title: Expression, purification, crystallization and preliminary X-ray diffraction analysis of galactokinase from Pyrococcus horikoshii.
abstract: Galactokinase (EC 2.7.1.6) catalyzes the ATP-dependent phosphorylation of alpha-D-galactose to alpha-D-galactose-1-phosphate, in an additional metabolic branch of glycolysis. The apo-form crystal structure of the enzyme has not yet been elucidated. Crystals of galactokinase from Pyrococcus horikoshii were prepared in both the apo form and as a ternary complex with alpha-D-galactose and an ATP analogue. Diffraction data sets were collected to 1.24 A resolution for the apo form and to 1.7 A for the ternary complex form using synchrotron radiation. The apo-form crystals belong to space group C2, with unit-cell parameters a = 108.08, b = 38.91, c = 81.57 A, beta = 109.8 degrees. The ternary complex form was isomorphous with the apo form, except for the length of the a axis. The galactokinase activity of the enzyme was confirmed and the kinetic parameters at 323 K were determined.
authors.affiliation: RIKEN SPring-8 Center, Harima Institute, 1-1-1 Kouto, Sayo, Hyogo 679-5148, Japan.
authors.forename: Eiji
authors.initials: E
authors.lastname: Inagaki
authors.forename: Keiko
authors.initials: K
authors.lastname: Sakamoto
authors.forename: Naomi
authors.initials: N
authors.lastname: Obayashi
authors.forename: Takaho
authors.initials: T
authors.lastname: Terada
authors.forename: Mikako
authors.initials: M
authors.lastname: Shirouzu
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Bessho
authors.forename: Chizu
authors.initials: C
authors.lastname: Kuroishi
authors.forename: Seiki
authors.initials: S
authors.lastname: Kuramitsu
authors.forename: Akeo
authors.initials: A
authors.lastname: Shinkai
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
mesh_terms: D001051:Apoenzymes / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D019843:Archaeal Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005686:Galactokinase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D019848:Gene Expression Regulation, Archaeal; D007700:Kinetics; D043902:Pyrococcus horikoshii / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D001051:Apoenzymes; D019843:Archaeal Proteins; D011994:Recombinant Proteins; D005686:Galactokinase