Glycobiology Advance Access first published online on June 8, 2006
This version published online on August 4, 2006
Glycobiology, doi:10.1093/glycob/cwl008
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1 Department of Biochemistry and Molecular Biology Oklahoma Center for Medical Glycobiology University of Oklahoma Health Sciences Center Oklahoma City, Oklahoma 73104
* To whom correspondence should be addressed. The common O-glycan core structure in animal glycoproteins is the core 1 disaccharide Gal
Received February 27, 2006
Revised May 25, 2006
Accepted May 26, 2006
Article
Identification of Core 1 O-Glycan T-synthase from Caenorhabditis elegans
Tongzhong Ju 1, Qinlong Zheng 1, and Richard D. Cummings 1 *
Richard D. Cummings, E-mail: richard-cummings{at}ouhsc.edu
Abstract 
1-3GalNAc
1-Ser/Thr, which is generated by addition of Gal to GalNAc1-Ser/Thr by core 1 UDP-Gal:GalNAc1-Ser/Thr 1,3-galactosyltransferase (core 1 3-Gal-T or T-synthase, EC2.4.1.122)2. Although O-glycans play important roles in vertebrates, much remains to be learned from model organisms such as the free-living nematode Caenorhabditis elegans, which offer many advantages in exploring O-glycan structure/function. Here we report the cloning and enzymatic characterization of T-synthase from C. elegans (Ce-T-synthase). A putative C. elegans gene for T-synthase, C38H2.2, was identified in GenBank by a BlastP search using the human T-synthase protein sequence. The full-length cDNA for Ce-T-synthase, which was generated by PCR using a C. elegans cDNA library as the template, contains 1,170 bp including the stop TAA. The cDNA encodes a protein of 389 amino acids with typical type-II membrane topology and a remarkable 42.7% identity to the human T-synthase. Ce-T-synthase has 7 Cys residues in the lumenal domain including 6 conserved Cys residues in all of the orthologs. The Ce-T-synthase has 4 potential N-glycosylation sequons, whereas the mammalian orthologs lack N-glycosylation sequons. Only one gene for Ce-T-synthase was identified in the genome-wide search and it contains 8 exons. Promoter analysis of the Ce-T-synthase using green fluorescent protein constructs show that the gene is expressed at all developmental stages and appears to be in all cells. Unexpectedly, only minimal activity was recovered in the recombinant, soluble Ce-T-synthase secreted from a wide variety of mammalian cell lines, whereas robust enzyme activity was recovered in the soluble Ce-T-synthase expressed in Hi-5 insect cells. Vertebrate T-synthase requires the molecular chaperone Cosmc, but our results show that Ce-T-synthase does not require Cosmc, and might require invertebrate-specific factors for formation of the optimally active enzyme. These results show that the Ce-T-synthase is a functional ortholog to the human T-synthase in generating core 1 O-glycans and opens new avenues to explore O-glycan function in this model organism.
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