DNA replication is a highly accurate process designed to duplicate the entire genome of a cell during each cell division. The accuracy of DNA replication is derived from the balance between three important components: base selectivity by the replicative DNA polymerases (pols), exonucleolytic proofreading, and post-replicative mismatch repair. Previously we identified a human 3′-5′ exonuclease (exoN) whose properties suggested it may function as a proofreader for the exonuclease-deficient replicative DNA pol α. Purified exoN has no associated pol activity and catalyzes removal of mispaired nucleotides from DNA duplexes. Consistent with previous reports, it was found that mammalian pol α is inefficient at extending from mispaired DNA terminals. However, in similar reactions that included exoN, there was a 4.4-15.7-fold increase in pol α-catalyzed elongation from mispaired base pairs. In contrast, exoN did not have a dramatic impact on the ability of exonuclease-deficient variants of Klenow (K-) and T7 polymerase to catalyze extension from mispaired DNA. Continuous DNA replication catalyzed by either pol α or K- generated base substitutions at a frequency of 24.3×10-4and 38×10-4, respectively. ExoN restored error-free DNA replication in reactions with pol α whereas it did not significantly improve the accuracy of K-. These results are consistent with a functional interaction between exoN and pol α to ensure accurate DNA replication. © 2002 Elsevier Science B.V. All rights reserved.
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