Ensemble refinement of protein crystal structures in PHENIX

Abstract

X-ray crystallography typically uses a single set of coordinates and B-factors to describe macromolecular conformations. Refinement of multiple copies of the entire structure has been previously used in specific cases as an alternative means of representing structural flexibility. Here, we systematically validate this method using simulated diffraction data, and find ensemble refinement produces better representations of the distributions of atomic positions in the simulated structures than single conformer refinements. Comparison of principal components calculated from the refined ensembles and simulations shows that concerted motions are captured locally, but correlations dissipate over long distances. Ensemble refinement is also used on 50 experimental structures of varying resolution, and leads to decreases in R-free, implying that improvements in the representation of flexibility observed for the simulated structures may apply to real structures. These gains are essentially independent of resolution or data-to-parameter ratio, suggesting even structures at moderate resolution can benefit from ensemble refinement.