSII reactions center or antenna proteins. Moreover, non-protein bound Zx is supposed to affect the membrane properties with respect to fluidity and/or stability, similar to tocopherols (Havaux, 1998 ), and changes in the membrane fluidity are supposed to be involved in the PSII repair cycle (Goral et al., 2010 Yoshioka-Nishimura, 2016 ). The less flexible thylakoid membrane structure in response to HL (Figure 5) observed in the two species with a small VAZ pool size, Arabidopsis and tobacco, thus strongly supports such a role of Zx. Since an increase of the VAZ pool size is a typical long-term HL acclimation response of plants (Bailey et al., 2004 Demmig-Adams et al., 2012 Mishra et al., 2012 Schumann et al., 2017 ), we propose that this parameter is an important factor determining the different HL sensitivities of the studied plant species.


                                    Light Absorption by Reaction-Center Chlorophylls Causes a Charge Separation across the Thylakoid Membrane

                                    The absorption of a quantum of light of wavelength � nm causes a chlorophyll a molecule to enter the first excited state. The energy of such photons increases the energy of chlorophyll a by 42 kcal/mol. In the reaction center, this excited-state energy is used to promote a charge separation across the thylakoid membrane: an electron is transported from a chlorophyll molecule to the primary electron acceptor, the quinone Q, on the stromal surface of the membrane, leaving a positive charge on the chlorophyll close to the luminal surface (Figure 16-38). The reduced primary electron acceptor becomes a powerful reducing agent, with a strong tendency to transfer the electron to another molecule. The positively charged chlorophyll, a strong oxidizing agent, will attract an electron from an electron donor on the luminal surface. These potent biological reductants and oxidants provide all the energy needed to drive all subsequent reactions of photosynthesis: electron transport, ATP synthesis, and CO2 fixation.

                                    Figure 16-38

                                    The primary event in photosynthesis. After a photon of light of wavelength � nm is absorbed by one of the many chlorophyll molecules in one of the light-harvesting complexes (LHCs) of an antenna (only one is shown), some of the absorbed energy (more. )

                                    The significant features of the primary reactions of photosynthesis are summarized in the following model, in which P represents the chlorophyll a in the reaction center, and Q represents the primary electron acceptor:

                                    According to this model, the ground state of the reaction-center chlorophyll, P, is not a strong enough reductant to reduce Q that is, an electron will not move spontaneously from P to Q. However, the excited state of the reactioncenter chlorophyll, P*, is an excellent reductant and rapidly (in about 10 � seconds) donates an electron to Q, generating P + and Q − . This photochemical electron movement, which depends on the unique environment of both the chlorophylls and the acceptor within the reaction center, occurs nearly every time a photon is absorbed. The acceptor, Q − , is a powerful reducing agent capable of transferring the electron to still other molecules, ultimately to NADP + . The powerful oxidant P + can remove electrons from other molecules to regenerate the original P. In plants, the oxidizing power of four molecules of P + is used, by way of intermediates, to remove four electrons from H2O to form O2:

                                    Chlorophyll a also absorbs light at discrete wavelengths shorter than 680 nm (see Figure 16-37b). Such absorption raises the molecule into one of several higher excited states, which decay within 10 � seconds (1 picosecond, ps) to the first excited state P*, with loss of the extra energy as heat. Photochemical charge separation occurs only from the first excited state of the reaction-center chlorophyll a, P*. This means that the quantum yield — the amount of photosynthesis per absorbed photon — is the same for all wavelengths of visible light shorter than 680 nm.

                                    The chlorophyll a molecules within reaction centers are capable of directly absorbing light and initiating photosynthesis. However, even at the maximum light intensity encountered by photosynthetic organisms (tropical noontime sun, 𢒁.2 ×� 20 photons/m 2 /s), each reaction-center chlorophyll a absorbs about one photon per second, which is not enough to support photosynthesis sufficient for the needs of the plant. To increase the efficiency of photosynthesis, especially at more typical light intensities, organisms utilize additional light-absorbing pigments.


                                    Light Independent Reactions and Carbon Fixation

                                    A short introduction

                                    The general principle of carbon fixation is that some cells under certain conditions can take CO2 and reduce it to a usable cellular form. Most of us are aware that green plants can take up CO2 and produce O2 in during photosynthesis. We have just discussed the ability of a cell to transfer light energy onto chemicals and ultimately to produce the energy carriers ATP and NADPH in a process known as the light reactions. In photosynthesis, the plant cells use the ATP and NADPH formed during photophosphorylation to reduce CO2 to sugar, (as we will see, specifically G3P, an intermediate in glycolysis also) in what are called the dark reactions. While we appreciate that this process happens in green plants, photosynthesis had its evolutionary origins in the bacterial world. In this module we will go over the general reactions of the Calvin Cycle, a reductive pathway that incorporates CO2 into cellular material.

