title: Role of high-fidelity Escherichia coli DNA polymerase I in replication bypass of a deoxyadenosine DNA-peptide cross-link.
abstract: Reaction of bifunctional electrophiles with DNA in the presence of peptides can result in DNA-peptide cross-links. In particular, the linkage can be formed in the major groove of DNA via the exocyclic amino group of adenine (N⁶-dA). We previously demonstrated that an A family human polymerase, Pol ν, can efficiently and accurately synthesize DNA past N⁶-dA-linked peptides. Based on these results, we hypothesized that another member of that family, Escherichia coli polymerase I (Pol I), may also be able to bypass these large major groove DNA lesions. To test this, oligodeoxynucleotides containing a site-specific N⁶-dA dodecylpeptide cross-link were created and utilized for in vitro DNA replication assays using E. coli DNA polymerases. The results showed that Pol I and Pol II could efficiently and accurately bypass this adduct, while Pol III replicase, Pol IV, and Pol V were strongly inhibited. In addition, cellular studies were conducted using E. coli strains that were either wild type or deficient in all three DNA damage-inducible polymerases, i.e., Pol II, Pol IV, and Pol V. When single-stranded DNA vectors containing a site-specific N⁶-dA dodecylpeptide cross-link were replicated in these strains, the efficiencies of replication were comparable, and in both strains, intracellular bypass of the lesion occurred in an error-free manner. Collectively, these findings demonstrate that despite its constrained active site, Pol I can catalyze DNA synthesis past N⁶-dA-linked peptide cross-links and is likely to play an essential role in cellular bypass of large major groove DNA lesions.
authors.affiliation: Center for Research on Occupational and Environmental Toxicology, Oregon Health & Science University, Portland, OR 97239, USA.
authors.forename: Kinrin
authors.initials: K
authors.lastname: Yamanaka
authors.forename: Irina G
authors.initials: IG
authors.lastname: Minko
authors.forename: Steven E
authors.initials: SE
authors.lastname: Finkel
authors.forename: Myron F
authors.initials: MF
authors.lastname: Goodman
authors.forename: R Stephen
authors.initials: RS
authors.lastname: Lloyd
mesh_terms: D003432:Cross-Linking Reagents / Q000737:chemistry; D018736:DNA Adducts / Q000737:chemistry / Q000235:genetics; D004249:DNA Damage; D004256:DNA Polymerase I / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D004261:DNA Replication; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics; D003839:Deoxyadenosines / Q000737:chemistry / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry*
chemical_list: D003432:Cross-Linking Reagents; D018736:DNA Adducts; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D003839:Deoxyadenosines; D029968:Escherichia coli Proteins; D010455:Peptides; D004256:DNA Polymerase I