Instrumentation and experimental procedures for robust collection of X-ray diffraction data from protein crystals across physiological temperatures.
J Appl Crystallogr
; 53(Pt 6): 1493-1501, 2020 Dec 01.
Article
in En
| MEDLINE
| ID: mdl-33312102
ABSTRACT
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 â¼5â
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 1.09-1.54â
Å resolution data over a temperature range of 293-363â
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.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Language:
En
Journal:
J Appl Crystallogr
Year:
2020
Document type:
Article
Affiliation country:
United States