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Expression, purification and crystallization of two endonuclease III enzymes from Deinococcus radiodurans
Author(s) -
Sarre Aili,
Ökvist Mats,
Klar Tobias,
Moe Elin,
Timmins Joanna
Publication year - 2014
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x14024935
Subject(s) - deinococcus radiodurans , monoclinic crystal system , deinococcus , endonuclease , crystallography , dna , enzyme , chemistry , microbiology and biotechnology , biology , stereochemistry , crystal structure , biochemistry
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 C 2, 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 C 2, 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.