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

TitleA Bacterial Expression Platform for Production of Therapeutic Proteins Containing Human-like O-Linked Glycans
Publication TypeJournal Article
Year of Publication2019
AuthorsDu, T, Buenbrazo, N, Kell, L, Rahmani, S, Sim, L, Withers, SG, DeFrees, S, Wakarchuk, W
JournalCELL CHEMICAL BIOLOGY
Volume26
Pagination203+
Date PublishedFEB 21
ISSN2451-9448
Abstract

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 beta 1,3-galactosyl transferase and UDP-Glc (NAc)-4-epimerase, both from Campylobacter jejuni, and a disulfide bond isomerase of bacterial or human origin. The effectiveness of this two-plasmid synthetic operon system has been tested on three proteins with therapeutic potential: the native and an engineered version of the naturally O-glycosylated human interferon alpha-2b, as well as human growth hormone with one engineered site of glycosylation. Having established proof of principle for the addition of the core-1 glycan onto proteins, we are now developing this system as a platform for producing and modifying human protein therapeutics with more complex O-glycan structures in E. coli.

DOI10.1016/j.chembiol.2018.10.017