title: Fidelity of the human mitochondrial DNA polymerase.
abstract: We have quantified the fidelity of polymerization of DNA by human mitochondrial DNA polymerase using synthetic DNA oligonucleotides and recombinant holoenzyme and examining each of the possible 16-base pair combinations. Although the kinetics of incorporation for all correct nucleotides are similar, with an average Kd of 0.8 microM and an average k(pol) of 37 s(-1), the kinetics of misincorporation vary widely. The ground state binding Kd of incorrect bases ranges from a low of 25 microM for a dATP:A mispair to a high of 360 microM for a dCTP:T mispair. Similarly, the rates of incorporation of incorrect bases vary from 0.0031 s(-1) for a dCTP:C mispair to 1.16 s(-1) for a dGTP:T mispair. Due to the variability in the kinetic parameters for misincorporation, the estimates of fidelity range from 1 error in 3563 nucleotides for dGTP:T to 1 error in 2.3 x 10(6) nucleotides for dCTP:C. Interestingly, the discrimination against a dGTP:T mismatch is 16.5 times lower than that of a dTTP:G mismatch due to a tighter Kd for ground state binding and a faster rate of incorporation of the dGTP:T mismatch relative to the dTTP:G mismatch. We calculate an average fidelity of 1 error in 440,000 nucleotides.
authors.affiliation: Department of Chemistry and Biochemistry, Institute for Cellular and Molecular Biology, University of Texas, Austin, Texas 78712, USA.
authors.forename: Harold R
authors.initials: HR
authors.lastname: Lee
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Johnson
mesh_terms: D000249:Adenosine Monophosphate / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D020137:Base Pair Mismatch; D004247:DNA / Q000737:chemistry; D004260:DNA Repair; D004272:DNA, Mitochondrial / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D008928:Mitochondria / Q000201:enzymology*; D009154:Mutation; D009711:Nucleotides / Q000737:chemistry; D013997:Time Factors
chemical_list: D004272:DNA, Mitochondrial; D009711:Nucleotides; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D004247:DNA; D004259:DNA-Directed DNA Polymerase