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bioRxiv · 10.1101/322461

Bacterial Glycosyltransferase-mediated Cell-surface Chemoenzymatic Glycan Editing: Methods and Applications

Hong, S.·Shi, Y.·Wu, N. C.·Grande, G.·Douthit, L.·Wilson, I. A.·Xie, J.·Wu, P.
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Abstract

AbstractChemoenzymatic glycan editing that modifies glycan structures directly on the cell surface has emerged as a complementary tool to metabolic oligosaccharide engineering. In this article, we report the discovery that three bacterial enzymes--Pasteurella multocida 2-3-sialyltransferase M144D mutant (Pm2,3ST-M144D), Photobacterium damsel 2-6-sialyltransferase (Pd2,6ST) and Helicobacter mustelae 1-2-fucosyltransferase (Hm1,2FT)--can serve as highly efficient tools for cell-surface glycan editing. Among these three enzymes, the two sialyltransferases were also found to be tolerant to large substituents introduced to the C-5 position of the cytidine monophosphate N-acetylneuraminic acid donor, including biotin and fluorescent dyes. Combining these enzymes with our previously discovered Helicobacter pylori 1-3-FT, we developed a live cell-based assay to probe host-cell glycan-mediated influenza A virus (IAV) infection including both wild-type and mutant strains of human H1N1 and H3N2 influenza subtypes. At high SiaNAc2-6-Gal levels, the ability of a viral strain to induce the host cell death is positively correlated with the SiaNAc2-6-Gal binding affinity of its haemagglutinin. Surprisingly, the creation of sLeX on the host cell surface via in situ 1-3-Fuc editing also exacerbated the killing induced by several wild-type IAV strains as well as a mutant known as HK68-MTA. Structural alignment of HAs from the wild-type HK68 and HK68-MTA revealed the formation of a putative hydrogen bond between Trp222 of HA-HK68-MTA and the C-4 hydroxyl group of the 1-3-linked fucose of sLeX. This interaction is likely to be responsible for the better binding affinity of HA-HK68-MTA to sLeX and accordingly the enhanced host-cell killing compared with the wild-type HK68.

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Hong, S., Shi, Y., Wu, N. C., Grande, G., Douthit, L., Wilson, I. A., Xie, J., Wu, P.. 2018-05-15. Bacterial Glycosyltransferase-mediated Cell-surface Chemoenzymatic Glycan Editing: Methods and Applications. https://doi.org/10.1101/322461

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