Characterization of a novel cis-syn and trans-syn-II pyrimidine dimer glycosylase/AP lyase from a eukaryotic algal virus, Paramecium bursaria chlorella virus-1

Amanda K. McCullough, Matthew T. Romberg, Simon Nyaga, Yuanfen Wei, Thomas G. Wood, John Stephen Taylor, James L. Van Etten, M. L. Dodson, R. Stephen Lloyd

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42 Scopus citations

Abstract

Endonuclease V from bacteriophage T4, is a cis-syn pyrimidine dimer- specific glycosylase. Recently, the first sequence homolog of T4 endonuclease V was identified from chlorella virus Paramecium bursaria chlorella virus-1 (PBCV-1). Here we present the biochemical characterization of the chlorella virus pyrimidine dimer glycosylase, cv-PDG. Interestingly, cv-PDG is specific not only for the cis-syn cyclobutane pyrimidine dimer, but also for the trans-syn-II isomer. This is the first trans-syn-II-specific glycosylase identified to date. Kinetic analysis demonstrates that DNAs containing both types of pyrimidine dimers are cleaved by the enzyme with similar catalytic efficiencies. Cleavage analysis and covalent trapping experiments demonstrate that the enzyme mechanism is consistent with the model proposed for glycosylase/AP lyase enzymes in which the glycosylase action is mediated via an imino intermediate between the C1' of the sugar and an amino group in the enzyme, followed by a β-elimination reaction resulting in cleavage of the phosphodiester bond. cv-PDG exhibits processive cleavage kinetics which are diminished at salt concentrations greater than those determined for T4 endonuclease V, indicating a possibly stronger electrostatic attraction between enzyme and DNA. The identification of this new enzyme with broader pyrimidine dimer specificity raises the intriguing possibility that there may be other T4 endonuclease V-like enzymes with specificity toward other DNA photoproducts.

Original languageEnglish (US)
Pages (from-to)13136-13142
Number of pages7
JournalJournal of Biological Chemistry
Volume273
Issue number21
DOIs
StatePublished - May 22 1998

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    McCullough, A. K., Romberg, M. T., Nyaga, S., Wei, Y., Wood, T. G., Taylor, J. S., Van Etten, J. L., Dodson, M. L., & Lloyd, R. S. (1998). Characterization of a novel cis-syn and trans-syn-II pyrimidine dimer glycosylase/AP lyase from a eukaryotic algal virus, Paramecium bursaria chlorella virus-1. Journal of Biological Chemistry, 273(21), 13136-13142. https://doi.org/10.1074/jbc.273.21.13136