title: Chondroitin sulfate synthase-3. Molecular cloning and characterization.
abstract: Recently, it has become evident that chondroitin sulfate (CS) glycosyltransferases, which transfer glucuronic acid and/or N-acetylgalactosamine residues from each UDP-sugar to the nonreducing terminus of the CS chain, form a gene family. We report here a novel human gene (GenBank trade mark accession number AB086062) that possesses a sequence homologous with the human chondroitin sulfate synthase-1 (CSS1) gene, formerly known as chondroitin synthase. The full-length open reading frame consists of 882 amino acids and encodes a typical type II membrane protein. This enzyme contains a beta 3-glycosyltransferase motif and a beta 4-glycosyltransferase motif similar to that found in CSS1. Both the enzymes were expressed in COS-7 cells as soluble proteins, and their enzymatic natures were characterized. Both glucuronyltransferase and N-acetylgalactosaminyltransferase activities were observed when chondroitin, CS polymer, and their corresponding oligosaccharides were used as the acceptor substrates, but no polymerization reaction was observed as in the case of CSS1. The new enzyme was thus designated chondroitin sulfate synthase-3 (CSS3). However, the specific activity of CSS3 was much lower than that of CSS1. The reaction products were shown to have a GlcUA beta 1-3GalNAc linkage and a GalNAc beta 1-4GlcUA linkage in the nonreducing terminus of chondroitin resulting from glucuronyltransferase activity and N-acetylgalactosaminyltransferase activity, respectively. Quantitative real time PCR analysis revealed that the transcript level of CSS3 was much lower than that of CSS1, although it was ubiquitously expressed in various human tissues. These results indicate that CSS3 is a glycosyltransferase having both glucuronyltransferase and N-acetylgalactosaminyltransferase activities. It may make a contribution to CS biosynthesis that differs from that of CSS1.
authors.affiliation: Institute for Molecular Science of Medicine, Aichi Medical University, Nagakute, Aichi 480-1195, Japan.
authors.forename: Toshikazu
authors.initials: T
authors.lastname: Yada
authors.forename: Takashi
authors.initials: T
authors.lastname: Sato
authors.forename: Hiromi
authors.initials: H
authors.lastname: Kaseyama
authors.forename: Masanori
authors.initials: M
authors.lastname: Gotoh
authors.forename: Hiroko
authors.initials: H
authors.lastname: Iwasaki
authors.forename: Norihiro
authors.initials: N
authors.lastname: Kikuchi
authors.forename: Yeon-Dae
authors.initials: YD
authors.lastname: Kwon
authors.forename: Akira
authors.initials: A
authors.lastname: Togayachi
authors.forename: Takashi
authors.initials: T
authors.lastname: Kudo
authors.forename: Hideto
authors.initials: H
authors.lastname: Watanabe
authors.forename: Hisashi
authors.initials: H
authors.lastname: Narimatsu
authors.forename: Koji
authors.initials: K
authors.lastname: Kimata
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002809:Chondroitin Sulfates / Q000096:biosynthesis; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D016695:Glycosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006602:Hexosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014018:Tissue Distribution
chemical_list: D018076:DNA, Complementary; D008565:Membrane Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D002809:Chondroitin Sulfates; D016695:Glycosyltransferases; D006602:Hexosyltransferases; C478997:CHSY3 protein, human