Catalytic effects of mutations of distant protein residues in human DNA polymerase β: theory and experiment.

ABSTRACT: We carried out free-energy calculations and transient kinetic experiments for the insertion of the right (dC) and wrong (dA) nucleotides by wild-type (WT) and six mutant variants of human DNA polymerase β (Pol β). Since the mutated residues in the point mutants, I174S, I260Q, M282L, H285D, E288K, and K289M, were not located in the Pol β catalytic site, we assumed that the WT and its point mutants share the same dianionic phosphorane transition-state structure of the triphosphate moiety of deoxyribonucleotide 5'-triphosphate (dNTP) substrate. On the basis of this assumption, we have formulated a thermodynamic cycle for calculating relative dNTP insertion efficiencies, Ω = (k(pol)/K(D))(mut)/(k(pol)/K(D))(WT) using free-energy perturbation (FEP) and linear interaction energy (LIE) methods. K

SUBMITTER: Klvana M 

PROVIDER: S-EPMC3982232 | biostudies-literature | 2012 Nov

REPOSITORIES: biostudies-literature