Shafiqur Rahman M, Qin Wensheng
Department of Biology, Lakehead University, Thunder Bay, ON, Canada.
Department of Microbiology, University of Chittagong, Chittagong, Bangladesh.
Methods Mol Biol. 2019;1954:237-243. doi: 10.1007/978-1-4939-9154-9_18.
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.
糖基转移酶（GTs）是一类重要的酶亚类，通过将糖残基从供体底物转移至受体底物，催化寡糖、多糖和糖缀合物中糖苷键的生物合成。膜相关糖基转移酶在细菌细胞壁多糖的生物合成中起着至关重要的作用。糖基转移酶活性的表征和定量对于多糖生物合成研究、药物靶点开发以及细菌产物生产具有重要意义。在本章中，将介绍用于测量糖基转移酶活性的比色测定法。作用于单糖衍生物的糖基转移酶测定法基于未反应的糖基对硝基苯酚受体的裂解，随后检测对硝基苯酚盐。与磷酸酶偶联并检测无机磷酸盐的糖基转移酶反应适用于大多数糖基转移酶。这些测定法无需使用放射性糖类即可方便地对糖基转移酶活性和动力学进行定量。
