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Instrumentation and experimental procedures for robust collection of X‐ray diffraction data from protein crystals across physiological temperatures
Author(s) -
Doukov Tzanko,
Herschlag Daniel,
Yabukarski Filip
Publication year - 2020
Publication title -
journal of applied crystallography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.429
H-Index - 162
ISSN - 1600-5767
DOI - 10.1107/s1600576720013503
Subject(s) - protein crystallization , diffraction , synchrotron radiation , crystallography , protein data bank , x ray crystallography , synchrotron , x ray , resolution (logic) , atmospheric temperature range , protein structure , chemistry , materials science , crystallization , physics , optics , computer science , thermodynamics , biochemistry , organic chemistry , artificial intelligence
Traditional X‐ray diffraction data collected at cryo‐temperatures have delivered invaluable insights into the three‐dimensional structures of proteins, providing the backbone of structure–function studies. While cryo‐cooling mitigates radiation damage, cryo‐temperatures can alter protein conformational ensembles and solvent structure. Furthermore, conformational ensembles underlie protein function and energetics, and recent advances in room‐temperature X‐ray crystallography have delivered conformational heterogeneity information that can be directly related to biological function. Given this capability, the next challenge is to develop a robust and broadly applicable method to collect single‐crystal X‐ray diffraction data at and above room temperature. This challenge is addressed herein. The approach described provides complete diffraction data sets with total collection times as short as ∼5 s from single protein crystals, dramatically increasing the quantity of data that can be collected within allocated synchrotron beam time. Its applicability was demonstrated by collecting 1.09–1.54 Å resolution data over a temperature range of 293–363 K for proteinase K, thaumatin and lysozyme crystals at BL14‐1 at the Stanford Synchrotron Radiation Lightsource. The analyses presented here indicate that the diffraction data are of high quality and do not suffer from excessive dehydration or radiation damage.