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Glycosyltransferase Activity Assay Using Colorimetric Methods

  • M. Shafiqur Rahman
  • Wensheng QinEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1954)

Abstract

The glycosyltransferases (GTs) are an important subclass of enzymes that catalyze the biosynthesis of glycosidic bonds in oligosaccharides, polysaccharides and glycoconjugates by transferring a sugar residue from a donor substrate to an acceptor substrate. The membrane-associated GTs play a vital role in the biosynthesis of bacterial cell-wall polysaccharides. Characterization and quantification of GT activities is important for studies of biosynthesis of polysaccharides, drug target development, and production of bacterial products. In this chapter, colorimetric assays for the measurement of GT activities will be presented. Assays for GTs acting on monosaccharide-derivatives are based on the cleavage of unreacted glycosyl-p-nitrophenol acceptors followed by detection of p-nitrophenolate. GT reactions coupled with phosphatases and detection of inorganic phosphate are suitable for most GTs. These assays permit convenient quantification of GT activities and kinetics without the use of radioactive sugars.

Key words

Bacterial glycosyltransferases Substrate specificity Polysaccharide glycosyltransferases Colorimetric assay Phosphate-coupled assay 

Notes

Acknowledgments

This work was supported by Natural Sciences and Engineering Research Council of Canada (RGPIN-2017-05366) to W. Q.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of BiologyLakehead UniversityThunder BayCanada
  2. 2.Department of MicrobiologyUniversity of ChittagongChittagongBangladesh

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