Preparing for a warmer future: Conformational ensembles, dynamics and interactions from variable temperature crystallography

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An understanding of the 3D structure of proteins and other molecules of life is essential to understanding their function and to the design of new medicines. Most atomic-resolution protein structures are determined using X-ray crystallography. But while proteins function at biological (generally, near room) temperature, in the last decade nearly all X-ray crystallography has been performed on protein crystals cooled to cryogenic temperatures, where many important details of structure and dynamics may be compromised. This project will explore the wealth of additional information that may be accessible using protein crystals with liquid solvent. Structural data sets collected at temperatures between ~330 and 200 K will allow exploration of how protein conformational ensembles and energy landscapes evolve, and of the signatures of and interactions leading to cold denaturation. Ultra-fast and variable-rate cooling will be used to characterize timescales of large and/or highly correlated protein motions, and to better preserve room temperature structure for examination at 100 K. The project builds on recent developments in sample handling and data collection and in refinement and analysis tools, and aims to substantially extend these developments. Broader Impacts This project will enable a broadening in the scope of crystallographic studies of proteins, and contribute to a comprehensive understanding of the relation between protein structure and function, relevant to the understanding biology and to the development of new biologically active compounds. It builds on past efforts in fundamental studies and methods development that have led to commercial tools used by crystallographers worldwide. Undergraduate and graduate student researchers will receive cross-disciplinary training in advanced methods and will be critical in propagating these methods to the broader structural biology community. This research program is complemented by vigorous efforts in undergraduate introductory course curriculum reform aimed at improving the retention and subject mastery of life science and engineering students. It is also complemented by a broad effort in STEM teacher recruiting and training and in institutional change that provides a model for how research-focused STEM departments can address critical teacher shortages in our middle and high schools.