CHEM 445 / BIOL 445
Biochemistry II

J. D. Cronk
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12. header

Lecture 12. CO2 fixation by RuBisCO

Wednesday 14 February 2007

Introduction to the Calvin cycle. What is the metabolite formed when CO2 is fixed? The reactions catalyzed by RuBisCO. RuBisCO structure and function. The mechanism of RuBisCO and the role of magnesium.

Reading: BTS6, Ch.20, pp.565-570.

 

12. Summary

Lecture 12 Summary

In today's lecture, we are introduced to the Calvin cycle, the process by which plants use the energy (ATP) and reducing power (NADPH) generated by the light reactions to "fix" carbon dioxide and synthesize carbohydrates. Then we undertake a close examination of the important enzyme that incorporates carbon dioxide into a five-carbon phosphosugar, ribulose 1,5-bisphosphate carboxylase/oxygenase, or RuBisCO. What is the metabolite formed when CO2 is fixed? Carbon dioxide fixation is catalyzed by Ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO). The RuBisCO carboxylase reaction: reactants, products, and mechanism. The role of magnesium (Mg2+) in the RuBisCo mechanism. RuBisCo catalyzes a "wasteful" oxygenase reaction.

 

RuBisCO - the carboxylase reaction

RuBisCO is an acronym for ribulose 1,5-bisphosphate carboxylase/oxygenase [EC 4.1.1.39]. This name acknowleges the fact that the enzyme carries out two distinct reactions - carboxylation of ribulose 1,5-bisphosphate (RuBP) and oxygenation of RuBP. RuBisCO is arguably one of the most important proteins of the earth's biosphere, since all life on the planet essentially depends on the ability of photosynthetic organisms to "fix" CO2 as carbohydrates. In fact, RuBisCO was the PDB's Molecule of the Month for November 2000.

The carboxylation reaction carried out by RuBisCO initiates the photosynthetic conversion of carbon dioxide into carbohydrates. Calvin's experiments suggested that the carboxylation by CO2 of a 2-carbon acceptor occurs as the first step of CO2 fixation in photosynthetic green algae. In fact, the CO2 acceptor is a 5-carbon compound, ribulose 1,5-bisphosphate (RuBP), which subsequently splits into two 3-carbon products, as illustrated in the equation below.

 
Equation for RuBisCO carboxylase reaction  

RuBisCO structure

RuBisCO from plant chloroplasts is composed of eight copies each of a large (L) and small (S) chain. The structure has 422 symmetry, meaning the presence of a four-fold symmetry axis and two two-fold symmetry axes that are all mutually perpendicular.

RuBisCO mechanism and the role of magnesium

The mechanism of RuBisCO depends critically upon the presence of a magnesium ion in the active site. The carboxylation of a lysine residue by addition of carbon dioxide to the e-amino group of Lys provides a negatively-charged carbamide ligand to Mg2+. Thus, carbon dioxide is not only a substrate, but also participates as an activating component that is required for enzyme function. The bound magnesium is essential in facilitating the conversion of the RuBP substrate to an enediolate form of RuBP. The formation of the enediolate is the rate-limiting step of the RuBisCO mechanism, and is also the first step in the mechanism of the competing oxygenase reaction. The enediolate acts as a nucleophile toward a substrate CO2 molecule (second step in the figure below). Here the magnesium again facilitates the reaction, this time helping to stabilize the developing negative charge on CO2 as it forms the new carbon-carbon bond to RuBP. An anology can be made between the role of magnesium in rubisco and that of zinc in carbonic anhydrase. The carboxylation of RuBP forms an unstable b-keto acid intermediate that beomes hydrated. The latter then undergoes a cleavage reaction that leads directly to the formation of a 3-phosphoglycerate molecule, and the carbanion form of another.

Mechanism of RuBisCO reaction

RuBisCO - the oxygenase reaction

When oxygen replaces carbon dioxide in the active site of RuBisCO, it can catalyze the oxygenase reaction. A mechanistic portrayal of this reaction is diagrammed below.

Diagram of mechanism for oxygenase reaction catalyzed by RuBisCO
Note that the oxygenase reaction depends on the formation of the same enediolate form of RuBP as in the carboxylase reaction. The products, on the other hand, are the 2-carbon molecule phosphoglycolate and the 3-carbon 3-phosphoglycerate (3PG), instead of the two 3PG molecules that result from the carboxylase reaction. The oxygenase reaction is certainly nonproductive, in terms of carbon fixation. Moreover, it is held to be wasteful due to the energy lost as the carbon atoms from two molecules of 2-phosphoglycolate are reconfigured into serine plus CO2. Since oxygen is input and carbon dioxide given off, this process is called photorespiration.

Study questions

  • Explain the catalytic role of Mg2+ in the RuBisCO mechanism.
  • Explain the role of CO2 in the function of RuBisCO other than its role as a substrate.
  • Describe the oxygenase reaction catalyzed by RuBisCO and its mechanism.
  • Think of a plausible factor helping to account for RuBisCO's apparent lack of substrate specificity.

Page updated 12-27-06

References

1. Berg, Tymoczko, and Stryer. Biochemistry (BTS): 6th edition (2007, Freeman) Ch.20, pp.565-570.

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