Photosynthesis

1. Photosynthesis The carbon reactions (Dark Reactions)

2. Overall Perspective • Dark Reactions: • Expend chemical energy • Fix Carbon [convert CO2 to organic form] • Light reactions: • Harvest light energy • Convert light energy to chemical energy

3. At the end of the light reactions • The reaction of the light reaction is: • CO2 +H2O (CH2O) + O2 • Recent estimates indicate that about 200 billion tones of CO2 (Mr = 44) are converted to biomass each year • 40 % of this is from marine phytoplankton • The bulk of the carbon is incorporated into organic compounds by the carbon reducing reactions (dark reactions) of photosynthesis

4. At the end of the light reactions • The reactions catalyzing the reduction of CO2 to carbohydrates are coupled to the consumption of NADPH and ATP by enzymes found in the stroma • fluid environment • These reactions were thought to be independent of the light reactions • So the name “dark reactions”stuck • However, these chemical reactions are regulated by light • So are called the “carbon reactions” of photosynthesis

5. Overview of the carbon reactions • The Calvin cycle: • Stage 1: • CO2 accepted by Ribulose-1,5-bisphosphate. • This undergoes carboxylation • Has a carboxyl group (-COOH) attached to it • At the end of stage 1, CO2 covalently linked to a carbon skeleton forming two 3-phosphycerate molecules.

6. Carboxylation: The first step is the most important • Step 1: The enzyme RUBISCO (Ribulose bis-phosphate carboxylase oxygenase) carries out this conversion • Rubisco accounts for 40% of the protein content of chloroplasts • is likely the most abundant protein on Earth • Rubisco is, in fact, very inefficient, and that a mechanism has evolved to deal with this handicap

7. Overview of the carbon reactions • The Calvin cycle: • Stage 2: • Each of the two 3-phosphycerate molecules are altered. • First phosphorylated through the use of the 3 ATPs generated during the light reaction. • Then reduced through the use of the 2 NADPHs generated during the light reaction. • Forms a carbohydrate • glyceraldehyde-3-phosphate

8. 3-phosphycerate molecules are altered • First phosphorylated through the use of the 3 ATP molecules generated during the light reaction • Forms 1,3-bisphosphoglycerate • Then reduced through the use of the 2 NADPH molecules generated during the light reaction • Forms glyceraldehyde-3-phosphate • Note the formation of triose phosphate

9. Overview of the carbon reactions • The Calvin cycle: • Stage 3: • Regeneration of Ribulose-1,5-bisphosphate. • This requires the coordinated action of eight reaction steps • And thus eight specific enzymes • Three molecules of Ribulose-1,5-bisphosphate are formed from the reshuffling of carbon atoms from triose phosphate.

10. Regeneration of Ribulose-1,5-bisphosphate • The Calvin cycle reactions regenerate the biochemical intermediates needed for operation • More importantly, the cycle is Autocatalytic • Rate of operation can be enhanced by increasing the concentration of the intermediates in the cycle • So, Calvin cycle has the metabolically desirable of producing more substrate than is consumed • Works as long as the produced triose phosphate is NOT diverted elsewhere (as in times of stress or disease)

11. Overview of the carbon reactions • The Calvin cycle: • The cycle runs six times: • Each time incorporating a new carbon. Those six carbon dioxides are reduced to glucose: • Glucose can now serve as a building block to make: • polysaccharides • other monosaccharides • fats • amino acids • nucleotides

12. Only one-sixth of the triose phosphate is used for polysaccharide production • Synthesis of polysaccharides, such as starch and sucrose, provide a sink • Ensures an adequate flow of carbon atoms through the cycleIFCO2 is constantly available • During a steady rate of photosynthesis 5/6 of the triose phosphates are used for the regeneration of Ribulose-1,5-bisphosphate • 1/6 is transported to the cytosol for the synthesis of sucrose or other metabolites that are converted to starch in the chloroplast

13. Regulation of the Calvin cycle • The high energy efficiency of the Calvin cycle indicates that some form of regulation ensures that all intermediates in cycle: • Are present at adequate concentrations • The cycle is turned off when it is not needed in the dark • Remember: • These are the “carbon reactions”, NOT the “dark reactions” • Many factors regulate the Calvin cycle

14. Regulation of the Calvin cycle • 1: The pH of the stroma increases as protons are pumped out of it through the membrane assembly of the light reactions. • The enzymes of the Calvin Cycle function better at this higher pH. • 2: The reactions of the Calvin cycle have to stop when they run out of substrate • as photosynthesis stops, there is no more ATP or NADPH in the stroma for the dark reactions to take place.

15. Regulation of the Calvin cycle • 3: The light reactions increase the permeability of the stromal membrane to required cofactors • Mg ions are required for the Calvin Cycle. • 4: Several enzymes of the Calvin Cycle are activated by the breaking of disulphide bridges of enzymes involved in the working of the cycle. • the activity of the light reactions is communicated to the dark reactions by an enzyme intermediate

16. When conditions are not optimum

17. Photorespiration • Occurs when the CO2 levels inside a leaf become low • This happens on hot dry days when a plant is forced to close its stomata to prevent excess water loss • If the plant continues to attempt to fix CO2 when its stomata are closed • CO2 will get used up and the O2 ratio in the leaf will increase relative to CO2 concentrations • When the CO2 levels inside the leaf drop to around 50 ppm, • Rubisco starts to combine O2 with Ribulose-1,5-bisphosphate instead of CO2

18. Photorespiration • Instead of producing 2 3C PGA molecules, only one molecule of PGA is produced and a toxic 2C molecule called phosphoglycolateis produced • The plant must get rid of the phosphoglycolate • The plant immediately gets rid of the phosphate group • converting the molecule to glycolic acid

