388271 Effect of Glycosylation on the Precipitation of Antibodies By Non Ionic Polymers

Wednesday, November 19, 2014
Galleria Exhibit Hall (Hilton Atlanta)
Daniel Forciniti, Cbe, Missouri University of Science and Technology, Rolla, MO

Post-translational modification of many proteins results in the attachment of various oligosaccharides to the peptide chain. The biological activity and stability of the protein are often affected by the degree of glycosylation. Glycosylation is relatively common, with antibodies and integral membrane proteins being two important protein types undergoing this post-translational modification.

     Proteins glycosylated in vitro or in transgenic plants show greater diversity than their in vivo counterpart and this diversity may differ from batch to batch.  A straightforward approach to eliminate this problem is to shut down the glycosylation machinery all together. Although the heterogeneity problem is of course eliminated, the fact that the native and transgenic proteins are not exactly the same (in spite of both having the same biological activity) raises questions about the stability of the non-glycosylated antibody. 

     We have studied the stability of a fully glycosylated and a non-glycosylated human antibody expressed in corn. The glycosylated protein was a mixture of fully glycosylated, GlcNac, and non-glycosylated proteins (isoelectric point: ~9.5).  We were particularly concerned about the effect that glycosylation may have on the solubility of the protein.  Therefore, we conducted a solubility study of both antibodies by inducing precipitation by adding poly(ethylene)glycol (PEG).  A resolution V experimental design that included as parameters temperature, pH, polymer molecular weight and level of protein glycosylation was pursued.  Our studies clearly show that glycosylation greatly affects the stability of the antibody and identifies non-trivial two parameters interactions.   An  anomalous behavior was observed as the solubility of the antibody does not go through a minimum at its isoelectric point.  The results of the model were statistically fitted and the parameters compared with existing theoretical predictions.

Extended Abstract: File Not Uploaded
See more of this Session: Poster Session: Bioengineering
See more of this Group/Topical: Food, Pharmaceutical & Bioengineering Division