Molecular cloning and expression of a novel beta-1, 6-N-acetylglucosaminyltransferase that forms core 2, core 4, and I branches

J Biol Chem. 1999 Jan 29;274(5):3215-21. doi: 10.1074/jbc.274.5.3215.

Abstract

Mucin-type O-glycans are classified according to their core structures. Among them, cores 2 and 4 are important for having N-acetyllactosamine side chains, which can be further modified to express various functional oligosaccharides. Previously, we discovered by cloning cDNAs that the core 2 branching enzyme, termed core 2 beta-1,6-N-acetylglucosaminyltransferase-leukocyte type (C2GnT-L), is highly homologous to the I branching beta-1, 6-N-acetylglucosaminyltransferase (IGnT) (Bierhuizen, M. F. A., Mattei, M.-G., and Fukuda, M. (1993) Genes Dev. 7, 468-478). Using these homologous sequences as probes, we identified an expressed sequence tag in dbEST, which has significant homology to C2GnT-L and IGnT. This approach, together with 5'and 3' rapid amplification of cDNA ends, yielded a human cDNA that encompasses a whole coding region of an enzyme, termed C2GnT-mucin type (C2GnT-M). C2GnT-M has 48.2 and 33.8% identity with C2GnT-L and IGnT at the amino acid levels. The expression of C2GnT-M cDNA directed the expression of core 2 branched oligosaccharides and I antigen on the cell surface. Moreover, a soluble chimeric C2GnT-M had core 4 branching activity in addition to core 2 and I branching activities. A soluble chimeric C2GnT-L, in contrast, almost exclusively contains core 2 branching activity. Northern blot analysis demonstrated that the C2GnT-M transcripts are heavily expressed in colon, small intestine, trachea, and stomach, where mucin is produced. In contrast, the transcripts of C2GnT-L were more widely detected, including the lymph node and bone marrow. These results indicate that the newly cloned C2GnT-M plays a critical role in O-glycan synthesis in mucins and might have distinctly different roles in oligosaccharide ligand formation compared with C2GnT-L.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • CHO Cells
  • Cloning, Molecular
  • Cricetinae
  • DNA, Complementary / chemistry
  • DNA, Complementary / isolation & purification
  • Humans
  • Molecular Sequence Data
  • Mucins / genetics*
  • N-Acetylglucosaminyltransferases / chemistry
  • N-Acetylglucosaminyltransferases / genetics*
  • Sequence Homology, Amino Acid
  • Surface Properties

Substances

  • DNA, Complementary
  • Mucins
  • N-acetylglucosaminyltransferase IGnT
  • N-Acetylglucosaminyltransferases
  • beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-N-acetylglucosaminyltransferase 3
  • beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-acetylglucosaminyl transferase

Associated data

  • GENBANK/AF102542