Molecular cloning and characterization of a novel UDP-Gal:GalNAc(alpha) peptide beta 1,3-galactosyltransferase (C1Gal-T2), an enzyme synthesizing a core 1 structure of O-glycan

J Biol Chem. 2002 Dec 6;277(49):47724-31. doi: 10.1074/jbc.M205839200. Epub 2002 Oct 1.

Abstract

Recently, a UDP-Gal:GalNAc(alpha) peptide beta1,3-galactosyltransferase (core 1 synthase 1; C1Gal-T1) has been purified from rat liver and its complementary DNA cloned from several species. We isolated a second candidate for core 1 synthase from a Colo205 cDNA library and named it C1Gal-T2. The deduced amino acid sequence of C1Gal-T2, having 26% homology to C1Gal-T1, showed a topology typical of a type II membrane protein. Real time PCR analysis revealed that the expression of C1Gal-T2 transcripts was widespread in many tissues and of relatively high level in salivary gland, stomach, small intestine, kidney, testis, thymus, and spleen. LSC cells, having no core 1 synthase activity, were transfected stably with the C1Gal-T2 gene. Their microsome fraction showed beta1,3-galactosyltransferase activity toward GalNAc-alpha-para-nitrophenyl and GalNAc(alpha)1 peptides resulting in the synthesis of the core 1 structure. The core 1 synthesizing activity of C1Gal-T2 was also determined by flow cytometry and lectin blotting using the LSC cells stably expressing C1Gal-T2. Finally, LSC cells, and Jurkat cells that also lack the core 1 synthase activity, were found to have null alleles of C1Gal-T2. These results indicated that C1Gal-T2 is the second candidate for core 1 synthase that plays an important role in synthesizing O-glycans in digestive organs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Chromatography, High Pressure Liquid
  • Cloning, Molecular
  • DNA, Complementary / metabolism
  • Databases as Topic
  • Exons
  • Expressed Sequence Tags
  • Flow Cytometry
  • Galactosyltransferases / chemistry*
  • Galactosyltransferases / genetics*
  • Gene Library
  • Humans
  • Jurkat Cells
  • Lectins
  • Molecular Sequence Data
  • Mutation
  • Open Reading Frames
  • Peptides / chemistry
  • Polysaccharides / chemistry
  • Reverse Transcriptase Polymerase Chain Reaction
  • Subcellular Fractions
  • Tissue Distribution
  • Transfection
  • Tumor Cells, Cultured

Substances

  • DNA, Complementary
  • Lectins
  • Peptides
  • Polysaccharides
  • Galactosyltransferases
  • UDP-galactose N-acetylgalactosamine peptide beta1,3-galactosyltransferase

Associated data

  • GENBANK/AB084170