title: Heterologous expression and biochemical characterization of soluble human xylosyltransferase II.
abstract: Human xylosyltransferase II (EC 2.4.2.26, XT-II) represents an isoform of xylosyltransferase I (XT-I). Recently, we and others provided first evidence that XT-II is capable of initiating the biosynthesis of glycosaminoglycan chains in proteoglycans. Here, a soluble form of human XT-II was expressed in the yeast Pichia pastoris and the substrate specificity for various acceptors was investigated, pointing to a modified bikunin peptide to be the optimal XT-II acceptor (K(M)=1.9 microM). Furthermore, biochemical characterization of XT-II showed that this enzyme was strongly inhibited by nucleotides and glycosaminoglycans. Its temperature optimum, stability, and ion dependency were further examined, demonstrating necessity for Mg(2+) or Mn(2+) ions for its enzymatic activity. Our data show for the first time that XT-I and XT-II are xylosyltransferases with similar but not identical properties, pointing to their potential role in modulating the cellular proteoglycan pool.
authors.affiliation: Institut für Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum Nordrhein-Westfalen, Universitätsklinik der Ruhr-Universität Bochum, Georgstrasse 11, 32545, Bad Oeynhausen, Germany.
authors.forename: Javier Carrera
authors.initials: JC
authors.lastname: Casanova
authors.forename: Joachim
authors.initials: J
authors.lastname: Kuhn
authors.forename: Knut
authors.initials: K
authors.lastname: Kleesiek
authors.forename: Christian
authors.initials: C
authors.lastname: Götting
mesh_terms: D004789:Enzyme Activation; D004795:Enzyme Stability; D006801:Humans; D010430:Pentosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012995:Solubility; D014162:Transfection / Q000379:methods*; D000097608:UDP Xylose-Protein Xylosyltransferase
chemical_list: D010430:Pentosyltransferases; D011994:Recombinant Proteins; D000097608:UDP Xylose-Protein Xylosyltransferase