title: Use of a repetitive seeding protocol to obtain diffraction-quality crystals of a putative human D-xylulokinase.
abstract: In mammals, the enzyme D-xylulokinase (XK; EC 2.7.1.17) catalyses the last step of the glucuronate-xylulose pathway, in which the ketopentose sugar D-xylulose is phosphorylated to yield D-xylulose 5-phosphate (Xu5P). Xu5P is also a metabolite of the pentose phosphate pathway and acts as a signalling molecule that regulates lipogenesis and glycolysis in the liver. To date, no eukaryotic XK has been structurally characterized. A putative human XK was expressed in Escherichia coli aided by molecular chaperones, purified and crystallized. A seeding procedure involving repeated rounds of seeding was developed and proved to be essential for obtaining diffraction-quality crystals. Preliminary X-ray diffraction analysis was performed using synchrotron radiation. This resulted in the collection of a complete diffraction data set to 2.7 Å resolution from a crystal belonging to the trigonal space group P3(1) or P3(2) with unit-cell parameters a = b = 101.87, c = 158.85 Å.
authors.affiliation: Maurice Wilkins Centre for Molecular Biodiscovery and School of Biological Sciences, University of Auckland, Private Bag 92019, Auckland, New Zealand.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Bunker
authors.forename: James M J
authors.initials: JM
authors.lastname: Dickson
authors.forename: Tom T
authors.initials: TT
authors.lastname: Caradoc-Davies
authors.forename: Kerry M
authors.initials: KM
authors.lastname: Loomes
authors.forename: Edward N
authors.initials: EN
authors.lastname: Baker
mesh_terms: D018360:Crystallography, X-Ray; D006801:Humans; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry*
chemical_list: D017853:Phosphotransferases (Alcohol Group Acceptor); C032700:xylulokinase