title: Structural and kinetic studies of induced fit in xylulose kinase from Escherichia coli.
abstract: The primary metabolic route for D-xylose, the second most abundant sugar in nature, is via the pentose phosphate pathway after a two-step or three-step conversion to xylulose-5-phosphate. Xylulose kinase (XK; EC 2.7.1.17) phosphorylates D-xylulose, the last step in this conversion. The apo and D-xylulose-bound crystal structures of Escherichia coli XK have been determined and show a dimer composed of two domains separated by an open cleft. XK dimerization was observed directly by a cryo-EM reconstruction at 36 A resolution. Kinetic studies reveal that XK has a weak substrate-independent MgATP-hydrolyzing activity, and phosphorylates several sugars and polyols with low catalytic efficiency. Binding of pentulose and MgATP to form the reactive ternary complex is strongly synergistic. Although the steady-state kinetic mechanism of XK is formally random, a path is preferred in which D-xylulose binds before MgATP. Modelling of MgATP binding to XK and the accompanying conformational change suggests that sugar binding is accompanied by a dramatic hinge-bending movement that enhances interactions with MgATP, explaining the observed synergism. A catalytic mechanism is proposed and supported by relevant site-directed mutants.
authors.affiliation: Section of Molecular and Cellular Biology, University of California, Davis, CA 95616, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Di Luccio
authors.forename: Barbara
authors.initials: B
authors.lastname: Petschacher
authors.forename: Jennifer
authors.initials: J
authors.lastname: Voegtli
authors.forename: Hui-Ting
authors.initials: HT
authors.lastname: Chou
authors.forename: Henning
authors.initials: H
authors.lastname: Stahlberg
authors.forename: Bernd
authors.initials: B
authors.lastname: Nidetzky
authors.forename: David K
authors.initials: DK
authors.lastname: Wilson
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry; D000595:Amino Acid Sequence; D002241:Carbohydrates / Q000737:chemistry; D002384:Catalysis; D017124:Conserved Sequence; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D005991:Glycerol Kinase / Q000737:chemistry; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D002241:Carbohydrates; D004791:Enzyme Inhibitors; D017853:Phosphotransferases (Alcohol Group Acceptor); C032700:xylulokinase; D005991:Glycerol Kinase; D000251:Adenosine Triphosphatases