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 ami noacyl-tRNA site (A site) of 16 S rRNA on the 30 S subunit. Change of a sin gle nucleotide, A1408 to G, reduces the affinity of many aminoglycosides fo r the ribosome; G1408 distinguishes between prokaryotic and eukaryotic ribo somes. The structures of a prokaryotic decoding region A-site oligonucleoti de free in solution and bound to the aminoglycosides paromomycin and gentam icin Cia were determined previously. Here, the structure of a eukaryotic de coding region A-site oligonucleotide bound to paromomycin has been determin ed using NMR spectroscopy and compared to the prokaryotic A-site-paromomyci n structure. A conformational change in three adenosine residues of an inte rnal loop, critical for high-affinity antibiotic binding, was observed in t he prokaryotic RNA-paromomycin complex in comparison to its free form. This conformational change is not observed in the eukaryotic RNA-paromomycin co mplex, disrupting the binding pocket for ring I of the antibiotic. The lack of the conformational change supports footprinting and titration calorimet ry data that demonstrate approximately 25-50-fold weaker binding of paromom ycin to the eukaryotic decoding-site oligonucleotide. Neomycin, which is mu ch less active against Escherichia coli ribosomes with an A1408G mutation, binds non-specifically to the oligonucleotide. These results suggest that e ukaryotic ribosomal RNA has a shallow binding pocket for aminoglycosides, w hich accommodates only certain antibiotics. (C) 2001 Academic Press.