title: Chlamydophila pneumoniae endonuclease IV prefers to remove mismatched 3' ribonucleotides: implication in proofreading mismatched 3'-terminal nucleotides in short-patch repair synthesis.
abstract: DNA polymerase I (DNApolI) catalyzes DNA synthesis during Okazaki fragment maturation, base excision repair, and nucleotide excision repair. Some bacterial DNApolIs are deficient in 3'-5' exonuclease, which is required for removing an incorrectly incorporated 3'-terminal nucleotide during DNA elongation by DNA polymerase activity. The key amino acid residues in the exonuclease center of Chlamydophila pneumoniae DNApolI (CpDNApolI) are naturally mutated, resulting in the loss of 3'-5' exonuclease. Hence, the manner by which CpDNApolI proofreads the incorrectly incorporated nucleotide during DNA synthesis warrants clarification. C. pneumoniae encodes three 3'-5' exonuclease activities: one endonuclease IV and two homologs of the epsilon subunit of replicative DNA polymerase III. The three proteins were biochemically characterized using single- and double-stranded DNA substrate. Among them, C. pneumoniae endonuclease IV (CpendoIV) possesses 3'-5' exonuclease activity that prefers to remove mismatched 3'-terminal nucleotides in the nick, gap, and 3' recess of a double-stranded DNA (dsDNA). Finally, we reconstituted the proofreading reaction of the mismatched 3'-terminal nucleotide using the dsDNA with a nick or 3' recess as substrate. Upon proofreading of the mismatched 3'-terminal nucleotide by CpendoIV, CpDNApolI can correctly reincorporate the matched nucleotide and the nick is further sealed by DNA ligase. Based on our biochemical results, we proposed that CpendoIV was responsible for proofreading the replication errors of CpDNApolI.
authors.affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 800 Dong-Chuan Road, Shanghai, China.
authors.forename: Juan-Juan
authors.initials: JJ
authors.lastname: Xie
authors.forename: Xi-Peng
authors.initials: XP
authors.lastname: Liu
authors.forename: Zhong
authors.initials: Z
authors.lastname: Han
authors.forename: Hui
authors.initials: H
authors.lastname: Yuan
authors.forename: You
authors.initials: Y
authors.lastname: Wang
authors.forename: Jing-Li
authors.initials: JL
authors.lastname: Hou
authors.forename: Jian-Hua
authors.initials: JH
authors.lastname: Liu
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D020137:Base Pair Mismatch; D016993:Chlamydophila pneumoniae / Q000201:enzymology*; D053904:DNA Breaks, Single-Stranded; D053843:DNA Mismatch Repair; D004269:DNA, Bacterial / Q000096:biosynthesis; D004277:DNA, Single-Stranded / Q000378:metabolism*; D043223:Deoxyribonuclease IV (Phage T4-Induced) / Q000378:metabolism*; D012265:Ribonucleotides / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D012265:Ribonucleotides; D043223:Deoxyribonuclease IV (Phage T4-Induced)