Structural origins of aminoglycoside specificity for prokaryotic ribosomes.
Aminoglycoside antibiotics, including paromomycin, neomycin and gentamicin, target a region of highly conserved nucleotides in the decoding region aminoacyl-tRNA site (A site) of 16 S rRNA on the 30 S subunit. Change of a single nucleotide, A1408 to G, reduces the affinity of many aminoglycosides for the ribosome; G1408 distinguishes between prokaryotic and eukaryotic ribosomes. The structures of a prokaryotic decoding region A-site oligonucleotide free in solution and bound to the aminoglycosides paromomycin and gentamicin C1a were determined previously. Here, the structure of a eukaryotic decoding region A-site oligonucleotide bound to paromomycin has been determined using NMR spectroscopy and compared to the prokaryotic A-site-paromomycin structure. A conformational change in three adenosine residues of an internal loop, critical for high-affinity antibiotic binding, was observed in the prokaryotic RNA-paromomycin complex in comparison to its free form. This conformational change is not observed in the eukaryotic RNA-paromomycin complex, disrupting the binding pocket for ring I of the antibiotic. The lack of the conformational change supports footprinting and titration calorimetry data that demonstrate approximately 25-50-fold weaker binding of paromomycin to the eukaryotic decoding-site oligonucleotide. Neomycin, which is much less active against Escherichia coli ribosomes with an A1408G mutation, binds non-specifically to the oligonucleotide. These results suggest that eukaryotic ribosomal RNA has a shallow binding pocket for aminoglycosides, which accommodates only certain antibiotics.
Department of Structural Biology, Stanford University School of Medicine, Stanford, CA 94305-5126, USA.
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Puglisi
D000241:Adenosine / Q000737:chemistry; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000378:metabolism; D004352:Drug Resistance, Microbial / Q000235:genetics; D006151:Guanosine / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D015394:Molecular Structure; D009355:Neomycin / Q000378:metabolism; D010303:Paromomycin / Q000378:metabolism*; D012329:RNA, Bacterial / Q000737:chemistry*; D012336:RNA, Ribosomal, 16S / Q000737:chemistry* / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry* / Q000378:metabolism; D013045:Species Specificity; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D000900:Anti-Bacterial Agents; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D012346:RNA, Transfer, Amino Acyl; D006151:Guanosine; D010303:Paromomycin; D009355:Neomycin; D000241:Adenosine