Biology subjects

Boddington, M. E.

Publications and source records attributed to Boddington, M. E..

2 recordsLinked to original sources

bioRxiv2023

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

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.

biochemistry↗

bioRxiv2023

One-Step Selective Labeling of Native Cell-Surface Sialoglycans by Exogenous α2,8-Sialylation

Exo-enzymatic glycan labeling strategies have emerged as versatile tools for efficient and selective installation of glycan terminal motifs onto live cell-surfaces. Through employing specific enzymes and nucleotide sugar probes, cells can be equipped with defined glyco-epitopes for modulating cell function or selective visualization and enrichment of glycoconjugates. Here, we identify Campylobacter jejuni sialyltransferase Cst-II I53S as a tool for cell-surface glycan modification, expanding the exo-enzymatic labeling toolkit to include installation of 2,8-disialyl epitopes. Labeling with Cst-II was achieved with biotin- and azide-tagged CMP-Neu5Ac derivatives on a model glycoprotein and on native sialylated cell-surface glycans across a panel of cell lines. The introduction of modified Neu5Ac derivatives onto cells by Cst-II was also retained on the surface for 6 h. By examining the specificity of Cst-II on cell surfaces, it was revealed that the 2,8-sialyltransferase primarily labeled N-glycans, with O-glycans labeled to a lesser extent, and there was an apparent preference for 2,3-linked sialosides. This approach thus broadens the scope of tools for selective exo-enzymatic labeling of native sialylated glycans and is highly amenable for construction of cell-based arrays.

biochemistry↗