UCSF

The large scale motions that are necessary for the function of many proteins are controlled by atomic level fluctuations. Understanding what causes these fluctuations and how they are translated to the larger motions is essential for understanding the basic mechanism of many biological systems. The ability to control the fluctuations in a predictable way would significantly improve our rational drug and protein design efforts. But, often the important fluctuation that control conformational of proteins are not known. This is in part due to lack of experimental techniques that bridge these scales. Furthermore, computational efforts to meet this need are a work in progress. This thesis describes efforts to understand the mechanistic basis of large-scale motions in different biological systems by developing and applying new tools for the measurement of conformational dynamics. We probed the conformational dynamics of the enzyme CypA using X-ray free electron lasers (XFELs) to validate a known allosteric pathway of the enzyme with this radiation damage free technique. We studied differences in conformation dynamics and allosteric ligand accessibility in αI-domain containing integrins using NMR and room temperature crystallography. By comparing the conformational heterogeneity of two homologous integrins, we have found evidence that the conformational landscape critically influences their ability to bind allosteric modulators. Finally, we visualized the solvent in the influenza M2 proton channel using XFELs in order to understand its mechanism of proton conduction. Taken together, our work highlights the essential role that conformational heterogeneity plays in the function of disparate biological systems, including the atomic-level motions that enable allosteric control of enzyme activity in CypA, the dependence of ligand binding on the conformational heterogeneity of LFA-1, and the solvent conformational heterogeneity that allows proton conduction through the M2 channel. This work is a significant advance towards a mechanistic understanding of the basis of conformational dynamics in the systems described, and will enable future work to manipulate these biological systems to fight disease and improve human health.