Two proton transfers in the transition state for nucleotidyl transfer catalyzed by RNA- and DNA-dependent RNA and DNA polymerases.

ABSTRACT: The rate-limiting step for nucleotide incorporation in the pre-steady state for most nucleic acid polymerases is thought to be a conformational change. As a result, very little information is available on the role of active-site residues in the chemistry of nucleotidyl transfer. For the poliovirus RNA-dependent RNA polymerase (3D(pol)), chemistry is partially (Mg(2+)) or completely (Mn(2+)) rate limiting. Here we show that nucleotidyl transfer depends on two ionizable groups with pK(a) values of 7.0 or 8.2 and 10.5, depending upon the divalent cation used in the reaction. A solvent deuterium isotope effect of three to seven was observed on the rate constant for nucleotide incorporation in the pre-steady state; none was observed in the steady state. Proton-inventory experiments were consist

SUBMITTER: Castro C 

PROVIDER: S-EPMC1838591 | biostudies-literature | 2007 Mar

REPOSITORIES: biostudies-literature