title: Purification, crystallization, X-ray diffraction analysis and phasing of an engineered single-chain PvuII restriction endonuclease.
abstract: The restriction endonuclease PvuII from Proteus vulgaris has been converted from its wild-type homodimeric form into the enzymatically active single-chain variant scPvuII by tandemly joining the two subunits through the peptide linker Gly-Ser-Gly-Gly. scPvuII, which is suitable for the development of programmed restriction endonucleases for highly specific DNA cleavage, was purified and crystallized. The crystals diffract to a resolution of 2.35 A and belong to space group P4(2), with unit-cell parameters a = b = 101.92, c = 100.28 A and two molecules per asymmetric unit. Phasing was successfully performed by molecular replacement.
authors.affiliation: Department of Science, Agricultural University of Athens, Athens, Greece.
authors.forename: Chrysi
authors.initials: C
authors.lastname: Meramveliotaki
authors.forename: Dina
authors.initials: D
authors.lastname: Kotsifaki
authors.forename: Maria
authors.initials: M
authors.lastname: Androulaki
authors.forename: Athanasios
authors.initials: A
authors.lastname: Hountas
authors.forename: Elias
authors.initials: E
authors.lastname: Eliopoulos
authors.forename: Michael
authors.initials: M
authors.lastname: Kokkinidis
mesh_terms: D003460:Crystallization; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015202:Protein Engineering / Q000379:methods; D011514:Proteus vulgaris / Q000201:enzymology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D014961:X-Ray Diffraction
chemical_list: D011994:Recombinant Proteins; C025630:CAGCTG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific