A Novel Human β1,3-N-Acetylgalactosaminyltransferase That Synthesizes a Unique Carbohydrate Structure, GalNAcβ1-3GlcNAc

• Published in: The Journal of biological chemistry Vol. 279; no. 14; p. 14087
• Main Authors: Toru Hiruma, Akira Togayachi, Kayo Okamura, Takashi Sato, Norihiro Kikuchi, Yeon-Dae Kwon, Aya Nakamura, Katsuya Fujimura, Masanori Gotoh, Kouichi Tachibana, Yasuko Ishizuka, Toshiaki Noce, Hiroshi Nakanishi, Hisashi Narimatsu
• Format: Journal Article
• Language: English
• Published: American Society for Biochemistry and Molecular Biology 02.04.2004
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M310614200

We found, using a BLAST search, a novel human gene (GenBank™ accession number BC029564) that possesses β3-glycosyltransferase motifs. The full-length open reading frame consists of 500 amino acids and encodes a typical type II membrane protein. This enzyme has a domain containing β1,3-glycosyltransferase motifs, which are widely conserved in the β1,3-galactosyltransferase and β1,3- N -acetylglucosaminyltransferase families. The putative catalytic domain was expressed in human embryonic kidney 293T cells as a soluble protein. Its N -acetylgalactosaminyltransferase activity was observed when N -acetylglucosamine (GlcNAc) β1- O -benzyl was used as an acceptor substrate. The enzyme product was determined to have a β1,3-linkage by NMR spectroscopic analysis, and was therefore named β1,3- N -acetylgalactosaminyltransferase-II (β3GalNAc-T2). The acceptor substrate specificity of β3GalNAc-T2 was examined using various oligosaccharide substrates. Galβ1-3(GlcNAcβ1-6)GalNAcα1- O - para -nitrophenyl (core 2- p NP) was the best acceptor substrate for β3GalNAc-T2, followed by GlcNAcβ1-4GlcNAcβ1- O -benzyl, and GlcNAcβ1-6GalNAcα1- O - para -nitrophenyl (core 6- p NP), among the tested oligosaccharide substrates. Quantitative real time PCR analysis revealed that the β 3Gal-NAc-T2 transcripts was restricted in its distribution mainly to the testis, adipose tissue, skeletal muscle, and ovary. Its putative orthologous gene, m β 3GalNAc-T2 , was also found in a data base of mouse expressed sequence tags. In situ hybridization analysis with mouse testis showed that the transcripts are expressed in germ line cells. β3GalNAc-T2 efficiently transferred GalNAc to N -glycans of fetal calf fetuin, which was treated with neuraminidase and β-galactosidase. However, it showed no activity toward any glycolipid examined. Although the GalNAcβ1-3GlcNAcβ1-R structure has not been reported in humans or other mammals, we have discovered a novel human glycosyltransferase producing this structure on N - and O -glycans.