14] Synthetic neoglycoconjugates in glycosyltransferase assay and purification

Abstract

This chapter discusses the use of synthetic neoglycoconjugates for the assay and isolation of N-acetylglucosaminyltransferase V [GnT-V; EC 2.4.1.155, α(1,3(6)-mannosylglycoprotein β-1,6-N-acetylglucosaminyl transferase], an enzyme involved in the branching of asparagine-linked oligosaccharides. Synthetic neoglycoconjugates are useful tools for both the isolation and assay of glycosyltransferase enzymes. The main advantage in using a synthetic approach is that a variety of natural as well as unnatural oligosaccharide structures can be prepared. These will be free of cross reacting acceptors within the detection limits of chemical characterization, typically nuclear magnetic resonance (NMR) spectroscopy, high-performance liquid chromatography (HPLC), or mass spectrometry. In addition, incorporation levels can be controlled and a variety of proteins can be used for conjugation. In the laboratories, the most commonly used protein for conjugate preparation is the readily available bovine serum albumin (BSA).