A novel fluorescent assay for T-synthase activity

• Authors: Tongzhong Ju, Emory University; Baoyun Xia, Emory University; Rajindra P Aryal, Emory University; Wenyi Wang, Emory University; Yingchun Wang, Emory University; Xiaokun Ding, Emory University; Rongjuan Mi, Emory University; Miao He, Emory University; Richard Cummings, Emory University
• Journal Title: Glycobiology
• Volume: Volume 21, Number 3
• Publisher: Oxford University Press | 2011-03, Pages 352-362
• Type of Work: Article | Post-print: After Peer Review
• Publisher DOI: http://doi.org/10.1093/glycob/cwq168
• Subjects: Biology, Genetics; Chemistry, Biochemistry
• Keywords: Cosmc; fluorescent assay; glycosyltransferase assay; 4-methylumbelliferone; T-synthase

Abstract

Loss of T-synthase (uridine diphosphate galactose:N-acetylgalactosaminyl-α1-Ser/Thr β3galactosyltransferase), a key enzyme required for the formation of mucin-type core 1 O-glycans, is observed in several human diseases, including cancer, Tn syndrome and IgA nephropathy, but current methods to assay the enzyme use radioactive substrates and complicated isolation of the product. Here we report the development of a novel fluorescent assay to measure its activity in a variety of tumor cell lines. Deficiencies in T-synthase activity correlate with mutations in the gene encoding the molecular chaperone Cosmc that is required for folding the T-synthase. This new high-throughput assay allows for facile screening of tumor specimens and other biological material for T-synthase activity and could be used diagnostically.