Control of O-glycan branch formation. Molecular cloning and characterization of a novel thymus-associated core 2 beta1, 6-n-acetylglucosaminyltransferase.
Core 2 O-glycan branching catalyzed by UDP-N-acetyl-alpha-D-glucosamine: acceptor beta1, 6-N-acetylglucosaminyltransferases (beta6GlcNAc-Ts) is an important step in mucin-type biosynthesis. Core 2 complex-type O-glycans are involved in selectin-mediated adhesion events, and O-glycan branching appears to be highly regulated. Two homologous beta6GlcNAc-Ts functioning in O-glycan branching have previously been characterized, and here we report a third homologous beta6GlcNAc-T designated C2GnT3. C2GnT3 was identified by BLAST analysis of human genome survey sequences. The catalytic activity of C2GnT3 was evaluated by in vitro analysis of a secreted form of the protein expressed in insect cells. The results revealed exclusive core 2 beta6GlcNAc-T activity. The product formed with core 1-para-nitrophenyl was confirmed by (1)H NMR to be core 2-para-nitrophenyl. In vivo analysis of the function of C2GnT3 by coexpression of leukosialin (CD43) and a full coding construct of C2GnT3 in Chinese hamster ovary cells confirmed the core 2 activity and failed to reveal I activity. The C2GnT3 gene was located to 5q12, and the coding region was contained in a single exon. Northern analysis revealed selectively high levels of a 5.5-kilobase C2GnT3 transcript in thymus with only low levels in other organs. The unique expression pattern of C2GnT3 suggests that this enzyme serves a specific function different from other members of the beta6GlcNAc-T gene family.
Schwientek
T
T
School of Dentistry, University of Copenhagen, Norre Allé 20, 2200 Copenhagen N, Denmark. tsc@odont.ku.dk
Yeh
J C
JC
Levery
S B
SB
Keck
B
B
Merkx
G
G
van Kessel
A G
AG
Fukuda
M
M
Clausen
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D006224:Cricetinae; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry / Q000235:genetics*; D011134:Polysaccharides / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D011134:Polysaccharides; D017351:N-Acetylglucosaminyltransferases; C037649:N-acetyllactosaminide beta-1,6-N-acetylglucosaminyltransferase