Elsevier

DNA Repair

Volume 83, November 2019, 102643
DNA Repair

Replication fidelity in E. coli: Differential leading and lagging strand effects for dnaE antimutator alleles

https://doi.org/10.1016/j.dnarep.2019.102643Get rights and content
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Highlights

  • Pol III antimutator variants have higher fidelity than the wild-type enzyme.

  • Antimutators increase the replication fidelity preferentially on the leading strand.

  • dnaE antimutators promote fidelity by increasing “dissociability”.

  • When antimutators are faced with error-prone polymerases they turn into mutators.

  • Increased 'dissociability' of the Pol III serves as an important fidelity factor.

Abstract

DNA Pol III holoenzyme (HE) is the major DNA replicase of Escherichia coli. It is a highly accurate enzyme responsible for simultaneously replicating the leading- and lagging DNA strands. Interestingly, the fidelity of replication for the two DNA strands is unequal, with a higher accuracy for lagging-strand replication. We have previously proposed this higher lagging-strand fidelity results from the more dissociative character of the lagging-strand polymerase. In support of this hypothesis, an E. coli mutant carrying a catalytic DNA polymerase subunit (DnaE915) characterized by decreased processivity yielded an antimutator phenotype (higher fidelity). The present work was undertaken to gain deeper insight into the factors that influence the fidelity of chromosomal DNA replication in E. coli. We used three different dnaE alleles (dnaE915, dnaE911, and dnaE941) that had previously been isolated as antimutators. We confirmed that each of the three dnaE alleles produced significant antimutator effects, but in addition showed that these antimutator effects proved largest for the normally less accurate leading strand. Additionally, in the presence of error-prone DNA polymerases, each of the three dnaE antimutator strains turned into mutators. The combined observations are fully supportive of our model in which the dissociative character of the DNA polymerase is an important determinant of in vivo replication fidelity. In this model, increased dissociation from terminal mismatches (i.e., potential mutations) leads to removal of the mismatches (antimutator effect), but in the presence of error-prone (or translesion) DNA polymerases the abandoned terminal mismatches become targets for error-prone extension (mutator effect). We also propose that these dnaE alleles are promising tools for studying polymerase exchanges at the replication fork.

Abbreviations

Pol
DNA polymerase
DNA Pol III HE
DNA Pol III holoenzyme

Keywords

DNA Pol III HE
Antimutators
Leading and lagging strand
Replication fidelity
DnaE915

Cited by (0)

1

Present Address: Cancer Research Center of Marseille, CNRS, UMR7258; Inserm, U1068; Institut Paoli-Calmettes; Aix-Marseille University, Marseille, France.

2

Present Address: Department of Biochemistry and Molecular Biology, Centre of Postgraduate Medical Education, Warsaw, Poland.