On the fidelity of DNA replication. Lack of primer position effect on the fidelity of mammalian DNA polymerases.

J Abbotts,L A Loeb

doi:10.1016/s0021-9258(17)39786-7

Abstract

Mechanisms for the fidelity of DNA replication in eucaryotes are not adequately understood. Certain hypotheses can be tested by examining whether the first nucleotide inserted is incorporated with a significantly higher error rate than subsequent nucleotides. Using synthetic oligodeoxynucleotides, we have measured the effect of primer position on single-base misinsertion frequencies at an amber site in phi X174 DNA. Our results show a lack of position effect, indicating that processivity and the most direct "energy relay" proofreading mechanisms are not important determinants in eucaryotic replication fidelity.

Highlights

From the Department of Biochemistry and the Joseph Gottstein Memorial Cancer Research Laboratory, Department of Pathology SM-30, University of Washington, Seattle, Washington 98195
Certhayin- additions. potheses can be tested by examining whether tfhierst We have previously developed a system to examine replinucleotide insertedis incorporated with saignificantly cation fidelity using a natural 6x174 am3 DNA template higher error rate than subsequent nucleotides
An unusually high error rate for the first nucleotide incorporated during DNA replication is predicted by mechanisms for fidelity that are mediated by a conformational change in DNA polymerase at each nucleotide addition step

Summary

Introduction

An unusually high error rate for the first nucleotide incorporated during DNA replication is predicted by mechanisms for fidelity that are mediated by a conformational change in DNA polymerase at each nucleotide addition step. We present an analysis of primer position effect on the fidelity of DNA polymerases a and p. Many homogeneous procaryotic DNA polymerases possess a 3’+5’ exonuclease activity [2] which actsto proofread incorrect nucleotide insertions and contributes substantially to replication fidelity; inhibiting this exonuclease activity leads to a 20-500-fold increase in the error rate of a given polymerase [3].

Results

Conclusion