Molecular cloning and characterization of human GnT-IX, a novel beta1,6-N-acetylglucosaminyltransferase that is specifically expressed in the brain.
A novel beta1,6-N-acetylglucosaminyltransferase (beta1, 6GnT) cDNA was identified by a BLAST search using the amino acid sequence of human GnT-V as a query. The full-length sequence was determined by a combination of 5'-rapid amplification of cDNA end analysis and a further data base search. The open reading frame encodes a 792 amino acid protein with a type II membrane protein structure typical of glycosyltransferases. The entire sequence identity to human GnT-V is 42%. When pyridylaminated (PA) agalacto biantennary N-linked oligosaccharide was used as an acceptor substrate, the recombinant enzyme generated a novel product other than the expected GnT-V product, (GlcNAcbeta1,2-Manalpha1,3-)[GlcNAcbeta1,2-(GlcNAcbeta1,6-)Manalpha1,6-]Manbeta1,4-GlcNAcbeta1,4-GlcNAc-PA. This new product was identified as [GlcNAcbeta1,2-(GlcNAcbeta1,6-)Manalpha1,3-][Glc-NAcbeta1,2-(GlcNAcbeta1,6-)Manalpha1,6-]Manbeta1,4-GlcNAcbeta1,4-GlcNAc-PA by mass spectrometry and 1H NMR. Namely, the new GnT (designated as GnT-IX) has beta1,6GnT activity not only to the alpha1,6-linked mannose arm but also to the alpha1,3-linked mannose arm of N-glycan, forming a unique structure that has not been reported to date. Northern blot analysis showed that the GnT-IX gene is exclusively expressed in the brain, whereas the GnT-V gene is expressed ubiquitously. These results suggest that GnT-IX is responsible for the synthesis of a unique oligosaccharide structure in the brain.
Inamori
Kei-ichiro
K
Department of Biochemistry, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
Endo
Takeshi
T
Ide
Yoshihito
Y
Fujii
Shigeru
S
Gu
Jianguo
J
Honke
Koichi
K
Taniguchi
Naoyuki
N
D000595:Amino Acid Sequence; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D001921:Brain / Q000201:enzymology*; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D005822:Genetic Vectors; D006031:Glycosylation; D006801:Humans; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D008956:Models, Chemical; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics*; D009419:Nerve Tissue Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D009474:Neurons / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism; D016366:Open Reading Frames; D011233:Precipitin Tests; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014018:Tissue Distribution
D018076:DNA, Complementary; D008024:Ligands; D009419:Nerve Tissue Proteins; D009844:Oligosaccharides; D012333:RNA, Messenger; D004247:DNA; C479741:MGAT5B protein, human; D017351:N-Acetylglucosaminyltransferases; C052254:alpha-1,6-mannosylglycoprotein beta 1,6-N-acetylglucosaminyltransferase