Crystal cryocooling distorts conformational heterogeneity in a model Michaelis complex of DHFR

Structure. 2014 Jun 10;22(6):899-910. doi: 10.1016/j.str.2014.04.016. Epub 2014 May 29.


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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Cold Temperature
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli / enzymology
  • Escherichia coli Proteins / chemistry*
  • Models, Molecular
  • Molecular Dynamics Simulation
  • Protein Conformation
  • Protein Structure, Tertiary
  • Static Electricity
  • Tetrahydrofolate Dehydrogenase / chemistry*


  • Escherichia coli Proteins
  • Tetrahydrofolate Dehydrogenase