%0 Journal Article
%T A Bacterial Expression Platform for Production of Therapeutic Proteins Containing Human-like O-Linked Glycans.
%A Du T
%A Buenbrazo N
%A Kell L
%A Rahmani S
%A Sim L
%A Withers SG
%A DeFrees S
%A Wakarchuk W
%J Cell Chem Biol
%V 26
%N 2
%D 02 2019 21
%M 30503285
%F 9.039
%R 10.1016/j.chembiol.2018.10.017
%X 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. 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 α-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.