Resonance Raman spectroscopic analysis of the iron–sulfur cluster redox chain of the Ralstonia eutropha membrane‐bound [NiFe]‐hydrogenase

dc.contributor.authorSiebert, Elisabeth
dc.contributor.authorSchmidt, Andrea
dc.contributor.authorFrielingsdorf, Stefan
dc.contributor.authorKalms, Jacqueline
dc.contributor.authorKuhlmann, Uwe
dc.contributor.authorLenz, Oliver
dc.contributor.authorScheerer, Patrick
dc.contributor.authorZebger, Ingo
dc.contributor.authorHildebrandt, Peter
dc.date.accessioned2022-04-01T08:24:34Z
dc.date.available2022-04-01T08:24:34Z
dc.date.issued2021-06-03
dc.date.updated2022-03-21T14:39:08Z
dc.description.abstractIron–sulfur (Fe–S) centers are versatile building blocks in biological electron transfer chains because their redox potentials may cover a wide potential range depending on the type of the cluster and the specific protein environment. Resonance Raman (RR) spectroscopy is widely used to analyze structural properties of such cofactors, but it remains still a challenge to disentangle the overlapping signals of metalloproteins carrying several Fe–S centers. In this work, we combined RR spectroscopy with protein engineering and X‐ray crystallography to address this issue on the basis of the oxygen‐tolerant membrane‐bound hydrogenase from Ralstonia eutropha that catalyzes the reversible conversion of hydrogen into protons and electrons. Besides the NiFe‐active site, this enzyme harbors three different Fe–S clusters constituting an electron relay with a distal [4Fe–4S], a medial [3Fe–4S], and an unusual proximal [4Fe–3S] cluster that may carry a hydroxyl ligand in the superoxidized state. RR spectra were measured from protein crystals by varying the crystal orientation with respect to the electric field vector of the incident laser to achieve a preferential RR enhancement for individual Fe–S clusters. In addition to spectral discrimination by selective reduction of the proximal cluster, protein engineering allowed for transforming the proximal and medial cluster into standard cubane‐type [4Fe–4S] centers in the C19G/C120G and P242C variants, respectively. The latter variant was structurally characterized for the first time in this work. Altogether, the entirety of the RR data provided the basis for identifying the vibrational modes characteristic of the various cluster states in this “model” enzyme as a prerequisite for future studies of complex (FeS)‐based electron transfer chains.en
dc.description.abstracten
dc.description.sponsorshipDFG, 390540038, EXC 2008: Unifying Systems in Catalysis "UniSysCat"en
dc.description.sponsorshipEC/H2020/810856/EU/Twin to Illuminate Metals in Biology and Biocatalysis through Biospectroscopy/TIMB3en
dc.identifier.eissn1097-4555
dc.identifier.issn0377-0486
dc.identifier.urihttps://depositonce.tu-berlin.de/handle/11303/16648
dc.identifier.urihttp://dx.doi.org/10.14279/depositonce-15425
dc.language.isoenen
dc.rightsThis is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
dc.rights
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/en
dc.subject.ddc540 Chemie und zugeordnete Wissenschaftende
dc.subject.otherelectron transferen
dc.subject.otherhydrogenaseen
dc.subject.otheriron–sulfur clusteren
dc.subject.otherprotein crystalsen
dc.subject.otherRaman spectroscopyen
dc.titleResonance Raman spectroscopic analysis of the iron–sulfur cluster redox chain of the Ralstonia eutropha membrane‐bound [NiFe]‐hydrogenaseen
dc.typeArticleen
dc.type.versionpublishedVersionen
dcterms.bibliographicCitation.doi10.1002/jrs.6163en
dcterms.bibliographicCitation.issue12en
dcterms.bibliographicCitation.journaltitleJournal of Raman Spectroscopyen
dcterms.bibliographicCitation.originalpublishernameWileyen
dcterms.bibliographicCitation.originalpublisherplaceNew York, NYen
dcterms.bibliographicCitation.pageend2632en
dcterms.bibliographicCitation.pagestart2621en
dcterms.bibliographicCitation.volume52en
tub.accessrights.dnbfreeen
tub.affiliationFak. 2 Mathematik und Naturwissenschaften>Inst. Chemie>FG Physikalische Chemie / Biophysikalische Chemiede
tub.affiliation.facultyFak. 2 Mathematik und Naturwissenschaftende
tub.affiliation.groupFG Physikalische Chemie / Biophysikalische Chemiede
tub.affiliation.instituteInst. Chemiede
tub.publisher.universityorinstitutionTechnische Universität Berlinen
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