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Kinetic modeling of the Calvin cycle identifies flux control and stable metabolomes in Synechocystis carbon fixation.

ABSTRACT: Biological fixation of atmospheric CO2 via the Calvin-Benson-Bassham cycle has massive ecological impact and offers potential for industrial exploitation, either by improving carbon fixation in plants and autotrophic bacteria, or by installation into new hosts. A kinetic model of the Calvin-Benson-Bassham cycle embedded in the central carbon metabolism of the cyanobacterium Synechocystis sp. PCC 6803 was developed to investigate its stability and underlying control mechanisms. To reduce the uncertainty associated with a single parameter set, random sampling of the steady-state metabolite concentrations and the enzyme kinetic parameters was employed, resulting in millions of parameterized models which were analyzed for flux control and stability against perturbation. Our results show that the Calvin cycle had an overall high intrinsic stability, but a high concentration of ribulose 1,5-bisphosphate was associated with unstable states. Low substrate saturation and high product saturation of enzymes involved in highly interconnected reactions correlated with increased network stability. Flux control, that is the effect that a change in one reaction rate has on the other reactions in the network, was distributed and mostly exerted by energy supply (ATP), but also by cofactor supply (NADPH). Sedoheptulose 1,7-bisphosphatase/fructose 1,6-bisphosphatase, fructose-bisphosphate aldolase, and transketolase had a weak but positive effect on overall network flux, in agreement with published observations. The identified flux control and relationships between metabolite concentrations and system stability can guide metabolic engineering. The kinetic model structure and parameterizing framework can be expanded for analysis of metabolic systems beyond the Calvin cycle.

SUBMITTER: Janasch M

PROVIDER: S-EPMC6363089 | biostudies-other | 2019 Feb

REPOSITORIES: biostudies-other

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Kinetic modeling of the Calvin cycle identifies flux control and stable metabolomes in Synechocystis carbon fixation.

Janasch Markus M Asplund-Samuelsson Johannes J Steuer Ralf R Hudson Elton P EP

Journal of experimental botany 20190201 3

Biological fixation of atmospheric CO2 via the Calvin-Benson-Bassham cycle has massive ecological impact and offers potential for industrial exploitation, either by improving carbon fixation in plants and autotrophic bacteria, or by installation into new hosts. A kinetic model of the Calvin-Benson-Bassham cycle embedded in the central carbon metabolism of the cyanobacterium Synechocystis sp. PCC 6803 was developed to investigate its stability and underlying control mechanisms. To reduce the unce ...[more]

PMID: 30371804

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Project description:Very few bacteria are able to fix carbon via both the reverse tricarboxylic acid (rTCA) and the Calvin-Benson-Bassham (CBB) cycles, such as symbiotic, sulfur-oxidizing bacteria that are the sole carbon source for the marine tubeworm Riftia pachyptila, the fastest-growing invertebrate. To date, the coexistence of these two carbon fixation pathways had not been found in a cultured bacterium and could thus not be studied in detail. Moreover, it was not clear if these two pathways were encoded in the same symbiont individual, or if two symbiont populations, each with one of the pathways, coexisted within tubeworms. With comparative genomics, we show that Thioflavicoccus mobilis, a cultured, free-living gammaproteobacterial sulfur oxidizer, possesses the genes for both carbon fixation pathways. Here, we also show that both the CBB and rTCA pathways are likely encoded in the genome of the sulfur-oxidizing symbiont of the tubeworm Escarpia laminata from deep-sea asphalt volcanoes in the Gulf of Mexico. Finally, we provide genomic and transcriptomic data suggesting a potential electron flow toward the rTCA cycle carboxylase 2-oxoglutarate:ferredoxin oxidoreductase, via a rare variant of NADH dehydrogenase/heterodisulfide reductase in the E. laminata symbiont. This electron-bifurcating complex, together with NAD(P)+ transhydrogenase and Na+ translocating Rnf membrane complexes, may improve the efficiency of the rTCA cycle in both the symbiotic and the free-living sulfur oxidizer.IMPORTANCE Primary production on Earth is dependent on autotrophic carbon fixation, which leads to the incorporation of carbon dioxide into biomass. Multiple metabolic pathways have been described for autotrophic carbon fixation, but most autotrophic organisms were assumed to have the genes for only one of these pathways. Our finding of a cultivable bacterium with two carbon fixation pathways in its genome, the rTCA and the CBB cycle, opens the possibility to study the potential benefits of having these two pathways and the interplay between them. Additionally, this will allow the investigation of the unusual and potentially very efficient mechanism of electron flow that could drive the rTCA cycle in these autotrophs. Such studies will deepen our understanding of carbon fixation pathways and could provide new avenues for optimizing carbon fixation in biotechnological applications.

Project description:Hydrogen gas (H(2)) is a possible future transportation fuel that can be produced by anoxygenic phototrophic bacteria via nitrogenase. The electrons for H(2) are usually derived from organic compounds. Thus, one would expect more H(2) to be produced when anoxygenic phototrophs are supplied with increasingly reduced (electron-rich) organic compounds. However, the H(2) yield does not always differ according to the substrate oxidation state. To understand other factors that influence the H(2) yield, we determined metabolic fluxes in Rhodopseudomonas palustris grown on (13)C-labeled fumarate, succinate, acetate, and butyrate (in order from most oxidized to most reduced). The flux maps revealed that the H(2) yield was influenced by two main factors in addition to substrate oxidation state. The first factor was the route that a substrate took to biosynthetic precursors. For example, succinate took a different route to acetyl-coenzyme A (CoA) than acetate. As a result, R. palustris generated similar amounts of reducing equivalents and similar amounts of H(2) from both succinate and acetate, even though succinate is more oxidized than acetate. The second factor affecting the H(2) yield was the amount of Calvin cycle flux competing for electrons. When nitrogenase was active, electrons were diverted away from the Calvin cycle towards H(2), but to various extents, depending on the substrate. When Calvin cycle flux was blocked, the H(2) yield increased during growth on all substrates. In general, this increase in H(2) yield could be predicted from the initial Calvin cycle flux. Photoheterotrophic bacteria, like Rhodopseudomonas palustris, obtain energy from light and carbon from organic compounds during anaerobic growth. Cells can naturally produce the biofuel H(2) as a way of disposing of excess electrons. Unexpectedly, feeding cells organic compounds with more electrons does not necessarily result in more H(2). Despite repeated observations over the last 40 years, the reasons for this discrepancy have remained unclear. In this paper, we identified two metabolic factors that influence the H(2) yield, (i) the route taken to make biosynthetic precursors and (ii) the amount of CO(2)-fixing Calvin cycle flux that competes against H(2) production for electrons. We show that the H(2) yield can be improved on all substrates by using a strain that is incapable of Calvin cycle flux. We also contributed quantitative knowledge to the long-standing question of why photoheterotrophs must produce H(2) or fix CO(2) even on relatively oxidized substrates.

Dataset Information

Kinetic modeling of the Calvin cycle identifies flux control and stable metabolomes in Synechocystis carbon fixation.

Publications

Kinetic modeling of the Calvin cycle identifies flux control and stable metabolomes in Synechocystis carbon fixation.

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