Hammerhead ribozyme mechanism: A ribonucleotide 5 ' to the substrate cleavage site is not essential

Three hammerhead ribozymes with triplet specificities for cleavage 3' of CU C, GUC, and GUA have been evaluated for their sensitivity to the substituti on of thymidine or 2'-deoxyuridine at central nucleotide position 16.1 in t he substrate triplet. All three ribozymes cleaved their respective substrat es, containing uridine or the modifications, with comparable rates. This in dicates that the 2'-hydroxy group at position 16.1 is not essential for act ivity even though X-ray structure analysis shows it participates in H-bondi ng interactions. These H-bonds were considered to be of functional signific ance because an earlier report had provided data that thymidine at position 16.1 is deleterious for catalytic activity [Yang, J.-H., Perreault, J.-P., Labuda, D., Usman, N., and Cedergren, R. (1990) Biochemistry 29, 11156-111 60].