Please use this identifier to cite or link to this item: http://hdl.handle.net/2381/27892
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dc.contributor.authorUthman, Shanow-
dc.contributor.authorBär, Christian-
dc.contributor.authorScheidt, Viktor-
dc.contributor.authorLiu, Shihui-
dc.contributor.authorten Have, Sara-
dc.contributor.authorGiorgini, Flaviano-
dc.contributor.authorStark, Michael J.R.-
dc.contributor.authorSchaffrath, Raffael-
dc.date.accessioned2013-05-08T15:00:13Z-
dc.date.available2013-05-08T15:00:13Z-
dc.date.issued2013-02-28-
dc.identifier.citationPLoS Genetics, 2013, 9 (2), e1003334.en
dc.identifier.issn1553-7390-
dc.identifier.urihttp://www.plosgenetics.org/article/info%3Adoi%2F10.1371%2Fjournal.pgen.1003334en
dc.identifier.urihttp://hdl.handle.net/2381/27892-
dc.description.abstractDiphthamide is a highly modified histidine residue in eukaryal translation elongation factor 2 (eEF2) that is the target for irreversible ADP ribosylation by diphtheria toxin (DT). In Saccharomyces cerevisiae, the initial steps of diphthamide biosynthesis are well characterized and require the DPH1-DPH5 genes. However, the last pathway step-amidation of the intermediate diphthine to diphthamide-is ill-defined. Here we mine the genetic interaction landscapes of DPH1-DPH5 to identify a candidate gene for the elusive amidase (YLR143w/DPH6) and confirm involvement of a second gene (YBR246w/DPH7) in the amidation step. Like dph1-dph5, dph6 and dph7 mutants maintain eEF2 forms that evade inhibition by DT and sordarin, a diphthamide-dependent antifungal. Moreover, mass spectrometry shows that dph6 and dph7 mutants specifically accumulate diphthine-modified eEF2, demonstrating failure to complete the final amidation step. Consistent with an expected requirement for ATP in diphthine amidation, Dph6 contains an essential adenine nucleotide hydrolase domain and binds to eEF2. Dph6 is therefore a candidate for the elusive amidase, while Dph7 apparently couples diphthine synthase (Dph5) to diphthine amidation. The latter conclusion is based on our observation that dph7 mutants show drastically upregulated interaction between Dph5 and eEF2, indicating that their association is kept in check by Dph7. Physiologically, completion of diphthamide synthesis is required for optimal translational accuracy and cell growth, as indicated by shared traits among the dph mutants including increased ribosomal -1 frameshifting and altered responses to translation inhibitors. Through identification of Dph6 and Dph7 as components required for the amidation step of the diphthamide pathway, our work paves the way for a detailed mechanistic understanding of diphthamide formation.en
dc.language.isoenen
dc.publisherPublic Library of Scienceen
dc.rightsThis is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.en
dc.titleThe amidation step of diphthamide biosynthesis in yeast requires DPH6, a gene identified through mining the DPH1-DPH5 interaction network.en
dc.typeJournal Articleen
dc.identifier.doi10.1371/journal.pgen.1003334-
dc.identifier.eissn1553-7404-
dc.identifier.piiPGENETICS-D-12-02403-
dc.description.statusPeer-revieweden
dc.description.versionPublisher Versionen
dc.type.subtypeJournal Article;Research Support, N.I.H., Intramural;Research Support, Non-U.S. Gov't-
pubs.organisational-group/Organisationen
pubs.organisational-group/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGYen
pubs.organisational-group/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Biological Sciencesen
pubs.organisational-group/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Biological Sciences/Department of Geneticsen
pubs.organisational-group/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/Themesen
pubs.organisational-group/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/Themes/Genome Scienceen
pubs.organisational-group/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/Themes/Neuroscience & Behaviouren
Appears in Collections:Published Articles, Dept. of Genetics

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