title: Crystallization and preliminary X-ray diffraction studies of monomeric isocitrate dehydrogenase by the MAD method using Mn atoms.
abstract: NADP(+)-dependent isocitrate dehydrogenase (E.C. 1.1.1.42; IDH) is an enzyme of the Krebs cycle and catalyzes the oxidative decarboxylation reaction from DL-isocitrate to alpha-ketoglutarate, with a concomitant reduction of the coenzyme NADP(+) to NADPH. Single crystals of monomeric IDH from Azotobacter vinelandii in complex with DL-isocitrate and Mn(2+) were obtained by the hanging-drop vapour-diffusion method at room temperature. One crystal diffracted to a resolution of 2.9 A and was found to belong to the orthorhombic system; the space group was determined to be P2(1)2(1)2(1), with unit-cell parameters a = 108.4, b = 121.7, c = 129.7 A. The asymmetric unit contains two molecules of monomeric IDH, corresponding to a V(M) value of 2.66 A(3) Da(-1). The crystals were frozen in a capillary by a flash-cooling technique and MAD data were collected using Mn atoms as anomalous scatterers on beamline BL41XU at SPring-8, Japan. The positions of two Mn atoms binding to two independent IDH molecules were located from Bijvoet difference Patterson maps.
authors.affiliation: Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo 060-0810, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Yasutake
authors.forename: S
authors.initials: S
authors.lastname: Watanabe
authors.forename: M
authors.initials: M
authors.lastname: Yao
authors.forename: Y
authors.initials: Y
authors.lastname: Takada
authors.forename: N
authors.initials: N
authors.lastname: Fukunaga
authors.forename: I
authors.initials: I
authors.lastname: Tanaka
mesh_terms: D016948:Azotobacter vinelandii / Q000201:enzymology*; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods; D007521:Isocitrate Dehydrogenase / Q000737:chemistry*; D008345:Manganese / Q000737:chemistry*; D011487:Protein Conformation; D012984:Software
chemical_list: D008345:Manganese; D007521:Isocitrate Dehydrogenase