Exploring the structural dynamics of proteins by pressure perturbation using macromolecular crystallography.

High pressure is a convenient thermodynamic parameter to probe the dynamics of proteins as it is intimately related to volume which is essential for protein function. To be biologically active, a protein fluctuates between different substates. Pressure perturbation can promote some hidden substates by modifying the population between them. High pressure macromolecular crystallography (HPMX) is a perfect tool to capture and to characterize such substates at a molecular level providing new insights on protein dynamics. The present chapter describes the use of the diamond anvil cell to perform HPMX experiments. It also provides tips on sample preparation and optimal data collection as well as on efficient analysis of the resulting high-pressure structures.