Chemical glycosylation: new insights on the interrelation between protein structural mobility, thermodynamic stability, and catalysis

Abstract

Chemical protein glycosylation was employed to sequentially modulate the structural dynamics of the serine protease alpha-chymotrypsin as evidenced from amide H/D exchange kinetics. The reduction in alpha-CT's structural dynamics at increasing glycan molar contents statistically correlated with the increased thermodynamic stability (T(m)) and reduced rate of enzyme catalysis (k(cat)) exhibited by the enzyme upon chemical glycosylation. Temperature-dependent experiments revealed that native-like structural dynamics and function could be restored for the glycosylated conjugates at temperature values close to their thermodynamic stability suggesting that the concept of "corresponding states" can be extended to glycoproteins. These results demonstrate the value of chemical glycosylation as a tool for studying the role of protein structural dynamics on protein biophysical properties; e.g. enzyme stability and function.