Mechanism of Rubisco:  The Carbamate as General Base

Ww, Cleland; Tj, Andrews; Gutteridge, Steven; Fc, Hartman; Gh, Lorimer

doi:10.1021/cr970010r

Cited by 393 publications

(436 citation statements)

References 99 publications

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“…To carboxylate RuBP, rubisco active sites need to first bind two cofactors: a nonsubstrate CO 2 and a Mg 2+ ion (31,32). This binding results in the activated holoenzyme [enzyme-CO 2 -Mg 2+ (ECM)].…”

Section: Resultsmentioning

confidence: 99%

Characterization of the heterooligomeric red-type rubisco activase from red algae

Loganathan

Tsai

Mueller‐Cajar

2016

Proc. Natl. Acad. Sci. U.S.A.

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The photosynthetic CO 2 -fixing enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase (rubisco) is inhibited by nonproductive binding of its substrate ribulose-1,5-bisphosphate (RuBP) and other sugar phosphates. Reactivation requires ATP-hydrolysis-powered remodeling of the inhibited complexes by diverse molecular chaperones known as rubisco activases (Rcas). Eukaryotic phytoplankton of the red plastid lineage contain so-called red-type rubiscos, some of which have been shown to possess superior kinetic properties to green-type rubiscos found in higher plants. These organisms are known to encode multiple homologs of CbbX, the α-proteobacterial red-type activase. Here we show that the gene products of two cbbX genes encoded by the nuclear and plastid genomes of the red algae Cyanidioschyzon merolae are nonfunctional in isolation, but together form a thermostable heterooligomeric Rca that can use both α-proteobacterial and red algal-inhibited rubisco complexes as a substrate. The mechanism of rubisco activation appears conserved between the bacterial and the algal systems and involves threading of the rubisco large subunit C terminus. Whereas binding of the allosteric regulator RuBP induces oligomeric transitions to the bacterial activase, it merely enhances the kinetics of ATP hydrolysis in the algal enzyme. Mutational analysis of nuclear and plastid isoforms demonstrates strong coordination between the subunits and implicates the nuclearencoded subunit as being functionally dominant. The plastid-encoded subunit may be catalytically inert. Efforts to enhance crop photosynthesis by transplanting red algal rubiscos with enhanced kinetics will need to take into account the requirement for a compatible Rca.rubisco | activase | photosynthesis | AAA + proteins | red algae

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Section: Resultsmentioning

confidence: 99%

Characterization of the heterooligomeric red-type rubisco activase from red algae

Loganathan

Tsai

Mueller‐Cajar

2016

Proc. Natl. Acad. Sci. U.S.A.

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“…By sequestering carbonic anhydrase and RuBisCO together, the carboxysome increases the local concentration of CO 2 in order to drive its reaction with ribulose-1,5-bisphosphate (RuBP) at the active site of RuBisCO, an enzyme whose catalytic properties are notoriously poor. 14,15 Carboxysomes, which were first visualized more than 40 years ago, [16][17][18] can be classified into two types, a and b. 9 a-Carboxysomes contain Form 1A RuBisCO and generally have their genes for RuBisCO, carbonic anhydrase, and the shell proteins encoded together in a single operon.…”

Section: Introductionmentioning

confidence: 99%

Insights from multiple structures of the shell proteins from the β‐carboxysome

Sawaya²,

et al. 2008

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Carboxysomes are primitive bacterial organelles that function as a part of a carbon concentrating mechanism (CCM) under conditions where inorganic carbon is limiting. The carboxysome enhances the efficiency of cellular carbon fixation by encapsulating together carbonic anhydrase and the CO 2 -fixing enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO). The carboxysome has a roughly icosahedral shape with an outer shell between 800 and 1500 Å in diameter, which is constructed from a few thousand small protein subunits. In the cyanobacterium Synechocystis sp. PCC 6803, the previous structure determination of two homologous shell protein subunits, CcmK2 and CcmK4, elucidated how the outer shell is formed by the tight packing of CcmK hexamers into a molecular layer. Here we describe the crystal structure of the hexameric shell protein CcmK1, along with structures of mutants of both CcmK1 and CcmK2 lacking their sometimes flexible C-terminal tails. Variations in the way hexamers pack into layers are noted, while sulfate ions bound in pores through the layer provide further support for the hypothesis that the pores serve for transport of substrates and products into and out of the carboxysome. One of the new structures provides a high-resolution (1.3 Å ) framework for subsequent computational studies of molecular transport through the pores. Crystal and solution studies of the C-terminal deletion mutants demonstrate the tendency of the terminal segments to participate in proteinAprotein interactions, thereby providing a clue as to which side of the molecular layer of hexameric shell proteins is likely to face toward the carboxysome interior.

