A guanine nucleobase important for catalysis by the VS ribozyme

Abstract

A guanine (G638) within the substrate loop of the VS ribozyme plays a critical role in the cleavage reaction. Replacement by any other nucleotide results in severe impairment of cleavage, yet folding of the substrate is not perturbed, and the variant substrates bind the ribozyme with similar affinity, acting as competitive inhibitors. Functional group substitution shows that the imino proton on the N1 is critical, suggesting a possible role in general acid-base catalysis, and this in accord with the pH dependence of the reaction rate for the natural and modified substrates. We propose a chemical mechanism for the ribozyme that involves general acid-base catalysis by the combination of the nucleobases of guanine 638 and adenine 756. This is closely similar to the probable mechanism of the hairpin ribozyme, and the active site arrangements for the two ribozymes appear topologically equivalent. This has probably arisen by convergent evolution.