In vacuo glycation of enzymes: A novel approach for increasing enzyme stability

Abstract

A novel approach for the thermostabilization of proteins was investigated. It is well established that proteins that are naturally highly glycosylated show an increased resistance to inactivation at high temperatures. However, non-enzymatic attachment of carbohydrate to proteins, otherwise known as protein glycation, under aqueous conditions is very difficult and has not been used as a general approach for increasing the thermostability of proteins. In the present study, advantage was taken of the in vacuo protein glycation procedure recently developed by Kaplan and his co-workers by which proteins can be extensively glycated in the absence of water and chemical reagents. In this procedure, reducing sugars react with the epsilon-amino groups of lysine side-chains to form a stable ketoamine derivative. The primary objective of the research presented in this thesis, was to determine the extent to which the thermostability of proteins can be increased by in vacuo glycation with reducing monosaccharides such as glucose. Another objective of this study was to investigate possible practical applications of this thermostabilization technology. (Abstract shortened by UMI.)