1 Department of Chemistry and Biochemistry, Concordia University, 7141 Sherbrooke Street West, Montreal, Quebec, Canada H4B 1R6.
2 Department of Biology, Centre for Applied Synthetic Biology, and Centre for Structural and Functional Genomics, Concordia University, 7141 Sherbrooke Street West, Montreal, Quebec, Canada H4B 1R6.

Enzymatic Synthesis of a Fluorogenic Reporter Substrate and the Development of a High-Throughput Assay for Fucosyltransferase VIII Provide a Toolkit to Probe and Inhibit Core Fucosylation.

Biochemistry. 2020 Jun 01;:

Authors: Soroko M, Kwan DH

Abstract

We report a straightforward enzymatic synthesis of the 4-methylumbelliferyl glycoside of a complex-type oligosaccharide substrate for core fucosylation. We demonstrate the use of this synthetic glycoconjugate in a newly developed enzyme assay to probe the activity and inhibition of fucosyltransferase VIII, which catalyzes the core fucosylation of N-glycans on eukaryotic glycoproteins. In this fucosyltransferase assay, we use the fluorogenic probe and a specific glycosidase in a sequentially coupled enzyme reaction to distinguish an unmodified 4-methylumbelliferyl oligosaccharide probe from a fucosylated probe. Our findings show that this strategy is very sensitive and specific in its detection of enzyme activity and can even be used for analyzing impure tissue lysate samples.

PMID: 32441090 [PubMed - as supplied by publisher]