6GM8

[FeFe]-hydrogenase CpI from Clostridium pasteurianum, variant E282Q

Summary for 6GM8

• Entry DOI: 10.2210/pdb6gm8/pdb
• Related: 4XDC
• Descriptor: Iron hydrogenase 1, dicarbonyl[bis(cyanide-kappaC)]-mu-(iminodimethanethiolatato-1kappaS:2kappaS)-mu-(oxomethylidene)diiron(2+), IRON/SULFUR CLUSTER, ... (6 entities in total)
• Functional Keywords: hydrogenase, h-cluster, oxidoreductase, semisynthetic enzyme
• Biological source: Clostridium pasteurianum
• Total number of polymer chains: 2
• Total formula weight: 134168.43

Authors

Duan, J.,Esselborn, J.,Hofmann, E.,Winkler, M.,Happe, T. (deposition date: 2018-05-24, release date: 2018-11-07, Last modification date: 2024-01-17)

Primary citation

Duan, J.,Senger, M.,Esselborn, J.,Engelbrecht, V.,Wittkamp, F.,Apfel, U.P.,Hofmann, E.,Stripp, S.T.,Happe, T.,Winkler, M.
Crystallographic and spectroscopic assignment of the proton transfer pathway in [FeFe]-hydrogenases.
Nat Commun, 9:4726-4726, 2018

PubMed Abstract: The unmatched catalytic turnover rates of [FeFe]-hydrogenases require an exceptionally efficient proton-transfer (PT) pathway to shuttle protons as substrates or products between bulk water and catalytic center. For clostridial [FeFe]-hydrogenase CpI such a pathway has been proposed and analyzed, but mainly on a theoretical basis. Here, eleven enzyme variants of two different [FeFe]-hydrogenases (CpI and HydA1) with substitutions in the presumptive PT-pathway are examined kinetically, spectroscopically, and crystallographically to provide solid experimental proof for its role in hydrogen-turnover. Targeting key residues of the PT-pathway by site directed mutagenesis significantly alters the pH-activity profile of these variants and in presence of H their cofactor is trapped in an intermediate state indicative of precluded proton-transfer. Furthermore, crystal structures coherently explain the individual levels of residual activity, demonstrating e.g. how trapped HO molecules rescue the interrupted PT-pathway. These features provide conclusive evidence that the targeted positions are indeed vital for catalytic proton-transfer.

PubMed: 30413719
DOI: 10.1038/s41467-018-07140-x

Experimental method

X-RAY DIFFRACTION (1.96 Å)