Expression, purification and crystallization of two endonuclease III enzymes from Deinococcus radiodurans

Acta Crystallogr F Struct Biol Commun. 2014 Dec 1;70(Pt 12):1688-92. doi: 10.1107/S2053230X14024935. Epub 2014 Nov 28.

Abstract

Endonuclease III is a bifunctional DNA glycosylase that removes a wide range of oxidized bases in DNA. Deinococcus radiodurans is an extreme radiation-resistant and desiccation-resistant bacterium and possesses three genes encoding endonuclease III enzymes in its genome: DR2438 (EndoIII-1), DR0289 (EndoIII-2) and DR0982 (EndoIII-3). Here, EndoIII-1 and an N-terminally truncated form of EndoIII-3 (EndoIII-3Δ76) have been expressed, purified and crystallized, and preliminary X-ray crystallographic analyses have been performed to 2.15 and 1.31 Å resolution, respectively. The EndoIII-1 crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 181.38, b = 38.56, c = 37.09 Å, β = 89.34° and one molecule per asymmetric unit. The EndoIII-3Δ76 crystals also belonged to the monoclinic space group C2, but with unit-cell parameters a = 91.47, b = 40.53, c = 72.47 Å, β = 102.53° and one molecule per asymmetric unit. The EndoIII-1 structure was determined by molecular replacement, while the truncated EndoIII-3Δ76 structure was determined by single-wavelength anomalous dispersion phasing. Refinement of the structures is in progress.

Keywords: Deinococcus radiodurans; EndoIII-1; EndoIII-3; endonuclease III.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallization
  • Crystallography, X-Ray
  • Deinococcus / enzymology*
  • Endonucleases / chemistry
  • Endonucleases / metabolism*

Substances

  • Endonucleases