SPECIFIC BINDING OF AMINOGLYCOSIDE ANTIBIOTICS TO RNA

Background: Aminoglycoside antibiotics interfere with ribosomal protei n synthesis and with intron splicing. Various lines of evidence sugges t that RNA is the molecular target for aminoglycosides, but little is known about the recognition process. Is recognition of a particular am inoglycoside specific for certain RNA structures? If so, what are the rules for recognition? We have begun to investigate this problem by in vitro selection of RNA molecules that can specifically bind to the am inoglycoside antibiotic tobramycin. Results: An RNA diversity library was used to select for sequences capable of binding to the aminoglycos ide antibiotic tobramycin. After six cycles of selection, 82 % of the RNA bound to tobramycin specifically. The selected RNA was reverse-tra nscribed into DNA, which was then cloned. At low selection stringency, an extremely large number of clones, on the order of 10(7), produced RNAs capable of binding tobramycin with K(d)s in the mu M range (value s similar to that observed for the binding of tobramycin to Escherichi a coli ribosomes). Sequencing of 18 of the clones revealed no obvious consensus sequence. At higher selection stringencies (K(d)s in the nM range) only two consensus sequences for binding were observed. Conclus ions: We have shown that RNA molecules can be readily selected that bi nd the aminoglycoside tobramycin. The RNAs that bind tobramycin with h igh affinity contain consensus binding regions that may be confined to predicted stern-loop structures. These studies open the way for under standing the basis of RNA-aminoglycoside recognition.