Roles of the F-domain in [FeFe] hydrogenase |
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Authors: | Charles Gauquelin Carole Baffert Pierre Richaud Emma Kamionka Emilien Etienne David Guieysse Laurence Girbal Vincent Fourmond Isabelle André Bruno Guigliarelli Christophe Léger Philippe Soucaille Isabelle Meynial-Salles |
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Institution: | 1. LISBP, Université de Toulouse, CNRS, INRA, INSA, Toulouse, France;2. Aix Marseille Univ, CNRS, BIP, Marseille, France;3. CEA, CNRS, Aix-Marseille Université, Institut de Biosciences et Biotechnologies Aix-Marseille, UMR 7265, Laboratoire de Bioénergétique et Biotechnologie des Bactéries et Microalgues, CEA Cadarache, Saint-Paul-lez-Durance F-13108, France |
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Abstract: | The role of accessory Fe-S clusters of the F-domain in the catalytic activity of M3-type FeFe] hydrogenase and the contribution of each of the two Fe-S surface clusters in the intermolecular electron transfer from ferredoxin are both poorly understood. We designed, constructed, produced and spectroscopically, electrochemically and biochemically characterized three mutants of Clostridium acetobutylicum CaHydA hydrogenase with modified Fe-S clusters: two site-directed mutants, HydA_C100A and HydA_C48A missing the FS4C and the FS2 surface Fe-S clusters, respectively, and a HydA_ΔDA mutant that completely lacks the F-domain. Analysis of the mutant enzyme activities clearly demonstrated the importance of accessory clusters in retaining full enzyme activity at potentials around and higher than the equilibrium 2H+/H2 potential but not at the lowest potentials, where all enzymes have a similar turnover rate. Moreover, our results, combined with molecular modelling approaches, indicated that the FS2 cluster is the main gate for electron transfer from reduced ferredoxin. |
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Keywords: | [Fe-Fe] hydrogenase H-cluster Electron transfer pathway Accessory domains Ferredoxin |
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