Effects of cryo-EM cooling on structural ensembles

被引:36
作者
Bock, Lars, V [1 ]
Grubmueller, Helmut [1 ]
机构
[1] Max Planck Inst Multidisciplinary Sci, Theoret & Computat Biophys Dept, Gottingen, Germany
关键词
GLASS-TRANSITION; CRYOELECTRON MICROSCOPY; PROTEIN-STRUCTURE; DATA-ACQUISITION; RIBONUCLEASE-A; DYNAMICS; TEMPERATURE; BEHAVIOR; WATER; SIMULATION;
D O I
10.1038/s41467-022-29332-2
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Structure determination by cryo electron microscopy (cryo-EM) provides information on structural heterogeneity and ensembles at atomic resolution. To obtain cryo-EM images of macromolecules, the samples are first rapidly cooled down to cryogenic temperatures. To what extent the structural ensemble is perturbed during cooling is currently unknown. Here, to quantify the effects of cooling, we combined continuum model calculations of the temperature drop, molecular dynamics simulations of a ribosome complex before and during cooling with kinetic models. Our results suggest that three effects markedly contribute to the narrowing of the structural ensembles: thermal contraction, reduced thermal motion within local potential wells, and the equilibration into lower free-energy conformations by overcoming separating free-energy barriers. During cooling, barrier heights below 10 kJ/mol were found to be overcome, which is expected to reduce B-factors in ensembles imaged by cryo-EM. Our approach now enables the quantification of the heterogeneity of room-temperature ensembles from cryo-EM structures. The rapid temperature drop during plunge-freezing affects the structural ensembles obtained by cryo-EM. To quantify the extent of perturbation, Bock and Grubmuller combined continuum calculations, MD simulations, and kinetic models.
引用
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页数:13
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