Date of Award
Doctor of Philosophy (PhD)
Chemistry & Biochemistry
Lesley H. Greene
Elucidating the mechanisms of protein folding and unfolding is one of the greatest scientific challenges in basic science. The overarching goal is to predict three-dimensional structures from their amino acid sequences. Understanding the determinants of protein folding and stability can be facilitated through the study of evolutionarily related but diverse proteins. Insights can also be gained through the study of proteins from extremophiles that may more closely resemble the primordial proteins. In this doctoral research, three aims were accomplished to characterize the structure, folding and unfolding behavior within the β-grasp superfamily. We propose that the determinants of structure, stability, and folding are conserved as sequence and interaction patterns in the β-grasp fold. To elucidate key residues, bioinformatics studies were conducted and identified nine structurally conserved amino acids in the core of the B1 domain of protein G (GB1). A network analysis of all long-range interactions in the structure of GB1 revealed the relative significance of each conserved amino acid. Within the β-grasp superfamily, two proteins, GB1 and the small archaeal modifier protein 1 (SAMP1), were investigated to elucidate the key determinants of structural stability at the level of individual interactions. They were subjected to high temperature molecular dynamics simulations and the detailed behavior of each long-range interaction was characterized. The results revealed that in GB1 the most stable region was the C-terminal hairpin and in SAMP1 it was the opposite, the N-terminal hairpin. The folding behavior of SAMP1 was investigated due to its nature as a divergent superfamily member and extremophile. The results revealed that SAMP1 at high ionic strength folds more rapidly than in low ionic strength. These findings clearly indicate that adaption at high salt produces rapid and less-frustrated folding. The results of these research aims provide insight into determinants of the β-grasp fold and the folding and unfolding behavior of two key members. Perhaps the most surprising finding is the presence of a significant number of non-native long-range interactions during unfolding which has largely gone unnoticed in the scientific community and appears to be pivotal.
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Bedford, John T..
"Computational and Experimental Investigation into the Determinants of Protein Structure, Folding, and Stability in the β-Grasp Superfamily"
(2021). Doctor of Philosophy (PhD), Dissertation, Chemistry & Biochemistry, Old Dominion University, DOI: 10.25777/apaz-yv79