Glyco-Engineering Cell Surfaces by Exo-Enzymatic Installation of GlcNAz and LacNAz Motifs

Abstract

Exo-enzymatic glyco-engineering of cell-surface glycoconjugates enables the selective display of well-defined glyco-motifs bearing bioorthogonal functional groups which can be used to study glycans and their interactions with glycan-binding proteins. While the installation of monosaccharides and their derivatives using glycosyltransferase enzymes has rapidly evolved, similar strategies to introduce chemical-reporter functionalized Type 2 LacNAc motifs have not been reported. Herein, we report the chemo-enzymatic synthesis of unnatural UDP-GlcNAc and UDP-GalNAc nucleotide-sugars, and the donor and acceptor substrate tolerance of the human glycosyltransferases B3GNT2 and B4GalT1, respectively, to form derivatized LacNAc moieties. We also demonstrate that B3GNT2 can be used to exo-enzymatically install GlcNAc and GlcNAz onto cell-surface glycans. GlcNAc- or GlcNAz-engineered cells can be further extended by B4GalT1, producing LacNAc or LacNAz-engineered cells. Our glyco-engineering labeling strategy is amenable to different cell types and our work expands the exo-enzymatic glycan editing toolbox to selectively introduce unnatural Type 2 LacNAc motifs.