Hammerhead ribozyme kinetics

Abstract

The hammerhead ribozyme is a small RNA motif that self cleaves at a specific phosphodiester bond to produce 2′,3′ cyclic phosphate and 5′ hydroxyl termini (Hutchins et al., 1986; Forster & Symons, 1987a). The secondary structure of the hammerhead consists of three helices of arbitrary sequence and length (designated I, II, and III) that intersect at 15 nucleotides termed the catalytic core (Fig. 1A) (Forster & Symons, 1987b; Hertel et al., 1992). The X-ray crystal structures of two hammerhead ribozyme–inhibitor complexes revealed that the core residues fold into two separate domains and the helices are arranged in a Y-shape conformation with helix I and helix II forming the upper portion of the Y (Pley et al., 1994; Scott et al., 1995). Although the hammerhead is found as an intramolecular motif embedded in several RNAs in vivo (Symons, 1989), it can be assembled from two separate oligonucleotides (Fig. 1B) in three different arrangements (Uhlenbeck, 1987; Haseloff & Gerlach, 1988; Koizumi et al., 1988; Jeffries & Symons, 1989). In these bimolecular formats, the hammerhead effects RNA cleavage in a similar manner to a true “enzyme,” proceeding through multiple rounds of substrate binding, cleavage, and product release (Uhlenbeck, 1987).