tRNAPhe binds aminoglycoside antibiotics Original Research Article

Aminoglycoside antibiotics have recently been found to bind to a variety of unrelated RNA molecules, including sequences that are important for retroviral replication. We report the binding of neomycin B, kanamycin A, and Neo–Neo (a synthetic neomycin–neomycin dimer) to tRNAPhe. Using thermal denaturation studies, fluorescence spectroscopy, Pb2+-mediated tRNAPhe cleavage, and gel mobility shift assays, we have established that aminoglycosides interact with yeast tRNAPhe and are likely to induce a conformational change. Thermal denaturation studies revealed that aminoglycosides have a substantial stabilizing effect on tRNAPhe secondary and tertiary structures, much greater than the stabilization effect of spermine, an unstructured polyamine. Aminoglycoside-induced inhibition of Pb2+-mediated tRNAPhe cleavage yielded IC50 values of: 5 μM for Neo–Neo, 100 μM for neomycin B, >1 mM for kanamycin A, and >10 mM for spermine. Enzymatic and chemical footprinting indicate that the anticodon stem as well as the junction of the TψC and D loops are preferred aminoglycoside binding sites.© Elsevier Science Ltd. All rights reserved.