title: Identification and characterization of three Drosophila melanogaster glucuronyltransferases responsible for the synthesis of the conserved glycosaminoglycan-protein linkage region of proteoglycans. Two novel homologs exhibit broad specificity toward oligosaccharides from proteoglycans, glycoproteins, and glycosphingolipids.
abstract: The Drosophila melanogaster genome contains three putative glucuronyltransferases homologous to human GlcAT-I and GlcAT-P. These enzymes are predicted to be beta1,3-glucuronyltransferases involved in the synthesis of the glycosaminoglycan (GAG)-protein linkage region of proteoglycans and the HNK-1 carbohydrate epitope of glycoproteins, respectively. The genes encode active enzymes, which we have designated DmGlcAT-I, DmGlcAT-BSI, and DmGlcAT-BSII (where BS stands for "broad specificity"). Protein A-tagged truncated soluble forms of all three enzymes efficiently transfer GlcUA from UDP-GlcUA to the linkage region trisaccharide Galbeta1-3Galbeta1-4Xyl. Strikingly, DmGlcAT-I has specificity for Galbeta1-3Galbeta1-4Xyl, whereas DmGlcAT-BSI and DmGlcAT-BSII act on a wide array of substrates with non-reducing terminal beta1,3- and beta1,4-linked Gal residues. Their highest activities are obtained with asialoorosomucoid with a terminal Galbeta1-4GlcNAc sequence, indicating their possible involvement in the synthesis of the HNK-1 epitope in addition to the GAG-protein linkage region. Galbeta1-3GlcNAc and Galbeta1-3GalNAc, disaccharide structures widely found in N- and O-glycans of glycoproteins and glycolipids, also serve as acceptors for DmGlcAT-BSI and -BSII. Transcripts of all three enzymes are ubiquitously expressed throughout the developmental stages and in adult tissues of Drosophila. Thus, all three glucuronyltransferases are likely involved in the synthesis of the GAG-protein linkage region in Drosophila, and DmGlcAT-BSI and -BSII appear to be involved in various GlcUA transfer reactions for the synthesis of proteoglycans, glycoproteins, and glycolipids. This activity distinguishes these glucuronyltransferases from their mammalian homologs GlcAT-P and GlcAT-D (or -S). Sequence alignment of the Drosophila glucuronyltransferases with homologs in human, rat, and Caenorhabditis elegans demonstrates the conservation of a majority of the critical amino acid residues in the active sites of the three Drosophila enzymes.
authors.affiliation: Department of Biochemistry, Kobe Pharmaceutical University, Higashinada-ku, Kobe 658-8558, Japan.
authors.forename: Byung-Taek
authors.initials: BT
authors.lastname: Kim
authors.forename: Kazunori
authors.initials: K
authors.lastname: Tsuchida
authors.forename: John
authors.initials: J
authors.lastname: Lincecum
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kitagawa
authors.forename: Merton
authors.initials: M
authors.lastname: Bernfield
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Sugahara
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D018076:DNA, Complementary / Q000378:metabolism; D019992:Databases as Topic; D029721:Drosophila Proteins / Q000737:chemistry*; D004331:Drosophila melanogaster / Q000201:enzymology*; D000939:Epitopes; D008040:Genetic Linkage; D016678:Genome; D014453:Glucuronosyltransferase / Q000737:chemistry* / Q000235:genetics*; D006025:Glycosaminoglycans / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry; D010802:Phylogeny; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D018076:DNA, Complementary; D029721:Drosophila Proteins; D000939:Epitopes; D006025:Glycosaminoglycans; D009844:Oligosaccharides; C067806:glucuronyltransferase GlcAT-1; C473770:GlcAT-P protein, Drosophila; C473769:GlcAT-S protein, Drosophila; D014453:Glucuronosyltransferase