Assay Methods for the Glycosyltransferases Involved in Synthesis of Bacterial Polysaccharides.

Glycans play many important roles in bacterial biology and the complexity of the glycan structures requires biochemical assays in place to help characterize the biosynthetic pathways. Our focus has been on the use of enzymes from pathogens which make molecular mimics of host glycans. We have been examining glycosyltransferases that make strategic linkages in biologically active glycans which can be also exploited for potential therapeutic glycoconjugate synthesis.

A Bacterial Expression Platform for Production of Therapeutic Proteins Containing Human-like O-Linked Glycans.

We have developed an Escherichia coli strain for the in vivo production of O-glycosylated proteins. This was achieved using a dual plasmid approach: one encoding a therapeutic protein target, and a second encoding the enzymatic machinery required for O-glycosylation. The latter plasmid encodes human polypeptide N-acetylgalactosaminyl transferase as well as a β1,3-galactosyl transferase and UDP-Glc(NAc)-4-epimerase, both from Campylobacter jejuni, and a disulfide bond isomerase of bacterial or human origin.