UCSF

Protein folding converts a disordered polymer to a globular structure, reducing many conformational degrees of freedom and incurring a significant conformational entropy penalty. However, residual conformational entropy is retained in a protein’s folded native state. Subtle changes between positions within the native state, mostly from sidechains, alters residual conformational entropy, leading to differences in binding affinity and allosteric communication. While NMR has provided measurements of conformational entropy, these measurements do not provide information on where this entropy is coming from, such as if this is coming from a sidechain moving harmonically or anharmonically. However, we can take advantage of the fact that X-ray crystallography and CryoEM experimental data capture the conformational ensemble allowing us to measure the motion of residues and their atomistic structure. This provides an unparalleled platform to answer how, where, and why conformational entropy. The first chapter of this thesis presents the improvements to the qFit algorithm. This algorithm allows for the automated modeling of multiple conformations per residue across a protein for high resolution X-ray crystallography or cryo-EM. We present algorithm improvements including the ability to run the program on a laptop. This algorithm was the basis for much of the future work of my thesis. The second chapter contains the findings of the relationship between conformational heterogeneity and ligand binding. Using qFit, we identified the changes in conformational heterogeneity between matched bound and unbound high resolution X-ray structures. We identified a reciprocal relationship upon ligand binding where as binding site residues become more rigid, distant residues become more flexible, indicating an entropic compensation. The third chapter contains my future outlook on the questions and techniques to probe conformational entropy mechanism. This chapter includes how to integrate new modeling techniques to understand how different motions of residues lead to differences in conformational entropy.