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“…Specifically, the CCM functions to supply CO 2 to ribulose bisphosphate carboxylase/oxygenase (RuBisCO), the primary carboxylating enzyme of the CBB cycle. High levels of CO 2 are essential to cyanobacterial metabolism because RuBisCO has relatively slow carboxylation kinetics and is promiscuous, catalyzing an off-pathway reaction with O 2 called oxygenation (2)(3)(4).…”

mentioning

confidence: 99%

pH determines the energetic efficiency of the cyanobacterial CO ₂ concentrating mechanism

Mangan

Flamholz

Hood

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

177

181

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Many carbon-fixing bacteria rely on a CO 2 concentrating mechanism (CCM) to elevate the CO 2 concentration around the carboxylating enzyme ribulose bisphosphate carboxylase/oxygenase (RuBisCO). The CCM is postulated to simultaneously enhance the rate of carboxylation and minimize oxygenation, a competitive reaction with O 2 also catalyzed by RuBisCO. To achieve this effect, the CCM combines two features: active transport of inorganic carbon into the cell and colocalization of carbonic anhydrase and RuBisCO inside proteinaceous microcompartments called carboxysomes. Understanding the significance of the various CCM components requires reconciling biochemical intuition with a quantitative description of the system. To this end, we have developed a mathematical model of the CCM to analyze its energetic costs and the inherent intertwining of physiology and pH. We find that intracellular pH greatly affects the cost of inorganic carbon accumulation. At low pH the inorganic carbon pool contains more of the highly cellpermeable H 2 CO 3 , necessitating a substantial expenditure of energy on transport to maintain internal inorganic carbon levels. An intracellular pH ≈8 reduces leakage, making the CCM significantly more energetically efficient. This pH prediction coincides well with our measurement of intracellular pH in a model cyanobacterium. We also demonstrate that CO 2 retention in the carboxysome is necessary, whereas selective uptake of HCO 3 − into the carboxysome would not appreciably enhance energetic efficiency. Altogether, integration of pH produces a model that is quantitatively consistent with cyanobacterial physiology, emphasizing that pH cannot be neglected when describing biological systems interacting with inorganic carbon pools.carbon fixation | RuBisCO | cyanobacteria | inorganic carbon | systems biology C yanobacteria and many other autotrophs use a CO 2 concentrating mechanism (CCM) to increase the cellular pool of inorganic carbon and facilitate the Calvin-Benson-Bassham (CBB) cycle (1). Specifically, the CCM functions to supply CO 2 to ribulose bisphosphate carboxylase/oxygenase (RuBisCO), the primary carboxylating enzyme of the CBB cycle. High levels of CO 2 are essential to cyanobacterial metabolism because RuBisCO has relatively slow carboxylation kinetics and is promiscuous, catalyzing an off-pathway reaction with O 2 called oxygenation (2-4).RuBisCO oxygenation produces 2-phosphoglycolate (2PG), which is not part of the CBB cycle and must be recycled. Recycling 2PG through photorespiratory pathways is costly, consuming reduced carbon and energy resources (5, 6). The problem of RuBisCO's limited specificity is all the more pronounced because there is ≈ 20 times more O 2 than CO 2 in aqueous solutions equilibrated with present-day atmosphere (SI Appendix, Fig. S1). CCMs overcome these problems by concentrating CO 2 near RuBisCO, favorably increasing the ratio of CO 2 to O 2 . High concentrations of CO 2 maximize the rate of carboxylation and competitively inhibit oxygenation. Indeed, it is widel...

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Mechanism of Rubisco: The Carbamate as General Base

Cited by 393 publications

References 99 publications

Characterization of the heterooligomeric red-type rubisco activase from red algae

Characterization of the heterooligomeric red-type rubisco activase from red algae

Insights from multiple structures of the shell proteins from the β‐carboxysome

pH determines the energetic efficiency of the cyanobacterial CO ₂ concentrating mechanism

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Mechanism of Rubisco: The Carbamate as General Base

Cited by 393 publications

References 99 publications

Characterization of the heterooligomeric red-type rubisco activase from red algae

Characterization of the heterooligomeric red-type rubisco activase from red algae

Insights from multiple structures of the shell proteins from the β‐carboxysome

pH determines the energetic efficiency of the cyanobacterial CO 2 concentrating mechanism

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pH determines the energetic efficiency of the cyanobacterial CO ₂ concentrating mechanism