Carbon Fixation in Plants

Question 1

What are primary producers?

Answer 1

Photosynthetic organisms, turning CO2 into biomass using the energy of the sun
The process where photosynthesisers fix CO2 into carbohydrate = the light independent reaction
Also called the Calvin cycle or the reductive pentose phosphate pathway
It is an autocatalytic cycle

Question 2

How was the Calvin cycle discovered?

Answer 2

Using algae and adding radioactively labelled 14CO2, in order to trace the carbon and determine the products produced

Question 3

What is the first stage of the Calvin cycle?

Answer 3

Carboxylation:
Ribulose-1,5-bisphosphate (RuBP) is carboxylated with CO2
Using Ribulose bisphosphate carboxylase (Rubisco)
Forms 3-phosphoglycerate (3PG)

3x RuBP + 3x CO2 = 6 x 3PG

Question 4

Describe Rubisco?

Answer 4

Most abundant protein in the biosphere
8 large subunits and 8 small subunits
L8S8 has D4 symmetry (symmetry of a square prism)
It requires an Mg2+ cofactor to stabilse negative charges

Question 5

What is the mechanism of Rubisco?

Answer 5

The reaction proceeds via an enediolate intermediate that nucleophilically attacks CO2 to form a beta-keto acid
The intermediate reacts with water to form 2 molecules of 3PG

Question 6

What is the second stage of the Calvin cycle?

Answer 6

Phosphorylation of 3PG:
3-phosphoglycerate is catalysed by phosphoglycerate kinase and 6x ATP to form 1,3-bisphosphoglycerate (BPG)

6x 3PG = 6x 1,3-BPG

Question 7

What is the third stage of the Calvin cycle?

Answer 7

1,3-Bisphosphoglycerate is catalysed by glyceraldehyde-3-phosphate dehydrogenase and 6x NADPH to form glyceraldehyde-3-phosphate (GAP)

6x 1,3-BPG = 6x GAP

Question 8

What happens to the 6x GAP after stage 3 of the Calvin cycle?

Answer 8

1x GAP from 3CO2 goes off for carbohydrate synthesis

5x GAP needs to be resynthesized into 3x RuBP

Question 9

What is stage 4 of the Calvin cycle?

Answer 9

The start of regeneration of RuBP:
GAP -> DHAP using TIM
DHAP can be converted into fructose-1,6-bisphosphate using aldolase
OR
combining with Erythrose-4-phosphate (E4P) to form Sedoheptulose-1,7-bisphosphate (SBP)

E4P is regenerated later

Question 10

How are the remaining 5x GAP resynthesised in stage 5 of the Calvin cycle?

Answer 10

The 5x 3C molcules are shuffled around in order to regenerate 3x 5C

DHAP + GAP -> F1,6BP (aldolase)
GAP + F1,6BP -> Erythrose-4-phosphate, E4P + Xu5P (transketolase)
DHAP + E4P -> sedoheptulose-1,7-bisphosphate, S1,7BP (aldolase)
GAP + S1,7BP -> Xu5P + R5P (transketolase)

Question 11

What is stage 6 of the Calvin cycle?

Answer 11

The final regeneration:
Using Xu5P and R5P from the previous reactions - they are converted back to ribulose-1,5-bisphosphate in order to continue the cycle

Ribose-5-phosphate (R5P) -> Ribulose-5-phosphate (Ru5P) (using ribose phosphate isomerase)
Xylulose-5-phosphate (Xu5P) -> Ribulose-5-phosphate (Ru5P) (using phosphopentose epimerase)
Finally:
Ribulose-5-phosphate (Ru5P) -> RuBP (using Phosphoribulose kinase and ATP hydrolysis)

Question 12

Give a rundown of the molcules in the Calvin cycle?

Answer 12

Ribulose-1,5-bisphosphate (RuBP)
3-phosphoglycerate (3PG)
1,3-bisphosphoglycerate
Glyceraldehyde-3-phosphate

Intermediates: DHAP, F1,6BP, S1,7BP, E4P

Xylulose-5-phosphate (Xu5P)
Ribose-5-phosphate (R5P)
Ribulose-5-phosphate (Ru5P)

Question 13

What is the overal stoichiometry of the Calvin cycle?

Answer 13

3 CO2 + 9 ATP + 6 NADPH → GAP + 9 ADP + 8 Pi + 6 NADP+

Question 14

How is the Cavlin cycle regulated?

Answer 14

The cycle needs ATP and NADPH produced by photosynthetic electron transport and photophosphorylation
Therefore the activity of the cycle is regulated by photosynthetic electron transport rates (the redox potential)
It is controlled indirectly by light

Ferredoxin (which is the electron acceptor from PSI) is the source of the electrons

Question 15

How can the Calvin cycle be controlled?

Answer 15

Control of Fructose-1,6-bisphosphate
Ferredoxin thioredoxin reductase (intermediate) with a disulphide bond - this reduces thiol groups of thioredoxin
Thioredoxin reduces the target enzyme bisphosphatase i.e. the reduced state with disulphide bonds is active
Many enzymes within the cycle are regulated in this way

Question 16

How can the Calvin cycle be controlled by pH and Mg2+?

