{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,19]],"date-time":"2026-02-19T00:53:40Z","timestamp":1771462420963,"version":"3.50.1"},"reference-count":37,"publisher":"American Society for Microbiology","issue":"5","license":[{"start":{"date-parts":[[2017,5,1]],"date-time":"2017-05-01T00:00:00Z","timestamp":1493596800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"funder":[{"DOI":"10.13039\/100000060","name":"HHS | NIH | National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01-AI088025"],"award-info":[{"award-number":["R01-AI088025"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Antimicrob Agents Chemother"],"published-print":{"date-parts":[[2017,5]]},"abstract":"<jats:title>ABSTRACT<\/jats:title><jats:p>The pathogen-associated 16S rRNA methyltransferase NpmA catalyzes m<jats:sup>1<\/jats:sup>A1408 modification to block the action of structurally diverse aminoglycoside antibiotics. Here, we describe the development of a fluorescence polarization binding assay and its use, together with complementary functional assays, to dissect the mechanism of NpmA substrate recognition. These studies reveal that electrostatic interactions made by the NpmA \u03b22\/3 linker collectively are critical for docking of NpmA on a conserved 16S rRNA tertiary surface. In contrast, other NpmA regions (\u03b25\/\u03b26 and \u03b26\/\u03b27 linkers) contain several residues critical for optimal positioning of A1408 but are largely dispensable for 30S binding. Our data support a model for NpmA action in which 30S binding and adoption of a catalytically competent state are distinct: docking on 16S rRNA via the \u03b22\/3 linker necessarily precedes functionally critical 30S substrate-driven conformational changes elsewhere in NpmA. This model is also consistent with catalysis being completely positional in nature, as the most significant effects on activity arise from changes that impact binding or stabilization of the flipped A1408 conformation. Our results provide a molecular framework for aminoglycoside resistance methyltransferase action that may serve as a functional paradigm for related enzymes and a starting point for development of inhibitors of these resistance determinants.<\/jats:p>","DOI":"10.1128\/aac.00077-17","type":"journal-article","created":{"date-parts":[[2017,3,14]],"date-time":"2017-03-14T00:42:27Z","timestamp":1489452147000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":7,"title":["Substrate Recognition and Modification by a Pathogen-Associated Aminoglycoside Resistance 16S rRNA Methyltransferase"],"prefix":"10.1128","volume":"61","author":[{"given":"Kellie","family":"Vinal","sequence":"first","affiliation":[{"name":"Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia, USA"},{"name":"Graduate Program in Microbiology and Molecular Genetics, Graduate Division of Biological and Biomedical Sciences, Emory University, Atlanta, Georgia, USA"},{"name":"Emory Antibiotic Resistance Center, Emory University School of Medicine, Atlanta, Georgia, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2419-380X","authenticated-orcid":false,"given":"Graeme L.","family":"Conn","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia, USA"},{"name":"Emory Antibiotic Resistance Center, Emory University School of Medicine, Atlanta, Georgia, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"235","reference":[{"key":"e_1_3_3_2_2","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.00926-07"},{"key":"e_1_3_3_3_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.drup.2012.05.001"},{"key":"e_1_3_3_4_2","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.44.12.3249-3256.2000"},{"key":"e_1_3_3_5_2","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-005-0279-0"},{"key":"e_1_3_3_6_2","doi-asserted-by":"publisher","DOI":"10.1021\/cr0301088"},{"key":"e_1_3_3_7_2","doi-asserted-by":"publisher","DOI":"10.1002\/cbic.200300657"},{"key":"e_1_3_3_8_2","doi-asserted-by":"publisher","DOI":"10.1038\/35030019"},{"key":"e_1_3_3_9_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gki862"},{"key":"e_1_3_3_10_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0969-2126(01)00629-3"},{"key":"e_1_3_3_11_2","doi-asserted-by":"publisher","DOI":"10.1038\/ncomms8896"},{"key":"e_1_3_3_12_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.mi.43.100189.001231"},{"key":"e_1_3_3_13_2","volume-title":"DNA and RNA modification enzymes: structure, mechanism, function, and evolution","author":"Conn GL","year":"2008","unstructured":"Conn GL SavicM MacmasterR. 2008. 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