Commissariat a l'Energie Atomique et aux Energies Alternatives (CEA), Grenoble (France); Univ. of Grenoble (France); Centre National de la Recherche Scientifique (CNRS), Grenoble (France). Inst. de Biologie Structurale (IBS)
Commissariat a l'Energie Atomique et aux Energies Alternatives (CEA), Grenoble (France); Univ. of Grenoble (France); Centre National de la Recherche Scientifique (CNRS), Grenoble (France). Inst. de Biologie Structurale (IBS); Inst. Laue-Langevin (ILL), Grenoble (France)
Univ. of Oxford (United Kingdom)
European Synchrotron Radiation Facility (ESRF), Grenoble (France)
Univ. of California, San Francisco, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); SLAC National Accelerator Lab., Menlo Park, CA (United States). Stanford Synchrotron Radiation Lightsource (SSRL)
Univ. of Notre Dame, IN (United States). Notre Dame Radiation Lab. (NDRL)
Radiation damage limits the accuracy of macromolecular structures in X-ray crystallography. Cryogenic (cryo-) cooling reduces the global radiation damage rate and, therefore, became the method of choice over the past decades. The recent advent of serial crystallography, which spreads the absorbed energy over many crystals, thereby reducing damage, has rendered room temperature (RT) data collection more practical and also extendable to microcrystals, both enabling and requiring the study of specific and global radiation damage at RT. Here, we performed sequential serial raster-scanning crystallography using a microfocused synchrotron beam that allowed for the collection of two series of 40 and 90 full datasets at 2- and 1.9-Å resolution at a dose rate of 40.3 MGy/s on hen egg white lysozyme (HEWL) crystals at RT and cryotemperature, respectively. The diffraction intensity halved its initial value at average doses (D1/2) of 0.57 and 15.3 MGy at RT and 100 K, respectively. Specific radiation damage at RT was observed at disulfide bonds but not at acidic residues, increasing and then apparently reversing, a peculiar behavior that can be modeled by accounting for differential diffraction intensity decay due to the nonuniform illumination by the X-ray beam. Specific damage to disulfide bonds is evident early on at RT and proceeds at a fivefold higher rate than global damage. The decay modeling suggests it is advisable not to exceed a dose of 0.38 MGy per dataset in static and time-resolved synchrotron crystallography experiments at RT. This rough yardstick might change for proteins other than HEWL and at resolutions other than 2 Å.
de la Mora, Eugenio, et al. "Radiation damage and dose limits in serial synchrotron crystallography at cryo- and room temperatures." Proceedings of the National Academy of Sciences of the United States of America, vol. 117, no. 8, Feb. 2020. https://doi.org/10.1073/pnas.1821522117
de la Mora, Eugenio, Coquelle, Nicolas, Bury, Charles S., Rosenthal, Martin, Holton, James M., Carmichael, Ian, Garman, Elspeth F., Burghammer, Manfred, Colletier, Jacques-Philippe, & Weik, Martin (2020). Radiation damage and dose limits in serial synchrotron crystallography at cryo- and room temperatures. Proceedings of the National Academy of Sciences of the United States of America, 117(8). https://doi.org/10.1073/pnas.1821522117
de la Mora, Eugenio, Coquelle, Nicolas, Bury, Charles S., et al., "Radiation damage and dose limits in serial synchrotron crystallography at cryo- and room temperatures," Proceedings of the National Academy of Sciences of the United States of America 117, no. 8 (2020), https://doi.org/10.1073/pnas.1821522117
@article{osti_1608636,
author = {de la Mora, Eugenio and Coquelle, Nicolas and Bury, Charles S. and Rosenthal, Martin and Holton, James M. and Carmichael, Ian and Garman, Elspeth F. and Burghammer, Manfred and Colletier, Jacques-Philippe and Weik, Martin},
title = {Radiation damage and dose limits in serial synchrotron crystallography at cryo- and room temperatures},
annote = {Radiation damage limits the accuracy of macromolecular structures in X-ray crystallography. Cryogenic (cryo-) cooling reduces the global radiation damage rate and, therefore, became the method of choice over the past decades. The recent advent of serial crystallography, which spreads the absorbed energy over many crystals, thereby reducing damage, has rendered room temperature (RT) data collection more practical and also extendable to microcrystals, both enabling and requiring the study of specific and global radiation damage at RT. Here, we performed sequential serial raster-scanning crystallography using a microfocused synchrotron beam that allowed for the collection of two series of 40 and 90 full datasets at 2- and 1.9-Å resolution at a dose rate of 40.3 MGy/s on hen egg white lysozyme (HEWL) crystals at RT and cryotemperature, respectively. The diffraction intensity halved its initial value at average doses (D1/2) of 0.57 and 15.3 MGy at RT and 100 K, respectively. Specific radiation damage at RT was observed at disulfide bonds but not at acidic residues, increasing and then apparently reversing, a peculiar behavior that can be modeled by accounting for differential diffraction intensity decay due to the nonuniform illumination by the X-ray beam. Specific damage to disulfide bonds is evident early on at RT and proceeds at a fivefold higher rate than global damage. The decay modeling suggests it is advisable not to exceed a dose of 0.38 MGy per dataset in static and time-resolved synchrotron crystallography experiments at RT. This rough yardstick might change for proteins other than HEWL and at resolutions other than 2 Å.},
doi = {10.1073/pnas.1821522117},
url = {https://www.osti.gov/biblio/1608636},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
issn = {ISSN 0027-8424},
number = {8},
volume = {117},
place = {United States},
publisher = {National Academy of Sciences},
year = {2020},
month = {02}}
SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States)
Sponsoring Organization:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22); National Institutes of Health (NIH); National Science Foundation (NSF)
Grant/Contract Number:
AC02-76SF00515; FC02-04ER15533; AC02-05CH11231
OSTI ID:
1608636
Journal Information:
Proceedings of the National Academy of Sciences of the United States of America, Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Issue: 8 Vol. 117; ISSN 0027-8424