Generation of Recombinant N-Linked Glycoproteins in E. coli

Abstract

The production of N-linked recombinant glycoproteins is possible in a variety of biotechnology host cells, and more recently in the bacterial workhorse, Escherichia coli. This methods chapter will outline the components and procedures needed to produce N-linked glycoproteins in E. coli, utilizing Campylobacter jejuni glycosylation machinery, although other related genes can be used with minimal tweaks to this methodology. To ensure a successful outcome, various methods will be highlighted that can confirm glycoprotein production to a high degree of confidence, including the gold standard of mass spectrometry analysis.

Keywords: E. coli; Glycoprotein validation; Glycosylation; N-Linked glycoproteins; Posttranslational modifications.