R964C Mutation of DNA Polymerase γ Imparts Increased Stavudine Toxicity by Decreasing Nucleoside Analog Discrimination and Impairing Polymerase Activity▿

摘要

The R964C mutation of human DNA polymerase γ was recently linked to stavudine (d4T)-mediated mitochondrial toxicity. We utilized pre-steady-state kinetics to determine the effect of this mutation on incorporation of natural substrate dTTP and the active metabolite of d4T (d4TTP). The R964C polymerase γ holoenzyme demonstrated a 33% decrease in dTTP incorporation efficiency and a threefold-lower d4TTP discrimination relative to that of the wild-type polymerase γ, providing a mechanistic basis for genetic predisposition to nucleoside reverse transcriptase inhibitor toxicity.