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DPH1 Gene Mutations Identify a Candidate SAM Pocket in Radical Enzyme Dph1 Dph2 for Diphthamide Synthesis on EF2

dc.date.accessioned2023-12-08T15:17:35Z
dc.date.available2023-12-08T15:17:35Z
dc.date.issued2023-11-16
dc.identifierdoi:10.17170/kobra-202312089194
dc.identifier.urihttp://hdl.handle.net/123456789/15279
dc.description.sponsorshipGefördert durch den Publikationsfonds der Universität Kasselger
dc.language.isoeng
dc.rightsNamensnennung 4.0 International*
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/*
dc.subjectSaccharomyces cerevisiaeeng
dc.subjectSAMeng
dc.subjectradical SAM enzymeseng
dc.subjectEF2 diphthamide modificationeng
dc.subjectDph1•Dph2eng
dc.subjectdiphtheria toxineng
dc.subjectADP ribosylationeng
dc.subject.ddc570
dc.titleDPH1 Gene Mutations Identify a Candidate SAM Pocket in Radical Enzyme Dph1 Dph2 for Diphthamide Synthesis on EF2eng
dc.typeAufsatz
dcterms.abstractIn eukaryotes, the Dph1•Dph2 dimer is a non-canonical radical SAM enzyme. Using iron-sulfur (FeS) clusters, it cleaves the cosubstrate S-adenosyl-methionine (SAM) to form a 3-amino-3-carboxy-propyl (ACP) radical for the synthesis of diphthamide. The latter decorates a histidine residue on elongation factor 2 (EF2) conserved from archaea to yeast and humans and is important for accurate mRNA translation and protein synthesis. Guided by evidence from archaeal orthologues, we searched for a putative SAM-binding pocket in Dph1•Dph2 from Saccharomyces cerevisiae. We predict an SAM-binding pocket near the FeS cluster domain that is conserved across eukaryotes in Dph1 but not Dph2. Site-directed DPH1 mutagenesis and functional characterization through assay diagnostics for the loss of diphthamide reveal that the SAM pocket is essential for synthesis of the décor on EF2 in vivo. Further evidence from structural modeling suggests particularly critical residues close to the methionine moiety of SAM. Presumably, they facilitate a geometry specific for SAM cleavage and ACP radical formation that distinguishes Dph1•Dph2 from classical radical SAM enzymes, which generate canonical 5′-deoxyadenosyl (dAdo) radicals.eng
dcterms.accessRightsopen access
dcterms.creatorSchaffrath, Raffael
dcterms.creatorÜtkür, Koray
dcterms.creatorMayer, Klaus
dcterms.creatorSchmidt, Sarina
dcterms.creatorKlassen, Roland
dcterms.creatorBrinkmann, Ulrich
dcterms.extent13 Seiten
dc.relation.doidoi:10.3390/biom13111655
dc.subject.swdSaccharomyces cerevisiaeger
dc.subject.swdRadikal <Chemie>ger
dc.subject.swdEnzymger
dc.subject.swdDiphtherieger
dc.subject.swdADP-Ribosylierungger
dc.type.versionpublishedVersion
dcterms.source.identifiereissn:2218-273X
dcterms.source.issueIssue 11
dcterms.source.journalBiomoleculeseng
dcterms.source.volumeVolume 13
kup.iskupfalse
dcterms.source.articlenumber1655


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