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    Refinement of atomic models in high resolution EM reconstructions using Flex-EM and local assessment

    Joseph, Agnel Praveen and Malhotra, S. and Burnley, T. and Wood, C. and Clare, D.K. and Winn, M. and Topf, Maya (2016) Refinement of atomic models in high resolution EM reconstructions using Flex-EM and local assessment. Methods 100 , pp. 42-49. ISSN 1046-2023.

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    Abstract

    As the resolutions of Three Dimensional Electron Microscopic reconstructions of biological macromolecules are being improved, there is a need for better fitting and refinement methods at high resolutions and robust approaches for model assessment. Flex-EM/MODELLER has been used for flexible fitting of atomic models in intermediate-to-low resolution density maps of different biological systems. Here, we demonstrate the suitability of the method to successfully refine structures at higher resolutions (2.5–4.5 Å) using both simulated and experimental data, including a newly processed map of Apo-GroEL. A hierarchical refinement protocol was adopted where the rigid body definitions are relaxed and atom displacement steps are reduced progressively at successive stages of refinement. For the assessment of local fit, we used the SMOC (segment-based Manders’ overlap coefficient) score, while the model quality was checked using the Qmean score. Comparison of SMOC profiles at different stages of refinement helped in detecting regions that are poorly fitted. We also show how initial model errors can have significant impact on the goodness-of-fit. Finally, we discuss the implementation of Flex-EM in the CCP-EM software suite.

    Metadata

    Item Type: Article
    Keyword(s) / Subject(s): Flexible fitting, Density fitting, Protein structure modelling, 3D-EM, Structure refinement, Model validation
    School: Birkbeck Schools and Departments > School of Science > Biological Sciences
    Research Centre: Bioinformatics, Bloomsbury Centre for, Structural Molecular Biology, Institute of (ISMB)
    Depositing User: Administrator
    Date Deposited: 12 Apr 2016 11:20
    Last Modified: 08 Dec 2017 14:22
    URI: http://eprints.bbk.ac.uk/id/eprint/14918

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