@article{55e39fadc3ce4cb4a6679bb1bce02161,
title = "Crystal cryocooling distorts conformational heterogeneity in a model michaelis complex of DHFR",
abstract = "Summary Most macromolecular X-ray structures are determined from cryocooled crystals, but it is unclear whether cryocooling distorts functionally relevant flexibility. Here we compare independently acquired pairs of high-resolution data sets of a model Michaelis complex of dihydrofolate reductase (DHFR), collected by separate groups at both room and cryogenic temperatures. These data sets allow us to isolate the differences between experimental procedures and between temperatures. Our analyses of multiconformer models and time-averaged ensembles suggest that cryocooling suppresses and otherwise modifies side-chain and main-chain conformational heterogeneity, quenching dynamic contact networks. Despite some idiosyncratic differences, most changes from room temperature to cryogenic temperature are conserved and likely reflect temperature-dependent solvent remodeling. Both cryogenic data sets point to additional conformations not evident in the corresponding room temperature data sets, suggesting that cryocooling does not merely trap preexisting conformational heterogeneity. Our results demonstrate that crystal cryocooling consistently distorts the energy landscape of DHFR, a paragon for understanding functional protein dynamics.",
author = "Keedy, {Daniel A.} and {Van Den Bedem}, Henry and Sivak, {David A.} and Petsko, {Gregory A.} and Dagmar Ringe and Wilson, {Mark A.} and Fraser, {James S.}",
note = "Funding Information: We thank Rahel Woldeyes and other Fraser lab members for helpful comments; Peter Wright and Gira Bhabha for discussions about DHFR; Tom Burnley for ensemble analysis scripts; James Holton, George Miegs, and Jane Tanamachi for technical support at ALS; and Nat Echols for PHENIX support. J.S.F. is supported by NIH OD009180, GM110580, and NSF STC-1231306. H.v.d.B. is supported by the US National Institute of General Medical Sciences Protein Structure Initiative (U54GM094586) at the Joint Center for Structural Genomics and a SLAC National Accelerator Laboratory LDRD (Laboratory Directed Research and Development) grant SLAC-LDRD-0014-13-2. D.A.S. is supported by NIH GM081879. G.A.P. and D.R. are supported by NIH GM32415. M.A.W. is supported by NIH GM092999. Funding Information: Diffraction data were collected at the Advanced Light Source, which is supported by the Director, Office of Science, Office of Basic Energy Sciences, of the US Department of Energy under contract no. DE-AC02-05CH11231, and at the Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, which is supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences under contract no. DE-AC02-76SF00515. The SSRL Structural Molecular Biology Program is supported by the DOE Office of Biological and Environmental Research, and by the National Institutes of Health, National Institute of General Medical Sciences (including P41GM103393). ",
year = "2014",
month = jun,
day = "10",
doi = "10.1016/j.str.2014.04.016",
language = "English (US)",
volume = "22",
pages = "899--910",
journal = "Structure",
issn = "0969-2126",
publisher = "Cell Press",
number = "6",
}