Glycosylation is one of the most common post-translational modifications of proteins, and proteins affected by this modification can have altered conformation, solubility, activity, and antigenicity. N-glycoproteins are a class of glycoproteins characterized by the attachment of a chain of sugars (glycans) to one or more asparagine residues through an amide bond. The study of N-glycoproteins has been hindered by the difficulty in obtaining homogeneous samples for testing their biological effects because natural samples of glycoproteins appear as complex mixtures of proteins with many different glycosylation patterns. Here, we describe our efforts toward chemospecific glycosylation of a recombinantly generated protein to produce a homogeneous sample of N-glycoprotein. The protein is produced recombinantly by E. coli which have been modified to insert a non-canonical amino acid containing a reactive thioester side chain at a specified internal site. This thioester handle can then undergo a chemoselective reaction with a modified glycan to give a cysteine-bridged N-glycoprotein. Because natural sources of glycoproteins contain a high degree of heterogeneity, this method of chemical synthesis is attractive because of its ability to generate homogeneous samples which can be used in biological research.

Synthesis of homogeneous cysteine-bridged N-glycoproteins via native chemical glycoligation

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

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