19. Photorespiration • Theglycolic acid is then transported to the peroxisome and there converted to glycine • Peroxisomes are ubiquitous organelles that function to rid cells of toxic substances • The glycine (4 carbons) is then transported into a mitochondria where it is converted into serine(3 carbons) • Releases CO2

20. Photorespiration • The serine is then used to make other organic molecules • All these conversions cost the plant energy and results in the net lost of CO2 from the plant • 75% of the carbon lost during the oxygenation of Rubisco is recovered during photorespiration and is returned to the Calvin cycle

21. The C4 Carbon cycle

22. The C4 carbon Cycle • The C4 carbon Cycle occurs in 16 families of both monocots and dicots. • Corn • Millet • Sugarcane • Maize • There are three variations of the basic C4 carbon Cycle • Due to the different four carbon molecule used

23. The C4 carbon Cycle • This is a biochemical pathway that prevents photorespiration • C4 leaves have TWO chloroplast containing cells • Mesophyll cells • Bundle sheath (deep in the leaf so atmospheric oxygen cannot diffuse easily to them) • C3 plants only have Mesophyll cells • Operation of the C4 cycle requires the coordinated effort of both cell types • No mesophyll cells is more than three cells away from a bundle sheath cells • Many plasmodesmata for communication

24. The C4 carbon Cycle • Four stages: • Stage 1: • In Mesophyll cell • Fixation of CO2 by the carboxylation of phosphenol-pyruvate (primary acceptor molecule) • forms a C4 acid molecule • Malic acid and/or aspartate • Stage 2: • Transport of the C4 acid molecule to the bundle sheath cell

25. The C4 carbon Cycle • Stage 3: • Decarboxylationof theC4 acid molecule (in bundle sheath) • Makes a C3 acid molecule • This generates CO2 • This CO2 is reduced to carbohydrate by the Calvin cycle • Stage 4: • The C3 acid molecule (pryuvate) is transported back to mesophyll cells • Regeneration of phosphenol-pyruvate

26. The C4 carbon Cycle • Regeneration of phosphenol-pyruvate consumestwo high energy bonds from ATP • Movement between cells is by diffusion via plasmodesmata • Movement within cells is regulated by concentration gradients • This system generates a higher CO2 conc in bundle sheath cells than would occur by equilibrium with the atmosphere • Prevents photorespiration!!!!!!!!!!

27. The C4 carbon Cycle • The net effect of the C4 carbon Cycle is to convert a dilute CO2 solution in the mesophyll into a concentrated solution in the bundle sheath cells • This requires more energy than C3 carbon plants • BUT – This energy requirement is constant no matter what the environmental conditions • Allows more efficient photosynthesis in hotter conditions

28. Crassulacean Acid Metabolism (CAM Plants)

29. CAM Plants • The CAM mechanism enables plants to improve water efficiency • CAM plant • Loses 50 – 100 g water for every gram of CO2 gained • C4 plant • Loses 250 – 300 g water for every gram of CO2 gained • C3 plant • Loses 400 – 500 g water for every gram of CO2 gained • Similar to C4 cycle • In CAM plants formation of the C4 acid is both temporally and spatially separated

30. CAM Plants • At night: • Stomata only open at night when it is cool • CO2 is captured by phosphenol-pyruvate carboxylase in the cytosol – leaves become acidic • The malic acid formed is stored in the vacuole • Amount of malic acid formed is equal to the amount of CO2 taken in

31. CAM Plants • During the day: • Stomata close, preventing water loss, and further uptake of CO2 • Malic acid is transported to the chloroplast and decarboxylated to release CO2 • This enters the Calvin cycle as it can not escape the leaf • Pyruvate is converted to starch in the chloroplast – regenerates carbon acceptor

32. Phosphorylation regulates phosphenol-pyruvate (PEP) carboxylase • CAM and C4 plants require a separation of the initial carboxylation from the following de-carboxylation • Diuranal regulation is used • IN CAM PLANTS:- • Phosphorylation of the serine residue of phosphenol-pyruvate (PEP) carboxylase (Ser-OP) yields a form of the enzyme which is active at night • This is relatively insensitive to malic acid

33. Photophorylation regulates phosphenol-pyruvate (PEP)carboxylase • During the day: • De-Phosphorylation of the serine (ser-OH) gives a form of the enzyme which is inhibited by malic acid • THIS IS THE OPPOSITE WAY AROUND FOR C4 PLANTS!

34. Summary • The reduction of CO2 to carbohydrate via photosynthesis is coupled to the consumption of ATP and NADPH • CO2 is reduced via the Calvin cycle • Takes place in the stroma (soluble phase) • CO2 and water combine with Ribulose-1,5-bisphosphatein the following reaction • CO2 +H2O (CH2O) + O2 • Regeneration of the carrier is required for the cycle to continue

35. Summary • The Calvin cycle requires the joint action of several light-dependant systems • Changes in ions (Mg+ and H+) • Changes in effector metabolites (enzyme substrates) • Changes in protein-mediated systems (rubisco activase) • Rubisco can also act as an oxygenase • The carboxylation & oxygenation reactions take place at the active sites of rubisco.

36. Summary • C4 and CAM plants Prevent photorespiration!!!!! • C4 leaves have TWO chloroplast containing cells • Mesophyll cells • Bundle sheath • CAM Plants drastically reduce water lass • CAM plant • Loses 50 – 100 g water for every gram of CO2 gained • C4 plant • Loses 250 – 300 g water for every gram of CO2 gained • C3 plant • Loses 400 – 500 g water for every gram of CO2 gained

37. Any Questions?