title: A unique dye-decolorizing peroxidase, DyP, from Thanatephorus cucumeris Dec 1: heterologous expression, crystallization and preliminary X-ray analysis.
abstract: The dye-decolorizing peroxidase DyP is a key enzyme in the decolorizing fungus Thanatephorus cucumeris Dec 1 that degrades azo and antraquinone dyes. The gene dyp from T. cucumeris Dec 1, which has low homology to other peroxidase genes, was cloned and transformed into Aspergillus oryzae and glycosylated DyP was expressed at high levels. Purified DyP was deglycosylated using GST Endo F1 and then crystallized in a strong magnetic field (10 T) at 283 K using ammonium sulfate as precipitant. X-ray diffraction data to 2.96 A resolution collected from a native crystal at the Photon Factory (Tsukuba, Japan) showed that the crystal belonged to the hexagonal space group P6(5)22, with unit-cell parameters a = b = 136.15, c = 363.46 A. The asymmetric unit of the crystal contained four DyP molecules, with a corresponding Matthews coefficient (V(M)) of 2.50 A(3) Da(-1) and a solvent content of 51%. Heavy-atom derivatives of DyP have been obtained and electron-density maps have been calculated. The haem is visible and continuous electron density between the haem and protein clearly indicates the location of the proximal histidine ligand.
authors.affiliation: Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Sato
authors.forename: Shusaku
authors.initials: S
authors.lastname: Hara
authors.forename: Takuro
authors.initials: T
authors.lastname: Matsui
authors.forename: Gen
authors.initials: G
authors.lastname: Sazaki
authors.forename: Shinya
authors.initials: S
authors.lastname: Saijo
authors.forename: Tadashi
authors.initials: T
authors.lastname: Ganbe
authors.forename: Nobuo
authors.initials: N
authors.lastname: Tanaka
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Sugano
authors.forename: Makoto
authors.initials: M
authors.lastname: Shoda
mesh_terms: D001236:Aspergillus oryzae / Q000235:genetics; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D005848:Geotrichum / Q000201:enzymology*; D006031:Glycosylation; D008958:Models, Molecular; D010544:Peroxidases / Q000737:chemistry*
chemical_list: D004271:DNA, Fungal; D010544:Peroxidases