Fig. 3.

Structural alignment at the 5′-OH leaving group of U1 and modeling of the cleavage site of the hatchet ribozyme. (A) Base stacking with U1 at the cleavage site and alignment of junctional structure. U1 is paired with A19 in stem P1 and stacked above the conserved reversed Watson–Crick G30-C20 pair. G30 and G31 adopt a splayed-apart conformation and are involved in stacking with parallel helixes H12 and H34, respectively. A74′ and A75′ are also splayed apart adjacent to the cleavage site and stacked in H12 and H34, respectively. (B) A surface representation of the hatchet ribozyme product with U1 shown in a stick representation. A cavity is formed within the hatchet ribozyme product adjacent to the leaving group 5′-OH of U1. The dimensions of the cavity appear to be of sufficient size so as to accommodate the cleavage step of the hatchet ribozyme. (C) The base stacking interaction of the modeled cleavage site between C(−1) and U1, in which C(−1) was stacked partially between G32-U62 from stem P3 and two conserved residues G63 and A75′, whereas U1 was stacked between the conserved G30-C20 base pair and the termini of stem P1. (D) A surface representation of the model of the hatchet ribozyme with the cleavage step between C(−1) and U1 shown in stick representation. C(−1) was modeled based on the shape of the cavity on the hatchet ribozyme surface. C(−1) and U1 adopt a splayed-apart conformation. (E) The proposed model of the cleavage site of the hatchet ribozyme, in which C(−1) forms extensive hydrogen bonds with nearby residues. The modeled in-line alignment conformation indicates the potential nucleotides that may contribute to general base and general acid catalysis in the cleavage process.