Macromolecular structures determined by X-ray crystallography are central to modern understanding of biological processes and disease mechanisms and to the design of new medicines. Crystal damage caused by the X-rays themselves and by the cryoprotective procedures used to minimize radiation damage reduces structure resolution and accuracy -- of particular importance to mechanistic studies -- and in unfavorable cases can prevent structure determination. Powerful diffraction and real-space imaging techniques have recently been used to dissect disorder in macromolecular crystals, and provide new tools for understanding this damage. ? ? The long term goals of this project are to characterize the systematics of radiation and cryoprotection-induced disorder using simple model proteins and viruses; to develop new procedures for reducing this disorder so that the full potential of high brilliance synchrotron sources for data collection can be exploited; and to apply these to selected systems of significant biological interest. Radiation damage and damage caused by flash cooling are complex phenomena, and no single factor is likely to produce dramatic improvements. Systematic experiments using X-ray diffraction and imaging, atomic force microscopy and other techniques will evaluate the most important factors, and explore how they may be combined to optimize overall data quality. This approach has significant and realistic potential to impact how macromolecular structures are determined, and thereby to contribute to their use in medical, pharmaceutical, and agricultural applications.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Biophysical Chemistry Study Section (BBCB)
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Edmonds, Charles G
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Cornell University
Other Domestic Higher Education
United States
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Warkentin, Matthew; Hopkins, Jesse B; Haber, Jonah B et al. (2014) Temperature-dependent radiation sensitivity and order of 70S ribosome crystals. Acta Crystallogr D Biol Crystallogr 70:2890-6
Warkentin, Matthew; Hopkins, Jesse B; Badeau, Ryan et al. (2013) Global radiation damage: temperature dependence, time dependence and how to outrun it. J Synchrotron Radiat 20:7-13
Meisburger, Steve P; Warkentin, Matthew; Chen, Huimin et al. (2013) Breaking the radiation damage limit with Cryo-SAXS. Biophys J 104:227-36
Warkentin, Matthew; Badeau, Ryan; Hopkins, Jesse B et al. (2012) Spatial distribution of radiation damage to crystalline proteins at 25-300 K. Acta Crystallogr D Biol Crystallogr 68:1108-17
Warkentin, Matthew; Badeau, Ryan; Hopkins, Jesse B et al. (2012) Global radiation damage at 300 and 260 K with dose rates approaching 1?MGy?s?ยน. Acta Crystallogr D Biol Crystallogr 68:124-33
Hopkins, Jesse B; Badeau, Ryan; Warkentin, Matthew et al. (2012) Effect of common cryoprotectants on critical warming rates and ice formation in aqueous solutions. Cryobiology 65:169-78
Warkentin, Matthew; Badeau, Ryan; Hopkins, Jesse et al. (2011) Dark progression reveals slow timescales for radiation damage between T = 180 and 240?K. Acta Crystallogr D Biol Crystallogr 67:792-803
Soliman, Ahmed S M; Warkentin, Matthew; Apker, Benjamin et al. (2011) Development of high-performance X-ray transparent crystallization plates for in situ protein crystal screening and analysis. Acta Crystallogr D Biol Crystallogr 67:646-56
Kmetko, Jan; Warkentin, Matthew; Englich, Ulrich et al. (2011) Can radiation damage to protein crystals be reduced using small-molecule compounds? Acta Crystallogr D Biol Crystallogr 67:881-93
Warkentin, Matthew; Thorne, Robert E (2010) Slow cooling and temperature-controlled protein crystallography. J Struct Funct Genomics 11:85-9

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