                                    In photosynthetic bacteria such as Cyanobacteria and purple non-sulfur bacteria, as well as in plants, the energy (ATP) and reducing power (NADPH) obtained from the light reactions is coupled to "Carbon Fixation", the incorporation of inorganic carbon (CO2) into organic molecules initially as glyceraldehyde-3-phosphate (G3P) and eventually into glucose. Organisms that can obtain all of their required carbon from an inorganic source (CO2) are referred to as autotrophs, while those organisms that require organic sources of carbon, such as glucose or amino acids, are referred to as heterotrophs. The biological pathway that leads to carbon fixation in green plants and cyanobacteria is called the Calvin Cycle and is a reductive pathway (consumes energy and electrons) which leads to the reduction of CO2 to G3P. There are at least five other pathways for carbon fixation, used by other autotrophic prokaryotes.

                                    The Calvin Cycle: the reduction of CO2 to Glyceraldehyde 3-Phosphate

                                    Light reactions harness energy from the sun to produce chemical bonds, ATP, and NADPH. A stylized chloroplast is shown above. These energy-carrying molecules are made in the stroma where carbon fixation (here the Calvin Cycle, drawn in an extremely simplified form on the right) takes place.

                                    In plant cells, the Calvin cycle takes place in the stroma (outside of the thylakoids) of the chloroplasts. While the process is similar in cyanobacteria, there are no specific organelles that house the Calvin Cycle and the reactions occur in the cytoplasm around a complex intracellular membrane system derived from the plasma membrane. There is strong evidence that supports the hypothesis that the origin (actually, several independent origins) of was from a symbiosis between cyanobacteria and nonphotosynthetic cells.

                                    Note that in Dr. Britt's class you will not have to memorize the Calvin cycle you only need to know what goes in, the first step, and what comes out. Dr. Britt suggests this video, which may lack style, but it's clear on the basics.

                                    Stage 1: Carbon Fixation

                                    In the stroma of plant chloroplasts, in addition to CO2, two other components are present to initiate the light-independent reactions: an enzyme called ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), and ribulose bisphosphate (RuBP), as shown in the figure below. Ribulose-1,5-bisphosphate (RuBP) is composed of five carbon atoms and includes two phosphates.

                                    The Calvin cycle has three stages. In stage 1, the enzyme RuBisCO incorporates carbon dioxide into an organic molecule, 3-PGA. In stage 2, the organic molecule is reduced using electrons supplied by NADPH. In stage 3, RuBP, the molecule that starts the cycle, is regenerated so that the cycle can continue. Only one carbon dioxide molecule is incorporated at a time, so the cycle must be completed three times to produce a single three-carbon GA3P molecule, and six times to produce a six-carbon glucose molecule. Here the Calvin cycle is presented in a (slightly less) symplified form.

                                    RuBisCO (ribulose bis phosphate carboxylase) catalyzes a reaction between CO2 and RuBP. For each CO2 molecule that reacts with one RuBP, a 6-Carbon molecules is formed. The resulting 6 C molecule will immediately split to form two molecules of another 3-carbon compound (3-PGA). Note that we have added only one C (Carbon) to an existing 5 C chain to make these 2 3-C molecules! Thus we will have to run this initial phase three times to "bleed off" a 3-C and regenerate the 3 RuBP's that were employed to capture each CO2.

                                    Stage 2: Reduction

                                    ATP and NADPH are used to convert the six molecules of 3-PGA into six molecules of a 3C molecule: glyceraldehyde 3-phosphate (G3P) - a carbon compound that you might or might not remember from glycolysis. Six molecules of both ATP and NADPH are used up in the process, helping to drive the reactions and produce the electrons required to reduce the incoming CO2. The "spent" molecules (ADP and NADP + ) return to the nearby thylakoids to be recycled back into ATP and NADPH.

                                    Stage 3: Regeneration

                                    Remember that we have consumed 3 molecules of RuBP to bleed off one G3P. Thus we need to somehow regenerate these 3 RuBP from the remaining 5 G3Ps. This regeneration phase might be politely described as "interesting", or more colloquially as "A Frigging Nightmare". Three more molecules of ATP are used in these regeneration reactions. It is included for your enjoyment below. Again, Dr. Britt will only quiz you on what goes into the Calvin cycle, its first step, and what comes out.


                                    Thylakoid vs Stroma

                                    The chloroplasts are flat structures found in the cytoplasm of plant cells. They consist of thylakoids which are small membrane-bound compartments. They are the sites of the light-dependent reaction of photosynthesis. Thylakoid is usually stacked to form structures called grana. Stroma is also an important component of the chloroplast. It is a colorless fluid matrix situated in the inner portion of the chloroplast. The thylakoids are surrounded by stroma. The stroma is the site where the light-independent reactions of photosynthesis take place. The enzymes and pigments which are essential for photosynthesis are usually embedded in both thylakoid and stroma. This can be described as the difference between Thylakoids and Stroma.

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                                    Reference:

                                    1.“Mitochondria and chloroplasts.” Khan Academy. Available here
                                    2.“Photophosphorylation (Cyclic and Non-Cyclic).” Photophosphorylation (Cyclic and Non-Cyclic) Available here
                                    3.The Editors of Encyclopædia Britannica. “Chloroplast.” Encyclopædia Britannica, Encyclopædia Britannica, inc., 17 Oct. 2016. Available here

                                    Image Courtesy:

                                    1.’Thylakoid2′ Public Domain via Commons Wikimedia
                                    2.’Chloroplast structure’ By Kelvinsong – Own work (CC BY-SA 3.0) via Commons Wikimedia