title: A second NAD(+)-dependent DNA ligase (LigB) in Escherichia coli.
abstract: Escherichia coli DNA ligase (LigA) is the prototype of the NAD(+)-dependent class of DNA ligases found in all bacteria. Here we report the characterization of E.coli LigB, a second NAD(+)-dependent DNA ligase identified by virtue of its sequence similarity to LigA. LigB differs from LigA in that it lacks the BRCA1 C-terminus domain (BRCT) and two of the four Zn-binding cysteines that are present in LigA and all other bacterial NAD(+) ligases. We found that recombinant LigB catalyzed strand joining on a singly-nicked DNA in the presence of a divalent cation and NAD(+), and that LigB reacted with NAD(+) to form a covalent ligase-adenylate intermediate. Alanine substitution for the motif I lysine ((126)KxDG) abolished nick joining and ligase-adenylate formation by LigB, thus confirming that the ligase and adenylyltransferase activities are intrinsic to the LigB protein.
authors.affiliation: Molecular Biology Program, Sloan-Kettering Institute, 1275 York Avenue, New York, NY 10021, USA.
authors.forename: V
authors.initials: V
authors.lastname: Sriskanda
authors.forename: S
authors.initials: S
authors.lastname: Shuman
mesh_terms: D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D011088:DNA Ligases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008239:Lysine / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D017386:Sequence Homology, Amino Acid
chemical_list: D004269:DNA, Bacterial; D007527:Isoenzymes; D011088:DNA Ligases; C120640:DNA ligase (NAD); D008239:Lysine; D000409:Alanine