PowerPoint PresentationSlide 2Three stages of the Calvin Cycle:Slide 4Basic scheme of the Calvin CycleSlide 6Stage 1: Fixation of CO2 to form 3-phosphoglycerateRubisco reaction can be broken down into four basic stepsSlide 9Stage 2: Reduction of 3-phosphoglycerate to form hexose sugarsSlide 11Stage 3: Regeneration of ribulose-1,5-bisphosphate“Carbon shuffle” reactionsThe net reaction of the Calvin Cycle can be broken down into two componentsWhy must the Calvin Cycle be regulated?Slide 16Slide 17Activation by thioredoxin-mediated reduction of disulfide bridgesPhotorespiration and RubiscoSlide 20Slide 21Slide 22C4 and CAM Carbon Fixation PathwaysTwo variations of the "Hatch-Slack" pathwayC4 Pathway in SugarcaneSlide 26Slide 27CAM PathwaySlide 29Carbon Fixation:The Calvin cycle, light regulation of carbon fixation, photorespiration in C4 and CAM plantsBioc 460 Spring 2008 - Lecture 32 (Miesfeld)Sugarcane plants use C4 carbon fixation to limit photorespirationMelvin Calvin won a 1961 Nobel Prize for discovering carbon fixationRibulose 1,5-bisphosphate carboxylase (Rubisco) is the most abundant enzyme on planet Earth•The photosynthetic electron transport chain operates in the light to generate chemical energy for use in the carbon fixation reactions of the Calvin cycle.•The Calvin cycle enzyme Rubisco carboxylates ribulose bisphosphate (RUBP) to form a C-6 intermediate that is rapidly cleaved to form two moles 3-phosphoglycerate; three turns of the cycle are needed to generate one mole of glyceraldehyde-3P (GAP) from three moles of CO2.•Light activates enzymes in the Calvin cycle by two primary mechanisms, 1) increased Rubisco activity in response to elevated pH and Mg2+ in the stroma, and 2) thioredoxin-mediated reduction of disulfide bonds.•Photorespiration is a wasteful side reaction of Rubisco that uses O2 to generate 2-phosphoglycerate which must be metabolized in peroxisomes. The C4 and CAM carbon fixation pathways minimize the effects of photorespiration by increasing the local concentratation of CO2 in the chloroplast stroma.Key Concepts in Carbon FixationThree stages of the Calvin Cycle:Fixation, Reduction, and Regeneration Plants store light energy in the form of carbohydrate, primarily starch and sucrose. The carbon and oxygen for this process comes from CO2, and the energy for the energy for carbon fixation is derived from the ATP and NADPH made during photosynthesis.The conversion of CO2 to carbohydrate is called the Calvin Cycle and is named after Melvin Calvin who discovered it. The Calvin Cycle requires the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase commonly called rubisco. The Calvin cycle generates the triose phosphates 3-phosphoglycerate, glyceraldehyde-3P (GAP) and dihydroxyacetone phosphate, all of which are used to synthesize the hexose phosphates fructose-1,6-bisphosphate and fructose 6-phosphate.Hexose phosphates produced by the Calvin Cycle are converted to:1) sucrose for transport to other plant tissues2) starch for energy stores within the cell3) cellulose for cell wall synthesis4) pentose phosphates for metabolic intermediatesBasic scheme of the Calvin Cycle Three turns of the cycle results in the fixation of three molecules of CO2 Key reaction in stage 1, catalyzed by rubisco enzyme, combines three molecules of ribulose-1,5-bisphosphate (RuBP), a five carbon (C5) compound, with three molecules of CO2 to form six molecules of the C3 compound 3-phosphoglycerateBasic scheme of the Calvin Cycle The Calvin Cycle is sometimes called the Dark Reactions, but do not be fooled by this name - the Calvin Cycle is the most active during the daylight hours when ATP and NADPH are plentiful.The net reaction of three turns of the Calvin cycle can be written as:3 CO2 + 3 RuBP + 6 NADPH + 9 ATP + 6 H2O →1 GAP + 3 RuBP + 6 NADP+ + 9 ADP + 9 PiIf we just look at the fate of the carbons coming from CO2 (C1) in this reaction, we see that one net C3 compound (glyceraldehydes-3P) is formed and three C5 molecules (RuBP) are regenerated:3 C1 + 3 C5 → 1 C3 + 3 C5Why property of Calvin Cycle reactions make them Dark Reactions?Stage 1: Fixation of CO2 to form 3-phosphoglycerate To identify the metabolic intermediates in this process, Calvin and his colleagues used radioactive labeling with 14CO2 to follow carbon fixation in photosynthetic algae cells grown in culture.They found that within a few seconds of adding 14CO2 to the culture, the cells accumulated 14C-labeled 3-phosphoglycerate, suggesting that this was the first product of the carboxylation reaction. Within a minute of adding 14CO2 to the culture, they found numerous compounds were labeled with 14C, many of which were later identified as Calvin Cycle intermediates.Rubisco reaction can be broken down into four basic steps 1) formation of an enediolate intermediate of RuBP2) carboxylation by nucleophilic attack on the CO23) hydration of 2-carboxy-3-keto-D-arabinitol-1,5-bisphosphate4) aldol cleavage to form two molecules of 3-phosphoglycerateThe rubisco reaction is very exergonic (ΔG°' = -35.1 kJ/mol), with the aldol cleavage step being a major contributor to the favorable change in free energyRubisco is a multisubunit enzyme consisting of eight identical catalytic subunits at the core surrounded by eight smaller subunits that function to stabilize the complex and presumably enhance enzyme activity. Considering that rubisco plays a central role in all photosynthetic autotrophic organisms on earth, of which ~85% are photosynthetic plants and microorganisms that inhabit the oceans, rubisco is the most abundant enzyme on this planet.Stage 2: Reduction of 3-phosphoglycerate to form hexose sugars 3-phosphoglycerate (product of the rubisco reaction) is converted to glyceraldehyde-3-phosphate (GAP) by two isozymes of phosphoglycerate kinase and glyceraldehyde-3P dehydrogenase.It is these two reactions that use the ATP and NADPH made during the light reactions. cleavage.Stage 2: Reduction of 3-phosphoglycerate to form hexose sugars Remember that for every 3 CO2 that are fixed by carboxylation of 3 RuBP molecules, six moles of 3-phosphoglycerate are generated by aldol cleavage.Therefore, 6 ATP and 6 NADPH are required for every 3 CO2 that are converted to one net glyceraldehyde-3P. An additional 3 ATP are used in stage 3 to regenerate these 3 RuBP molecules.3 CO2 + 3 RuBP + 6 NADPH + 9 ATP + 6 H2O →1 GAP + 3 RuBP + 6 NADP+ + 9 ADP + 9 PiStage 3: Regeneration of ribulose-1,5-bisphosphateIn this final
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