Improving the Accuracy of Macromolecular Structure Refinement at 7 Å Resolution

Abstract

SUMMARY In X-ray crystallography, molecular replacement and subsequent refinement is challenging at low resolution. We compared refinement methods using synchrotron diffraction data of photosystem I at 7.4 Å resolution, starting from different initial models with increasing deviations from the known high-resolution structure. Standard refinement spoiled the initial models moving them further away from the true structure and leading to high Rfree-values. In contrast, DEN-refinement improved even the most distant starting model as judged by Rfree, atomic root-mean-square differences to the true structure, significance of features not included in the initial model, and connectivity of electron density. The best protocol was DEN-refinement with initial segmented rigid-body refinement. For the most distant initial model, the fraction of atoms within 2 Å of the true structure improved from 24% to 60%. We also found a significant correlation between Rfree-values and the accuracy of the model, suggesting that Rfree is useful even at low resolution.

Structural Basis for Inhibitor-Induced Aggregation of HIV Integrase

“…We determined the structure by molecular replacement using the catalytic core domain dimer structure (without ligand) from the GSK1264-bound structure previously determined (PDB 4OJR, [7]) together with the previously reported IN CTD structure [7,25]. The structure was refined using deformable elastic network (DEN) restraints [26–28]. The catalytic core domains, CTDs, and connecting linkers displayed strong electron density in simulated annealing 2F o -F c composite omit maps, and thus, their positions are well defined in the structure (Fig 2A).…”

Structural evidence for Nap1‐dependent H2A–H2B deposition and nucleosome assembly

“…B factors from the high‐resolution search models were maintained during the initial cycles of refinement and the initial model was used as both the starting and reference model for subsequent Deformable Elastic Network (DEN) refinement using CNS over a grid‐enabled web server hosted by SBGrid (Schroder et al , 2010; O'Donovan et al , 2012). The refinement protocol was similar to that used previously (Brunger et al , 2012) with the following non‐default setting: Only one overall anisotropic B factor refinement per chain was carried out with the starting B factors maintained from the original high‐resolution models. DEN restraints and non‐crystallographic symmetry (NCS) restraints were maintained throughout the refinement procedure.…”

An Autoinhibited Structure of α-Catenin and Its Implications for Vinculin Recruitment to Adherens Junctions

“…The structure solution was solved by molecular replacement with PHASER (32) using the ␣E-catenin N domain residues 82-262 (PDB code 1DOV) as a search model for the N domain and a homology model of ␣E-catenin M II-III regions generated from the full-length vinculin structure (PDB code 1ST6) as a search model for the M domain. For refinement at 6.5-Å resolution, the deformable elastic network (DEN) refinement was performed by using CNS 1.3 (33) as previously described (34), except for the sole implementation of overall anisotropic B-factor refinement and the constitutive use of the DEN restraints throughout the process (35). The optimum values of the ␥ and W DEN parameters of DEN refinement were determined by a two-dimensional grid search using the SBGrid DEN refinement portal.…”