title: Effect of dNTP pool alterations on fidelity of leading and lagging strand DNA replication in E. coli.
abstract: The fidelity with which organisms replicate their chromosomal DNA is of considerable interest. Detailed studies in the bacterium Escherichia coli have indicated that the fidelity of leading- and lagging-strand DNA replication is not the same, based on experiments in which the orientation of certain mutational targets on the chromosome was inverted relative to the movement of the replication fork: different mutation rates for several base-pair substitutions were observed depending on this orientation. While these experiments are indicative of differential replication fidelity in the two strands, a conclusion whether leading or lagging strand is the more accurate depends on knowledge of the primary mispairing error responsible for the base substitutions in question. A broad analysis of in vitro base-pairing preferences of DNA polymerases led us to propose that lagging-strand is the more accurate strand. In the present work, we present more direct in vivo evidence in support of this proposal. We determine the orientation dependence of mutant frequencies in ndk and dcd strains, which carry defined dNTP pool alterations. As these pool alterations lead to predictable effects on the array of possible mispairing errors, they mark the strands in which the observed errors occur. The combined results support the proposed higher accuracy of lagging-strand replication in E. coli.
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw 02-106, Poland; Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, United States. Electronic address: gawel@cmkp.edu.pl.
authors.forename: Damian
authors.initials: D
authors.lastname: Gawel
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw 02-106, Poland.
authors.forename: Iwona J
authors.initials: IJ
authors.lastname: Fijalkowska
authors.affiliation: Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw 02-106, Poland.
authors.forename: Piotr
authors.initials: P
authors.lastname: Jonczyk
authors.affiliation: Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, United States.
authors.forename: Roel M
authors.initials: RM
authors.lastname: Schaaper
mesh_terms: D004261:DNA Replication; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D003854:Deoxyribonucleotides / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D007763:Lac Operon; D013942:Thymine Nucleotides / Q000378:metabolism
chemical_list: D003838:Deoxyadenine Nucleotides; D003845:Deoxycytosine Nucleotides; D003848:Deoxyguanine Nucleotides; D003854:Deoxyribonucleotides; D013942:Thymine Nucleotides; C024107:2'-deoxycytidine 5'-triphosphate; C029603:deoxyguanosine triphosphate; C026600:2'-deoxyadenosine triphosphate; C024157:thymidine 5'-triphosphate
keywords: Base selection; Base·base mispairs; DNA replication fidelity; Exonucleolytic proofreading; Leading and lagging strands; dNTP precursors