title: Reactivation from the Ni-B state in [NiFe] hydrogenase of Ralstonia eutropha is controlled by reduction of the superoxidised proximal cluster.
abstract: The tolerance towards oxic conditions of O2-tolerant [NiFe] hydrogenases has been attributed to an unusual [4Fe-3S] cluster that lies proximal to the [NiFe] active site. Upon exposure to oxygen, this cluster converts to a superoxidised (5+) state, which is believed to secure the formation of the so-called Ni-B state that is rapidly reactivated under reducing conditions. Here, the reductive reactivation of the membrane-bound [NiFe]-hydrogenase (MBH) from Ralstonia eutropha in a native-like lipid membrane was characterised and compared to a variant that instead carries a typical [4Fe-4S] proximal cluster. Reactivation from the Ni-B state was faster in the [4Fe-4S] variant, suggesting that the reactivation rate in MBH is limited by the reduction of the superoxidised [4Fe-3S] cluster. We propose that the [4Fe-3S] cluster plays a major role in protecting MBH by blocking the reversal of electron transfer to the [NiFe] active site, which would produce damaging radical oxygen species.
authors.affiliation: School of Biomedical Sciences, The Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds LS2 9JT, UK. L.J.C.Jeuken@leeds.ac.uk.
authors.forename: Valentin
authors.initials: V
authors.lastname: Radu
authors.forename: Stefan
authors.initials: S
authors.lastname: Frielingsdorf
authors.forename: Oliver
authors.initials: O
authors.lastname: Lenz
authors.forename: Lars J C
authors.initials: LJ
authors.lastname: Jeuken