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dc.contributor.authorSweeney, Pamela
dc.contributor.authorGalliford, Ashleigh
dc.contributor.authorKumar, Abhishek
dc.contributor.authorRaju, Dinesh
dc.contributor.authorKrishna, Naveen B.
dc.contributor.authorSutherland, Emmajay
dc.contributor.authorLeo, Caitlin J.
dc.contributor.authorFisher, Gemma
dc.contributor.authorLalitha, Roopa
dc.contributor.authorMuthuraj, Likith
dc.contributor.authorSigamani, Gladstone
dc.contributor.authorOehler, Verena
dc.contributor.authorSynowsky, Silvia Anna
dc.contributor.authorShirran, Sally Lorna
dc.contributor.authorGloster, Tracey
dc.contributor.authorMelo Czekster, Clarissa
dc.contributor.authorKumar, Pravin
dc.contributor.authorda Silva, R.G.
dc.date.accessioned2022-06-10T16:30:02Z
dc.date.available2022-06-10T16:30:02Z
dc.date.issued2022-05
dc.identifier.citationSweeney , P , Galliford , A , Kumar , A , Raju , D , Krishna , N B , Sutherland , E , Leo , C J , Fisher , G , Lalitha , R , Muthuraj , L , Sigamani , G , Oehler , V , Synowsky , S A , Shirran , S L , Gloster , T , Melo Czekster , C , Kumar , P & da Silva , R G 2022 , ' Structure, dynamics, and molecular inhibition of the Staphylococcus aureus m 1 A22-tRNA methyltransferase TrmK ' , Journal of Biological Chemistry , vol. 298 , no. 6 , 102040 . https://doi.org/10.1016/j.jbc.2022.102040en
dc.identifier.issn0021-9258
dc.identifier.otherPURE: 279617562
dc.identifier.otherPURE UUID: c09e36f3-ef2c-4c1f-85b3-299c08a8832b
dc.identifier.otherORCID: /0000-0002-7163-4057/work/114335689
dc.identifier.otherORCID: /0000-0003-3516-3507/work/114335856
dc.identifier.otherORCID: /0000-0002-1308-8190/work/114335921
dc.identifier.otherScopus: 85131461501
dc.identifier.otherWOS: 000829588600012
dc.identifier.urihttp://hdl.handle.net/10023/25512
dc.descriptionThis work was supported by a Wellcome Trust Seed Award in Science [208980/Z/17/Z] to RGdS; a University of St Andrews/Scottish Funding Council St Andrews Restarting Research Fund to RGdS; and a Wellcome Trust Institutional Strategic Support Fund [204821/Z/16/Z] to the University of St Andrews. ES is the recipient of a Cunningham Trust PhD studentship (PhD-CT-18-41).en
dc.description.abstractThe enzyme m1A22-tRNA methyltransferase (TrmK) catalyzes the transfer of a methyl group to the N1 of adenine 22 in bacterial tRNAs. TrmK is essential for Staphylococcus aureus survival during infection but has no homolog in mammals, making it a promising target for antibiotic development. Here, we characterize the structure and function of S. aureus TrmK (SaTrmK) using X-ray crystallography, binding assays, and molecular dynamics simulations. We report crystal structures for the SaTrmK apoenzyme as well as in complexes with methyl donor SAM and co-product product SAH. Isothermal titration calorimetry showed that SAM binds to the enzyme with favorable but modest enthalpic and entropic contributions, whereas SAH binding leads to an entropic penalty compensated for by a large favorable enthalpic contribution. Molecular dynamics simulations point to specific motions of the C-terminal domain being altered by SAM binding, which might have implications for tRNA recruitment. In addition, activity assays for SaTrmK-catalyzed methylation of A22 mutants of tRNALeu demonstrate that the adenine at position 22 is absolutely essential. In silico screening of compounds suggested the multifunctional organic toxin plumbagin as a potential inhibitor of TrmK, which was confirmed by activity measurements. Furthermore, LC-MS data indicated the protein was covalently modified by one equivalent of the inhibitor, and proteolytic digestion coupled with LC-MS identified Cys92 in the vicinity of the SAM-binding site as the sole residue modified. These results identify a cryptic binding pocket of SaTrmK, laying a foundation for future structure-based drug discovery.
dc.format.extent15
dc.language.isoeng
dc.relation.ispartofJournal of Biological Chemistryen
dc.rightsCopyright © 2022 The Authors. Published by Elsevier Inc on behalf of American Society for Biochemistry and Molecular Biology. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).en
dc.subjectRNA methyltransferaseen
dc.subjectTransfer RNA (tRNA)en
dc.subjectS-adenosylmethionine (SAM)en
dc.subjectStaphylococcus aureusen
dc.subjectX-ray crystallographyen
dc.subjectTrmKen
dc.subjectQR Microbiologyen
dc.subjectRM Therapeutics. Pharmacologyen
dc.subjectDASen
dc.subject.lccQRen
dc.subject.lccRMen
dc.titleStructure, dynamics, and molecular inhibition of the Staphylococcus aureus m1A22-tRNA methyltransferase TrmKen
dc.typeJournal articleen
dc.contributor.sponsorThe Wellcome Trusten
dc.contributor.sponsorThe Wellcome Trusten
dc.description.versionPublisher PDFen
dc.contributor.institutionUniversity of St Andrews. School of Biologyen
dc.contributor.institutionUniversity of St Andrews. Institute of Behavioural and Neural Sciencesen
dc.contributor.institutionUniversity of St Andrews. School of Chemistryen
dc.contributor.institutionUniversity of St Andrews. Biomedical Sciences Research Complexen
dc.identifier.doihttps://doi.org/10.1016/j.jbc.2022.102040
dc.description.statusPeer revieweden
dc.identifier.urlhttp://10.1101/2021.12.24.474102en
dc.identifier.grantnumber208980/Z/17/Zen
dc.identifier.grantnumber204821/Z/16/Zen


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