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On the fidelity of DNA replication. Nucleoside monophosphate generation during polymerization.

Authors :
Loeb LA
Dube DK
Beckman RA
Koplitz M
Gopinathan KP
Source :
The Journal of biological chemistry [J Biol Chem] 1981 Apr 25; Vol. 256 (8), pp. 3978-87.
Publication Year :
1981

Abstract

During catalysis by homogeneous procaryotic DNA polymerases, nucleoside monophosphates are generated by a 3' leads to 5'-exonucleolytic activity. Using Escherichia coli DNA polymerase I and poly[d(A-T)] as a template, the contribution of this activity to the fidelity of DNA synthesis has been evaluated by three different criteria. 1) The ratio between the rates of monophosphate generation and incorporation of the noncomplementary nucleotide with Mg2+ as an activating cation was 0.6 +/- 0.6, which is insufficient to account for the high fidelity of polymerization. 2) Inhibition of polymerization by pyrophosphate fails to diminish fidelity, although some kinetic models suggest that optimal error correction via monophosphate release requires the polymerization reaction to be strongly driven by pyrophosphate release. 3) The addition of deoxynucleoside monophosphates in concentrations as great as 10 mM to the reaction mixture does not alter the fidelity of DNA synthesis. These observations argue against the kinetic proofreading mode to account for the fidelity of E. coli DNA polymerase I when copying poly[d(A-T)] in a Mg2+-activated reaction. Furthermore, they suggest that the polymerase may enhance specificity at the base-selection step. However, the 3' leads to 5' exonuclease plays a larger role when the polymerase is activated with Mn2+ and may also be important in copying natural DNA where lower error rates are observed in vitro.

Details

Language :
English
ISSN :
0021-9258
Volume :
256
Issue :
8
Database :
MEDLINE
Journal :
The Journal of biological chemistry
Publication Type :
Academic Journal
Accession number :
7012147