Answer 16

Dark - pH 7 and 1-3 mM Mg2+
Light - pH 8 and 3-6 mM Mg2+
Rubisco has an optimum pH around 8 and Mg2+ increase stimulates it as well

pH increases as protons are being moved into the thylakoid lumen
This allows several Calvin cycle enzymes to be activated

Question 17

What are some outcomes of the GAP used for synthesis?

Answer 17

Converted into F6P and then G1P (by phosphoglucose isomerase and phosphoglucomutase)
Converted into F-1,6-BP, before being coupled with UDP-glucose/UDP reaction to form sucrose-6-phosphate and then sucrose

Question 18

What competes with photosynthesis?

Answer 18

Photorespiration

A process where plants consume oxygen and release carbon dioxide in the light (NOT MITOCHONDRIAL RESPIRATION)

Question 19

Why can photorespiration take place?

Answer 19

Rubisco is an oxygenase as well as carboxylase activity
Ribulose bisphosphate can either fix CO2 or O2

RuBP + CO2 = 2x 3PG
RuBP + O2 = 1x 3PG + 1x 2-phosphoglycolate

When it fixes O2 we don’t get that ‘extra’ molecule of GAP to produce carbohydrates with, therefore we essentially lose this carbon

Question 20

What is the pathway of converting 2-phosphoglycolate - Choloroplast and Peroxisome?

Answer 20

Chloroplast:
2-PG -> Glycolate (using phosphoglycolare phosphatase)
Glycolate is exported to the peroxisome

Peroxisome:
Glycolate -> Glyoxylate (using glycolate oxidase) - this also forms hydrogen peroxide which is dealt with by catalase
Glyoxylate -> glycine (in a transamination reaction) and is exported to the mitochondrion

Question 21

What is the pathway of converting 2-phosphoglycolate - Mitochondria, peroxisome and chloroplast?

Answer 21

Mitochondria:
2x glycine -> 1x serine + CO2, serine is transported back to the peroxisome
Peroxisome:
serine -> hydroxypyruvate (transamination)
hydroxypyruvate -> glycerate (using hydroxypyruvate reductase and NADH)
Cytosol:
Glycerate -> 3PG (using glycerate kinase and ATP)
Chloroplast:
3PG is reconverted into RuBP in the Calvin cycle

Question 22

What are the disadvantages of photorespiration?

Answer 22

Fixed C is lost, also fixed N
Photorespiration can reduce C3 crop photosynthetic efficiency by 20-50%
ATP and NADH are used up recycling

Question 23

What are the advantages of photorespiration?

Answer 23

It has evolved to protect the plants from excess light energy = exceeding the capacity of the Calvin cycle to use up the ATP and NADPH
This prevents hypereduction and the photosynthetic apparatus from damage = photoinhibition
It uses up ATP and releases CO2
It also detoxifies P-glycolate and allows recycling of C that would otherwise be lost

Question 24

What are C4 plants?

Answer 24

C4 plants have evolved independently to eliminate photorespiration
Stomata close under conditions of high temperature or low water availability
When stomata are closed CO2 can’t enter
Many tropical plants (sugar cane, maize, sorghum) have solved this problem by physically separating CO2 fixation and the Calvin cycle
This allows them to reduce stomatal opening to limit water loss while ensuring rubisco is saturated for CO2 and effectively eliminates photorespiration

Question 25

What is the structure of C4 plants?

Answer 25

The mesophyll cell carry out the initial CO2 fixation into C4 acid (no rubisco in the mesophyll)
Kranz anatomy - bundle sheath cells form bundles where the Calvin cycle is
The C4 compound is transported to bundle sheath cells where it is decarboxylated and the CO2 refixed by the Calvin cycle
This leads to lack of O2 fixation

They have an enzyme called PEP carboxylase
They have little PSII but lots of rubisco

Question 26

What is the mechanism of a C4 plant?

Answer 26

Mesophyll cell:
PEP -> oxaloacetate (using PEP carboxylase)
Oxaloacetate -> malate (using malate dehydrogenase)
Bundle sheath cell:
malate -> pyruvate + 3PG (using malic enzyme, NADP+ and CO2)
3PG goes to the Calvin cycle

Keeping the cycle going
Pyruvate -> PEP (using pyruvatephosphate dikinase)

Question 27

Why could C4 plants be useful in the future?

Answer 27

C4 plants photosynthesise more efficiently at low CO2 levels
Due to CO2 concentrating mechanism
C3 plants function efficiently at current CO2 concentrations and future levels
But rising CO2 = rising temperature and C4 plants function more efficiently at higher temperatures

Question 28

How can we convert C3 plants to C4 plants?

Answer 28

With increasing temperatures leads to an interest in reducing/eliminating photorespiration:

Engineer rubisco to limit O2 fixation - very hard
Change C3 into C4 plants - trying with rice
Provide an alternative/more efficient photorespiratory pathway e.g. Synthetic biology approaches e.g. In tobacco