Open AccessExpression, purification, crystallization and preliminary X‐ray diffraction analysis of galactokinase from Pyrococcus horikoshii
Author(s) -
Inagaki Eiji,
Sakamoto Keiko,
Obayashi Naomi,
Terada Takaho,
Shirouzu Mikako,
Bessho Yoshitaka,
Kuroishi Chizu,
Kuramitsu Seiki,
Shinkai Akeo,
Yokoyama Shigeyuki
Publication year - 2006
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
ISSN - 1744-3091
DOI - 10.1107/s1744309106001813
Subject(s) - galactokinase , pyrococcus horikoshii , crystallography , chemistry , ternary complex , stereochemistry , crystal structure , enzyme , biochemistry , escherichia coli , gene
Galactokinase (EC 2.7.1.6) catalyzes the ATP‐dependent phosphorylation of α‐ d ‐galactose to α‐ 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 α‐ d ‐galactose and an ATP analogue. Diffraction data sets were collected to 1.24 Å resolution for the apo form and to 1.7 Å for the ternary complex form using synchrotron radiation. The apo‐form crystals belong to space group C 2, with unit‐cell parameters a = 108.08, b = 38.91, c = 81.57 Å, β = 109.8°. 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.