Investigation of the conformational energetics of bacteriorhodopsin
Abstract
Membrane proteins are exist in lipid bilayers and perform various biochemical reactions that are essential for biological function. Many aspects of the forces that determine their conformation remain shrouded in mystery. This dissertation summarizes a number of studies that focus on the kinetics and thermodynamics of the α-helical membrane protein folding. I first describe the development of "pulse proteolysis" as a novel probe for membrane protein folding using the unfolding of H. halobium bacteriorhodopsin (bR) by SDS as a model system. I next describe its application to study the refolding of bR, which lead to the discovery of an unknown factor that controls the kinetics and thermodynamics of bR folding. I then assess the conformational relaxation kinetics of bR under various experimental conditions and utilize the results to develop an approach to investigate the conformational energetics of bR. Application of this approach to a series of bR mutants has clarified the effects of mutation on the folding and unfolding kinetics. Moreover, the relaxation kinetics of the mutants has provided insight into the rate limiting transition state for folding. Together, these findings unify a substantial body of research on the conformational energetics of bR and provide new tools for the future investigations of α-helical membrane protein folding.
Degree
Ph.D.
Advisors
Park, Purdue University.
Subject Area
Biochemistry|Physical chemistry|Biophysics
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