Crystallization and preliminary X-ray diffraction studies of a hyperthermophilic Rieske protein variant (SDX-triple) with an engineered rubredoxin-like mononuclear iron site.
In place of the Rieske [2Fe-2S] cluster, an archetypal mononuclear iron site has rationally been designed into a hyperthermophilic archaeal Rieske [2Fe-2S] protein (sulredoxin) from Sulfolobus tokodaii by three residue replacements with reference to the Pyrococcus furiosus rubredoxin sequence. The resulting sulredoxin variant, SDX-triple (H44I/A45C/H64C), has been purified and crystallized by the hanging-drop vapour-diffusion method using 65%(v/v) 2-methyl-2,4-pentanediol, 0.025 M citric acid and 0.075 M sodium acetate trihydrate pH 4.3. The crystals diffract to 1.63 A resolution and belong to the triclinic space group P1, with unit-cell parameters a = 43.56, b = 76.54, c = 80.28 A, alpha = 88.12, beta = 78.82, gamma = 73.46 degrees. The asymmetric unit contains eight protein molecules.
Iwasaki
Toshio
T
Department of Biochemistry and Molecular Biology, Nippon Medical School, Sendagi, Bunkyo-ku, Tokyo 113-8602, Japan. tiwasaki@nms.ac.jp
Kounosu
Asako
A
Ohmori
Daijiro
D
Kumasaka
Takashi
T
D000595:Amino Acid Sequence; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D014450:Electron Transport Complex III / Q000737:chemistry* / Q000235:genetics; D007501:Iron / Q000737:chemistry* / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D015202:Protein Engineering; D020027:Pyrococcus furiosus / Q000737:chemistry / Q000201:enzymology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D012416:Rubredoxins / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D016944:Sulfolobus / Q000737:chemistry / Q000201:enzymology; D014961:X-Ray Diffraction
D007506:Iron-Sulfur Proteins; D011994:Recombinant Proteins; C037989:Rieske iron-sulfur protein; D012416:Rubredoxins; D007501:Iron; D014450:Electron Transport Complex III