The effect of base mismatches in the substrate recognition helices of hammerhead ribozymes on binding and catalysis

Abstract

The ability of the hammerhead ribozyme to distinguish between matched and mismatched substrates was evaluated using two kinetically defined ribozymes that differed in the length and sequence of the substrate recognition helices. A mismatch in the innermost base pair of helix I affected k2, the chemical cleavage step, while more distal mismatches had no such effect. In contrast, mismatches in any of the four innermost base pairs of helix III affected k2. Chase experiments indicated that mismatches also increased the rate of substrate dissociation by at least 20-100-fold, as expected from the stabilities of RNA helices.