Figure 5.

Effect of functional group changes at position 638 on the rate of VS ribozyme cleavage. Variant substrates were synthesized with alternative nucleotide bases or analogs at position 638 as shown and radioactively 5′-³²P labelled. These were subjected to cleavage in trans by VS ribozyme under single-turnover conditions for 30 min at 37°C in standard buffer. Substrate and product were separated by electrophoresis in 20% polyacrylamide gels under denaturing conditions, and visualized by phosphorimaging. Tracks 1, natural substrate before cleavage; tracks 2–8, incubation with VS ribozyme; track 2, natural substrate; track 3, substitution by adenine; track 4, substitution by inosine; track 5, substitution by 2-aminopurine; track 6, substitution by 2,6-diaminopurine and track 7, substitution by purine.