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X-ray refinement significantly underestimates the level of microscopic heterogeneity in biomolecular crystals.

ABSTRACT: Biomolecular X-ray structures typically provide a static, time- and ensemble-averaged view of molecular ensembles in crystals. In the absence of rigid-body motions and lattice defects, B-factors are thought to accurately reflect the structural heterogeneity of such ensembles. In order to study the effects of averaging on B-factors, we employ molecular dynamics simulations to controllably manipulate microscopic heterogeneity of a crystal containing 216 copies of villin headpiece. Using average structure factors derived from simulation, we analyse how well this heterogeneity is captured by high-resolution molecular-replacement-based model refinement. We find that both isotropic and anisotropic refined B-factors often significantly deviate from their actual values known from simulation: even at high 1.0?Å resolution and Rfree of 5.9%, B-factors of some well-resolved atoms underestimate their actual values even sixfold. Our results suggest that conformational averaging and inadequate treatment of correlated motion considerably influence estimation of microscopic heterogeneity via B-factors, and invite caution in their interpretation.

SUBMITTER: Kuzmanic A

PROVIDER: S-EPMC3926004 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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X-ray refinement significantly underestimates the level of microscopic heterogeneity in biomolecular crystals.

Kuzmanic Antonija A Pannu Navraj S NS Zagrovic Bojan B

Nature communications 20140101

Biomolecular X-ray structures typically provide a static, time- and ensemble-averaged view of molecular ensembles in crystals. In the absence of rigid-body motions and lattice defects, B-factors are thought to accurately reflect the structural heterogeneity of such ensembles. In order to study the effects of averaging on B-factors, we employ molecular dynamics simulations to controllably manipulate microscopic heterogeneity of a crystal containing 216 copies of villin headpiece. Using average st ...[more]

PMID: 24504120

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Project description:This study aims to solve the site-occupancy issues of any crystalline substance, showing that the structure refinement results should not be limited to a simple description of the crystal geometry but should go beyond that, providing refined site-scattering values consistent with the chemical data. A detailed description of the structure of the amphibole-supergroup minerals is very challenging owing to their complex chemical composition that renders the process of cation partition extremely difficult, particularly because of the occurrence of multivalent elements. Since amphiboles naturally occur under a fibrous morphology and have largely been used to produce asbestos, there is a growing demand for detailed and accurate structural data in order to study the relationships between structure, composition and toxicity. The present study proposes a recommended refinement procedure for both X-ray single-crystal structure refinement (SREF) and Rietveld analysis for tremolite, selected as a test case. The corresponding structural results are compared to estimate the ‘degree of confidence’ of the Rietveld refinement with regard to SREF. In particular, it is shown that the interpretation of the electron density of the tremolite structure by SREF is model dependent. By assuming that the site-scattering values from SREF should be as close as possible to those from electron microprobe analysis, as a crucial constraint for the correct description of the final crystal-chemical model, it is found that it is best satisfied by using partially ionized scattering curves (SCs) for O and Si, and neutral SCs (neutral oxygen curves or NOCs) for other atoms. This combination leads to the best fit to the diffraction data. Moreover, it is found that Rietveld refinement using NOCs produces the best structural results, in excellent agreement with SREF. It is worth noting that, due to the complexity of the diffraction pattern and the fairly large number of freely refinable parameters, refinements with different combinations of SCs produce results almost indistinguishable from a statistical point of view, albeit showing significant differences from a structural point of view.

Dataset Information

X-ray refinement significantly underestimates the level of microscopic heterogeneity in biomolecular crystals.

Publications

X-ray refinement significantly underestimates the level of microscopic heterogeneity in biomolecular crystals.

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