title: Bacterial nonhomologous end joining ligases preferentially seal breaks with a 3'-OH monoribonucleotide.
abstract: Many bacterial species have a nonhomologous end joining system of DNA repair driven by dedicated DNA ligases (LigD and LigC). LigD is a multifunctional enzyme composed of a ligase domain fused to two other catalytic modules: a polymerase that preferentially adds ribonucleotides to double-strand break ends and a phosphoesterase that trims 3'-oligoribonucleotide tracts until only a single 3'-ribonucleotide remains. LigD and LigC have a feeble capacity to seal 3'-OH/5'-PO(4) DNA nicks. Here, we report that nick sealing by LigD and LigC enzymes is stimulated by the presence of a single ribonucleotide at the broken 3'-OH end. The ribonucleotide effect on LigD and LigC is specific for the 3'-terminal nucleotide and is either diminished or abolished when additional vicinal ribonucleotides are present. No such 3'-ribonucleotide effect is observed for bacterial LigA or Chlorella virus ligase. We found that in vitro repair of a double-strand break by Pseudomonas LigD requires the polymerase module and results in incorporation of an alkali-labile ribonucleotide at the repair junction. These results illuminate an underlying logic for the domain organization of LigD, whereby the polymerase and phosphoesterase domains can heal the broken 3'-end to generate the monoribonucleotide terminus favored by the nonhomologous end joining ligases.
authors.affiliation: Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10021, USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhu
authors.forename: Stewart
authors.initials: S
authors.lastname: Shuman
mesh_terms: D001483:Base Sequence; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D011088:DNA Ligases / Q000378:metabolism*; D004260:DNA Repair / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D012265:Ribonucleotides / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D012265:Ribonucleotides; D011088:DNA Ligases