Molecular recognition and modification of the 30S ribosome by the aminoglycoside-resistance methyltransferase NpmA.
Aminoglycosides are potent, broad spectrum, ribosome-targeting antibacterials whose clinical efficacy is seriously threatened by multiple resistance mechanisms. Here, we report the structural basis for 30S recognition by the novel plasmid-mediated aminoglycoside-resistance rRNA methyltransferase A (NpmA). These studies are supported by biochemical and functional assays that define the molecular features necessary for NpmA to catalyze m(1)A1408 modification and confer resistance. The requirement for the mature 30S as a substrate for NpmA is clearly explained by its recognition of four disparate 16S rRNA helices brought into proximity by 30S assembly. Our structure captures a "precatalytic state" in which multiple structural reorganizations orient functionally critical residues to flip A1408 from helix 44 and position it precisely in a remodeled active site for methylation. Our findings provide a new molecular framework for the activity of aminoglycoside-resistance rRNA methyltransferases that may serve as a functional paradigm for other modification enzymes acting late in 30S biogenesis.
Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322.
Jack A
JA
Dunkle
Kellie
K
Vinal
Pooja M
PM
Desai
Natalia
N
Zelinskaya
Miloje
M
Savic
Dayne M
DM
West
Graeme L
GL
Conn
Christine M
CM
Dunham
D000241:Adenosine / Q000031:analogs & derivatives / Q000737:chemistry / Q000494:pharmacology; D000596:Amino Acids / Q000378:metabolism; D000617:Aminoglycosides / Q000494:pharmacology*; D055162:Biocatalysis / Q000187:drug effects; D020134:Catalytic Domain; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D024881:Drug Resistance, Bacterial / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D022761:Gene Transfer, Horizontal / Q000187:drug effects; D008745:Methylation / Q000187:drug effects; D008780:Methyltransferases / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D009711:Nucleotides / Q000378:metabolism; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012336:RNA, Ribosomal, 16S / Q000737:chemistry; D054680:Ribosome Subunits, Small, Bacterial / Q000737:chemistry* / Q000378:metabolism*; D013329:Structure-Activity Relationship
D000596:Amino Acids; D000617:Aminoglycosides; D029968:Escherichia coli Proteins; D009711:Nucleotides; D012336:RNA, Ribosomal, 16S; D008780:Methyltransferases; C554940:NpmA protein, E coli; D000241:Adenosine; C006235:sinefungin
RNA modification; antibiotic resistance; base flipping