KINETICS AND THERMODYNAMICCS OF PROTEIN UNFOLDING

Abstract

The thermodynamics of protein denaturation due to heat or chemical denaturants is not well understood despite decades of study. Modern approaches to elucidate the process of protein folding such as fluorescent spectroscopy provide a reliable approach to examine the relationship between protein structure, conformational stability, and the denaturation process. There is a myriad of factors that affect the conformational stability of a protein, including hydrogen bonding, Van-der-Waals forces, proline isomerization, and the protein environment. The manipulation of the protein environment through heat or chaotropic denaturants destabilizes the protein and triggers unfolding through modification of the protein structure. In this study, heat, urea, and guanidine hydrochloride (GdmCl) were used to study the unfolding of cytochrome C (Cyt C), cytochrome P450 reductase (CPR) and bovine serum albumin (BSA). The exposure of the proteins to the different denaturants suggests that different proteins have differing conformational stability. In addition, the unfolding curves generated indicate that unfolding is complex and does not occur in a simple two-state mechanism is conceptualized of small proteins. This study also demonstrates the fluorescence quenching phenomenon when BSA is denatured by urea and GdmCl.