title: Molecular cloning and expression of a second chondroitin N-acetylgalactosaminyltransferase involved in the initiation and elongation of chondroitin/dermatan sulfate.
abstract: We identified a novel human chondroitin N-acetylgalactosaminyltransferase, designated chondroitin GalNAcT-2 after a BLAST analysis of the GenBank(TM) data base using the sequence of a previously described human chondroitin N-acetylgalactosaminyltransferase (chondroitin GalNAcT-1) as a probe. The new cDNA sequence contained an open reading frame encoding a protein of 542 amino acids with a type II transmembrane protein topology. The amino acid sequence displayed 60% identity to that of human chondroitin GalNAcT-1. Like chondroitin GalNAcT-1, the expression of a soluble form of the protein in COS-1 cells produced an active enzyme, which not only transferred beta1,4-N-acetylgalactosamine (GalNAc) from UDP-[(3)H]GalNAc to a polymer chondroitin representing growing chondroitin chains (beta-GalNAc transferase II activity) but also to GlcUA beta 1-3Gal beta 1-O-C(2)H(4)NHCbz, a synthetic substrate for beta-GalNAc transferase I that transfers the first GalNAc to the core tetrasaccharide in the protein-linkage region of chondroitin sulfate. In contrast, the tetrasaccharide serine (GlcUA beta 1-3Gal beta 1-3Gal beta 1-4Xyl beta 1-O-Ser) derived from the linkage region, which is an inert acceptor substrate for chondroitin GalNAcT-1, served as an acceptor substrate. The coding region of this enzyme was divided into seven discrete exons, which is similar to the genomic organization of the chondroitin GalNAcT-1 gene, and was localized to chromosome 10q11.22. Northern blot analysis revealed that the chondroitin GalNAcT-2 gene exhibited a ubiquitous but differing expression in human tissues, and the expression pattern differed from that of chondroitin GalNAcT-1. Thus, we demonstrated redundancy in the chondroitin GalNAc transferases involved in the biosynthetic initiation and elongation of chondroitin sulfate, which is important for understanding the biosynthetic mechanisms leading to the selective chain assembly of chondroitin/dermatan sulfate on the linkage region tetrasaccharide common to various proteoglycans containing chondroitin/dermatan sulfate and heparin/heparan sulfate chains.
authors.affiliation: Department of Biochemistry, Kobe Pharmaceutical University, Higashinada-ku, Kobe 658-8558, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Uyama
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kitagawa
authors.forename: Junko
authors.initials: J
authors.lastname: Tanaka
authors.forename: Jun-ichi
authors.initials: J
authors.lastname: Tamura
authors.forename: Tomoya
authors.initials: T
authors.lastname: Ogawa
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Sugahara
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002809:Chondroitin Sulfates / Q000096:biosynthesis*; D003001:Cloning, Molecular; D003871:Dermatan Sulfate / Q000096:biosynthesis*; D004331:Drosophila melanogaster; D006801:Humans; D008969:Molecular Sequence Data; D017350:N-Acetylgalactosaminyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D011994:Recombinant Proteins; D003871:Dermatan Sulfate; D002809:Chondroitin Sulfates; D017350:N-Acetylgalactosaminyltransferases; C107227:UDP-N-acetylgalactosamine-chondroitin sulfate N-acetylgalactosaminyltransferase