In vitro bypass of UV-induced lesions by Escherichia coli DNA polymerase I: specificity of nucleotide incorporation.

Publisher Website
Google Scholar
Abstract
A variety of DNA polymerases, synthesizing in vitro on an UV-irradiated .vphi.X174 DNA template, terminate synthesis one nucleotide before the 3'' pyrimidines of putative dimers on the template. A system using E. coli DNA polymerase I (Klenow fragment) that can synthesize past at least some of these dimers was devised. The bypass is carried out in a multistep process: first, the incorporation of nucleotides opposite the pyrimidines in the dimer and, then, the addition of nucleotides complementary to the bases distal to the dimer. The insertion of a nucleotide opposite the first (3'') pyrimidine of a putative dimer in the presence of Mn2+ occurs in a concentration-dependent fashion with a 3- to 4-fold preference for purine nucleotides over pyrimidine nucleotides. In the presence of Mg2+, insertion is less frequent. Correlation of these results with in vivo mutation data suggests a role for the polymerase in determining the spectrum of base substitution mutagenesis in SOS induced cells.