Labeling glycans on living cells by a chemoenzymatic glycoengineering approach.

Ruben T Almaraz,Yanhong Li

doi:10.1242/bio.021600

Abstract

ABSTRACTStructural glycobiology has traditionally been a challenging field due to a limited set of tools available to investigate the diverse and complex glycan molecules. However, we cannot ignore that glycans play critical roles in health as well as in disease, and are present in more than 50% of all proteins and on over 80% of all surface proteins. Chemoenzymatic glycoengineering (CGE) methods are a powerful set of tools to synthesize complex glycans, but the full potential of these methods have not been explored in cell biology yet. Herein, we report the labeling of live Chinese hamster ovary (CHO) cells by employing three highly specific glycosyltransferases: a sialyltransferase, a galactosyltransferase, and an N-acetyl-glucosaminyl transferase. We verified our results by bio-orthogonal blots and further rationalized them by computational modeling. We expect CGE applications in cell biology to rise and their implementation will assist in structural-functional discoveries in glycobiology. This research will contribute to this effort.

Highlights

The glycobiology of any given cell plays essential roles in many aspects of the cell daily functions
Metabolic glycoengineering (MGE), which consists in the ability to implement non-natural monosaccharides analogs, was introduced in 1992 and since a plethora of publications utilizing MGE technology has emerged (Nischan and Kohler, 2016)
Chemoenzymatic glycoengineering (CGE) methods became an important part of the toolbox to visualize and study the glycome (Mbua et al, 2013; Zheng et al, 2011)

Summary

Introduction

The glycobiology of any given cell plays essential roles in many aspects of the cell daily functions. We incubated the CHO K1, Lec2, Lec8 and Lec12 cells with PmST_M144D and different concentration of CMP-Neu5Az 1 (Fig. 1A) at 37°C for 10 and 20 min in a DPBS-Tris buffer.

Results

Conclusion