title: The excision of 3' penultimate errors by DNA polymerase I and its role in endonuclease V-mediated DNA repair.
abstract: Deamination of adenine can occur spontaneously under physiological conditions, and is enhanced by exposure of DNA to ionizing radiation, UV light, nitrous acid, or heat, generating the highly mutagenic lesion of deoxyinosine in DNA. Such DNA lesions tends to generate A:T to G:C transition mutations if unrepaired. In Escherichia coli, deoxyinosine is primarily removed through a repair pathway initiated by endonuclease V (endo V). In this study, we compared the repair of three mutagenic deoxyinosine lesions of A-I, G-I, and T-I using E. coli cell-free extracts as well as reconstituted protein system. We found that 3'-5' exonuclease activity of DNA polymerase I (pol I) was very important for processing all deoxyinosine lesions. To understand the nature of pol I in removing damaged nucleotides, we systemically analyzed its proofreading to 12 possible mismatches 3'-penultimate of a nick, a configuration that represents a repair intermediate generated by endo V. The results showed all mismatches as well as deoxyinosine at the 3' penultimate site were corrected with similar efficiency. This study strongly supports for the idea that the 3'-5' exonuclease activity of E. coli pol I is the primary exonuclease activity for removing 3'-penultimate deoxyinosines derived from endo V nicking reaction.
authors.affiliation: Department of Clinical Laboratory Sciences and Medical Biotechnology, College of Medicine, National Taiwan University, Taipei, 100-02, Taiwan, ROC.
authors.forename: Chia-Chia
authors.initials: CC
authors.lastname: Lee
authors.forename: Ya-Chien
authors.initials: YC
authors.lastname: Yang
authors.forename: Steven D
authors.initials: SD
authors.lastname: Goodman
authors.forename: Chien-Ju
authors.initials: CJ
authors.lastname: Lin
authors.forename: Yi-An
authors.initials: YA
authors.lastname: Chen
authors.forename: Yi-Ting
authors.initials: YT
authors.lastname: Wang
authors.forename: Wern-Cherng
authors.initials: WC
authors.lastname: Cheng
authors.forename: Liang-In
authors.initials: LI
authors.lastname: Lin
authors.forename: Woei-Horng
authors.initials: WH
authors.lastname: Fang
mesh_terms: D053904:DNA Breaks, Single-Stranded; D011088:DNA Ligases / Q000378:metabolism; D053843:DNA Mismatch Repair; D004256:DNA Polymerase I / Q000378:metabolism*; D003641:Deamination; D043224:Deoxyribonuclease (Pyrimidine Dimer) / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D007288:Inosine / Q000031:analogs & derivatives / Q000378:metabolism; D009154:Mutation; D013379:Substrate Specificity
chemical_list: D029968:Escherichia coli Proteins; D007288:Inosine; D004256:DNA Polymerase I; D043224:Deoxyribonuclease (Pyrimidine Dimer); D011088:DNA Ligases; C012271:deoxyinosine
keywords: 3′ Penultimate site; 3′-5′ Exonuclease; DNA mismatch; DNA polymerase I; Deoxyinosine; Endonuclease V