Characterization of an ATP-dependent DNA ligase encoded by Chlorella virus PBCV-1

C. Kiong Ho, James L. Van Etten, Stewart Shuman

Research output: Contribution to journalArticle

72 Scopus citations

Abstract

We report that Chlorella virus PBCV-1 encodes a 298-amino-acid ATP- dependent DNA ligase. The PBCV-1 enzyme is the smallest member of the covalent nucleotidyl transferase superfamily, which includes the ATP- dependent polynucleotide ligases and the GTP-dependent RNA capping enzymes. The specificity of PBCV-1 DNA ligase was investigated by using purified recombinant protein. The enzyme catalyzed efficient strand joining on a singly nicked DNA in the presence of magnesium and ATP (K(m), 75 μM). Other nucleoside triphosphates or deoxynucleoside triphosphates could not substitute for ATP. PBCV-1 ligase was unable to ligate across a 2-nucleotide gap and ligated poorly across a 1-nucleotide gap. A native gel mobility shift assay showed that PBCV-1 DNA ligase discriminated between nicked and gapped DNAs at the substrate-binding step. These findings underscore the importance of a properly positioned 3' OH acceptor terminus in substrate recognition and reaction chemistry.

Original languageEnglish (US)
Pages (from-to)1931-1937
Number of pages7
JournalJournal of virology
Volume71
Issue number3
DOIs
StatePublished - Mar 1997

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

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