Hammerhead Ribozymes with a Faster Cleavage Rate

Abstract

A hammerhead ribozyme that was previously reported to have a rate of chemical cleavage 10-fold faster than that of conventional hammerheads was analyzed in greater detail. Although originally found as a bimolecular hammerhead assembled through helices I and II, fast cleavage was observed in hammerheads in the more conventional helix I−helix III form, provided the sequence of helix I of the fast hammerhead was preserved. Mutations indicated that the fast rate of cleavage was due to the presence of both the U1.1-A2.1 and A1.2-U2.2 base pairs. The faster rate of cleavage was due to a small increase in the activation entropy of the reaction. In addition, we confirmed previous reports that increasing the length of helix I by greater than five base pairs inhibits cleavage slightly and have uncovered a similar effect in helix II.