Hairpin ribozyme cleavage catalyzed by aminoglycoside antibiotics and the polyamine spermine in the absence of metal ions

The hairpin ribozyme is a small catalytic RNA that achieves an active confi guration by docking of its two helical domains in an antiparallel fashion. Both docking and subsequent cleavage are dependent on the presence of dival ent metal ions, such as magnesium, but there is no evidence to date for dir ect participation of such ions in the chemical cleavage step. We show that aminoglycoside antibiotics inhibit cleavage of the hairpin ribozyme in the presence of metal ions with the most effective being 5-epi-sisomicin and ne omycin B, In contrast, in the absence of metal ions, a number of aminoglyco side antibiotics at 10 mM concentration promote hairpin cleavage with rates only 13-20-fold lower than the magnesium-dependent reaction. We show that neomycin B competes with metal ions by ion replacement with the postively c harged amino groups of the antibiotic. In addition, we show that the polyam ine spermine at 10 mM promotes efficient hairpin cleavage with rates simila r to the magnesium-dependent reaction. Low concentrations of either spermin e or the shorter polyamine spermidine synergize with 5 mM magnesium ions to boost cleavage rates considerably, In contrast, at 500 mu M magnesium ions , 4 mM spermine, but not spermidine, boosts the cleavage rate. The results have significance both in understanding the role of ions in hairpin ribozym e cleavage and in potential therapeutic applications in mammalian cells.