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Phonon-assisted electron-proton transfer in [FeFe] hydrogenases: Topological role of clusters.

ABSTRACT: [FeFe] hydrogenases are enzymes that have acquired a unique capacity to synthesize or consume molecular hydrogen (H₂). This function relies on a complex catalytic mechanism involving the active site and two distinct electron and proton transfer networks working in concert. By an analysis based on terahertz vibrations of [FeFe] hydrogenase structure, we are able to predict and identify the existence of rate-promoting vibrations at the catalytic site and the coupling with functional residues involved in reported electron and proton transfer networks. Our findings suggest that the positioning of the cluster is influenced by the response of the scaffold to thermal fluctuations, which in turn drives the formation of networks for electron transfer through phonon-assisted mechanisms. Thus, we address the problem of linking the molecular structure to the catalytic function through picosecond dynamics, while raising the functional gain brought by the cofactors or clusters, using the concept of fold-encoded localized vibrations.

SUBMITTER: Chalopin Y

PROVIDER: S-EPMC10147833 | biostudies-literature | 2023 Apr

REPOSITORIES: biostudies-literature

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Phonon-assisted electron-proton transfer in [FeFe] hydrogenases: Topological role of clusters.

Chalopin Yann Y Cramer Stephen P SP Arragain Simon S

Biophysical journal 20230323 8

[FeFe] hydrogenases are enzymes that have acquired a unique capacity to synthesize or consume molecular hydrogen (H<sub>2</sub>). This function relies on a complex catalytic mechanism involving the active site and two distinct electron and proton transfer networks working in concert. By an analysis based on terahertz vibrations of [FeFe] hydrogenase structure, we are able to predict and identify the existence of rate-promoting vibrations at the catalytic site and the coupling with functional res ...[more]

PMID: 36960530

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Project description:A non-bifurcating NADH-dependent, dimeric [FeFe]-hydrogenase (HydAB) from Syntrophus aciditrophicus was heterologously produced in Escherichia coli, purified and characterized. Purified recombinant HydAB catalyzed NAD+ reduction coupled to hydrogen oxidation and produced hydrogen from NADH without the involvement of ferredoxin. Hydrogen partial pressures (2.2-40.2 Pa) produced by the purified recombinant HydAB at NADH to NAD+ ratios of 1-5 were similar to the hydrogen partial pressures generated by pure and cocultures of S. aciditrophicus (5.9-36.6 Pa). Thus, the hydrogen partial pressures observed in metabolizing cultures and cocultures of S. aciditrophicus can be generated by HydAB if S. aciditrophicus maintains NADH to NAD+ ratios greater than one. The flavin-containing beta subunits from S. aciditrophicus HydAB and the non-bifurcating NADH-dependent S. wolfei Hyd1ABC share a number of conserved residues with the flavin-containing beta subunits from non-bifurcating NADH-dependent enzymes such as NADH:quinone oxidoreductases and formate dehydrogenases. A number of differences were observed between sequences of these non-bifurcating NADH-dependent enzymes and [FeFe]-hydrogenases and formate dehydrogenases known to catalyze electron bifurcation including differences in the number of [Fe-S] centers and in conserved residues near predicted cofactor binding sites. These differences can be used to distinguish members of these two groups of enzymes and may be relevant to the differences in ferredoxin-dependence and ability to mediate electron-bifurcation. These results show that two phylogenetically distinct syntrophic fatty acid-oxidizing bacteria, Syntrophomonas wolfei a member of the phylum Firmicutes, and S. aciditrophicus, a member of the class Deltaproteobacteria, possess functionally similar [FeFe]-hydrogenases that produce hydrogen from NADH during syntrophic fatty acid oxidation without the involvement of reduced ferredoxin. The reliance on a non-bifurcating NADH-dependent [FeFe]-hydrogenases may explain the obligate requirement that many syntrophic metabolizers have for a hydrogen-using partner microorganism when grown on fatty, aromatic and alicyclic acids.

Project description:The [FeFe]-hydrogenases are enzymes that catalyze the reversible activation of H2 coupled to the reduction-oxidation of electron carriers. Members of the different taxonomic groups of [FeFe]-hydrogenases display a wide range of preference, or bias, for H2 oxidation or H2 production reactions, despite sharing a common catalytic cofactor, or H-cluster. Identifying the properties that control reactivity remains an active area of investigation, and models have emerged that include diversity in the catalytic site coordination environments and compositions of electron transfer chains. The kinetics of proton-coupled electron transfer at the H-cluster might be expected to be a point of control of reactivity. To test this hypothesis, systematic changes were made to the conserved cysteine residue that functions in proton exchange with the H-cluster in the three model enzymes: CaI, CpII, and CrHydA1. CaI and CpII both employ electron transfer accessory clusters but differ in bias, whereas CrHydA1 lacks accessory clusters having only the H-cluster. Changing from cysteine to either serine (more basic) or aspartate (more acidic) modifies the sidechain pKa and thus the barrier for the proton exchange step. The reaction rates for H2 oxidation or H2 evolution were surveyed and measured for model [FeFe]-hydrogenases, and the results show that the initial proton-transfer step in [FeFe]-hydrogenase is tightly coupled to the control of reactivity; a change from cysteine to more basic serine favored H2 oxidation in all enzymes, whereas a change to more acidic aspartate caused a shift in preference toward H2 evolution. Overall, the changes in reactivity profiles were profound, spanning 105 in ratio of the H2 oxidation-to-H2 evolution rates. The fact that the change in reactivity follows a common trend implies that the effect of changing the proton-transfer residue pKa may also be framed as an effect on the scaling relationship between the H-cluster di(thiolmethyl)amine (DTMA) ligand pKa and E m values of the H-cluster. Experimental observations that support this relationship, and how it relates to catalytic function in [FeFe]-hydrogenases, are discussed.

Dataset Information

Phonon-assisted electron-proton transfer in [FeFe] hydrogenases: Topological role of clusters.

Publications

Phonon-assisted electron-proton transfer in [FeFe] hydrogenases: Topological role of clusters.

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