This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. X-ray cryocrystallography is the primary tool used to determine the 3D molecular structure of proteins and other biological macromolecules, and to date roughly 50,000 unique structures having been determined. We are investigating several factors that affect the quality of the structural information that is obtained, and attempting to develop improved protocols. Specific questions being investigated include: Why does cooling protein crystals disorder them? How do X-rays damage the crystal, and how do low temperatures reduce this damage? How do the method by which a protein crystal is cooled and the cooling rate affect its structure? How does protein structure evolve with temperature, especially in the vicinity of the glass transition? Can variable temperature data collection shed new light on protein dynamics? Our planned work is based on two significant discoveries that have allowed us to successfully cool crystals at rates from 0.1 K/s to 100,000 K/s, and to successfully collect structural quality data sets at arbitrary temperatures between 300 K and 100 K, in both cases without the use of penetrating cryoprotectants.
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