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Title: An investigation of protein-protein interactions during renaturation
Author: Ho, J. G. S.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2003
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The aim of the research presented in this thesis is to determine the applicability of measuring protein-protein interactions (the second virial coefficient) to protein renaturation (refolding). As shown subsequently in this thesis, the second virial coefficient provides an experimentally accessible link between empirical observations with molecular and thermodynamic insight into protein aggregation during renaturation. This thesis is the first instance that light scattering has been used to probe protein-protein interactions during the renaturation, and describes the theory, development and validation of this technique. Using this new approach, four major themes on protein-protein interactions and protein renaturation are explored. The first theme is the influence of surface properties of proteins (hydration and surface hydrophobicity) and how they affect protein-protein interactions. Secondly, a quantitative method for assessing both the sign and magnitude of protein-protein interactions in relation to empirical observations of protein aggregation and protein solubility are presented. This subsequently leads to the issue of the impact of protein-protein interactions and water structure on renaturation. A simple technique to assess water structure conducive to protein renaturation is also introduced. The final theme is to present the measure of second virial coefficients as a simple metric for assessing the propensity of inclusion body proteins to aggregate during renaturation. A variety of inclusion bodies are screened and relationships between protein molecular weight, hydropathicity, and fractional yield are presented. The key outcome of this thesis is the demonstration of the utility of the second virial coefficient as a tool to rationally screen renaturation conditions for new proteins. It will help our understanding of why certain conditions promote renaturation in preference to aggregation. Importantly, this technique provides a useful tool for determining refolding strategy, provides a link between empirical practice and fundamental thermodynamics and demonstrates an initial relationship between protein sequence and structure for the protein production/refolding process.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available