Dialing-Up Distal Sites in Proteins: The Search for the Mechanical Consequences of Thermodynamic Coupling in eglin C

Boyer, Joshua Alan

Dialing-Up Distal Sites in Proteins: The Search for the Mechanical Consequences of Thermodynamic Coupling in eglin C Public Deposited

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March 20, 2019

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Boyer, Joshua Alan
- Affiliation: School of Medicine, Department of Biochemistry and Biophysics

Abstract

Proteins have evolved to exploit long-range dynamic and structural effects as a means of regulating function. Understanding this communication between distal sites in proteins is therefore vital to our comprehension of such phenomena as allostery. Structural effects have been suggested but are insufficient for conclusively describing distal-site communication. We believe that understanding protein dynamics may be imperative to comprehending these remote effects. The intent of this dissertation and work herein, is to elucidate the bases underlying thermodynamic coupling/non-additivity, one such long-range effect. The absence of ligands or products in protein folding studies provides a simple background for examining this most basic communication, non-additivity. However, traditional coupling-measuring methods may be too insensitive to report on the idiosyncratic behavior linking distal sites. To increase sensitivity, we have developed hydrogen exchange (HX) techniques that employ NMR's atomic resolution to standard methodologies of double mutant cycles. Due to the large number of probes and their ability to report on equilibria that do not require global unfolding transitions, this method is more likely to perceive underlying dynamic bases than its traditional counterparts. Similarly, to increase the scope and sensitivity of dynamic probes, we applied aromatic 13C relaxation experiments. Together, these methods have identified the co-localization of dynamic and thermodynamic responses in eglin c, further insinuating a relationship between motion and non-additivity.

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