[Complex of repair DNA polymerase beta with autonomous 3'-->5'-exonuclease shows increased accuracy of DNA synthesis]

Izv Akad Nauk Ser Biol. 2007 Sep-Oct;(5):517-23.
[Article in Russian]

Abstract

The complexes of repair DNA polymerase beta with 3'-exonuclease and some other proteins were isolated from the chromatin of hepatocytes of normal rats for the first time. Biopolymers were extracted from the chromatin by the solution of NaCl and Triton X-100. The extract was fractionated by gel-filtration on Sephacryl S-300 columns successively in low and high ionic strength solutions, on hydroxyapatite, and on Sephadex G-100 columns. The complexes have molecular weights of 100 and 300 kDa. They dissociate to DNA polymerase and exonuclease in the course of chromatography on a DNA-cellulose column or after gel-filtration in the presence of 1 M NaCl. The co-purification of the polymerase and exonuclease is reconstituted in 0.1 M NaCl. The fidelity of monomeric and composite DNA polymerase beta was measured using phage phiX174 amber 3 as a primer/template. The products of the synthesis were transfected into Escherichia coli spheroplasts, and the frequency of reverse mutations was determined. The complex of DNA polymerase beta with 3'-exonuclease was shown to be 30 times more accurate than the monomeric polymerase, which can decrease the probability of repair mutagenesis and carcinogenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bacteriophage phi X 174 / chemistry*
  • Bacteriophage phi X 174 / genetics
  • DNA Polymerase beta / chemistry*
  • DNA Polymerase beta / isolation & purification
  • DNA, Viral / chemistry*
  • DNA, Viral / genetics
  • Escherichia coli / genetics
  • Exodeoxyribonucleases / chemistry*
  • Exodeoxyribonucleases / isolation & purification
  • Hepatocytes / enzymology*
  • Mutation
  • Rats
  • Spheroplasts / genetics

Substances

  • DNA, Viral
  • DNA Polymerase beta
  • Exodeoxyribonucleases