The role of DNA polymerase in base substitution mutagenesis on non-instructional templates

Abstract

In vitro DNA synthesis on phi X174 or M13 templates with non-instructional lesions such as UV dimers or AP (apurinic/apyrimidinic) sites terminates one base before the site of the lesion when synthesis is catalyzed by T4 DNA polymerase or E. coli polymerase I. E. Coli polymerase I also produces termination bands at the site of AP lesions. Substitution of Mn2+ for Mg2+ and increasing the concentration of dNTP's results in elongation of the newly synthesized strand opposite the site of the lesion and beyond. Purine deoxynucleoside triphosphates are utilized for insertion opposite lesions to a greater extent than are pyrimidine deoxynucleoside triphosphates. Deoxy ATP is used almost exclusively for elongation opposite AP sites with pol I-Klenow fragment in the presence of Mg2+. We suppose that these results illustrate the previously observed greater affinity of polymerases under template-free conditions for purine nucleotides. We also suppose that the results can be used to account for mutagenic base selection on noninstructional DNA templates. If purines are preferentially selected by polymerases, then treatments which inactivate pyrimidines will lead to an excess of transitions whereas inactivation of purines will produce more transversions. Data in the literature support this hypothesis.