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      Current developments in Coot for macromolecular model building of Electron Cryo‐microscopy and Crystallographic Data

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          Abstract

          Coot is a tool widely used for model building, refinement, and validation of macromolecular structures. It has been extensively used for crystallography and, more recently, improvements have been introduced to aid in cryo‐EM model building and refinement, as cryo‐EM structures with resolution ranging 2.5–4 A are now routinely available. Model building into these maps can be time‐consuming and requires experience in both biochemistry and building into low‐resolution maps. To simplify and expedite the model building task, and minimize the needed expertise, new tools are being added in Coot. Some examples include morphing, Geman‐McClure restraints, full‐chain refinement, and Fourier‐model based residue‐type‐specific Ramachandran restraints. Here, we present the current state‐of‐the‐art in Coot usage.

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          Structure validation by Calpha geometry: phi,psi and Cbeta deviation.

          Geometrical validation around the Calpha is described, with a new Cbeta measure and updated Ramachandran plot. Deviation of the observed Cbeta atom from ideal position provides a single measure encapsulating the major structure-validation information contained in bond angle distortions. Cbeta deviation is sensitive to incompatibilities between sidechain and backbone caused by misfit conformations or inappropriate refinement restraints. A new phi,psi plot using density-dependent smoothing for 81,234 non-Gly, non-Pro, and non-prePro residues with B < 30 from 500 high-resolution proteins shows sharp boundaries at critical edges and clear delineation between large empty areas and regions that are allowed but disfavored. One such region is the gamma-turn conformation near +75 degrees,-60 degrees, counted as forbidden by common structure-validation programs; however, it occurs in well-ordered parts of good structures, it is overrepresented near functional sites, and strain is partly compensated by the gamma-turn H-bond. Favored and allowed phi,psi regions are also defined for Pro, pre-Pro, and Gly (important because Gly phi,psi angles are more permissive but less accurately determined). Details of these accurate empirical distributions are poorly predicted by previous theoretical calculations, including a region left of alpha-helix, which rates as favorable in energy yet rarely occurs. A proposed factor explaining this discrepancy is that crowding of the two-peptide NHs permits donating only a single H-bond. New calculations by Hu et al. [Proteins 2002 (this issue)] for Ala and Gly dipeptides, using mixed quantum mechanics and molecular mechanics, fit our nonrepetitive data in excellent detail. To run our geometrical evaluations on a user-uploaded file, see MOLPROBITY (http://kinemage.biochem.duke.edu) or RAMPAGE (http://www-cryst.bioc.cam.ac.uk/rampage). Copyright 2003 Wiley-Liss, Inc.
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            JSME: a free molecule editor in JavaScript

            Background A molecule editor, i.e. a program facilitating graphical input and interactive editing of molecules, is an indispensable part of every cheminformatics or molecular processing system. Today, when a web browser has become the universal scientific user interface, a tool to edit molecules directly within the web browser is essential. One of the most popular tools for molecular structure input on the web is the JME applet. Since its release nearly 15 years ago, however the web environment has changed and Java applets are facing increasing implementation hurdles due to their maintenance and support requirements, as well as security issues. This prompted us to update the JME editor and port it to a modern Internet programming language - JavaScript. Summary The actual molecule editing Java code of the JME editor was translated into JavaScript with help of the Google Web Toolkit compiler and a custom library that emulates a subset of the GUI features of the Java runtime environment. In this process, the editor was enhanced by additional functionalities including a substituent menu, copy/paste, drag and drop and undo/redo capabilities and an integrated help. In addition to desktop computers, the editor supports molecule editing on touch devices, including iPhone, iPad and Android phones and tablets. In analogy to JME the new editor is named JSME. This new molecule editor is compact, easy to use and easy to incorporate into web pages. Conclusions A free molecule editor written in JavaScript was developed and is released under the terms of permissive BSD license. The editor is compatible with JME, has practically the same user interface as well as the web application programming interface. The JSME editor is available for download from the project web page http://peter-ertl.com/jsme/
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              Model-based local density sharpening of cryo-EM maps

              Atomic models based on high-resolution density maps are the ultimate result of the cryo-EM structure determination process. Here, we introduce a general procedure for local sharpening of cryo-EM density maps based on prior knowledge of an atomic reference structure. The procedure optimizes contrast of cryo-EM densities by amplitude scaling against the radially averaged local falloff estimated from a windowed reference model. By testing the procedure using six cryo-EM structures of TRPV1, β-galactosidase, γ-secretase, ribosome-EF-Tu complex, 20S proteasome and RNA polymerase III, we illustrate how local sharpening can increase interpretability of density maps in particular in cases of resolution variation and facilitates model building and atomic model refinement.
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                Author and article information

                Contributors
                pemsley@mrc-lmb.cam.ac.uk
                Journal
                Protein Sci
                Protein Sci
                10.1002/(ISSN)1469-896X
                PRO
                Protein Science : A Publication of the Protein Society
                John Wiley & Sons, Inc. (Hoboken, USA )
                0961-8368
                1469-896X
                02 March 2020
                April 2020
                02 March 2020
                : 29
                : 4 ( doiID: 10.1002/pro.v29.4 )
                : 1069-1078
                Affiliations
                [ 1 ] MRC Laboratory of Molecular Biology Cambridge Biomedical Campus Cambridge UK
                [ 2 ] Division of Molecular Structural Biology, Department of Medical Biochemistry and Biophysics Karolinska Institutet Stockholm Sweden
                Author notes
                [*] [* ] Correspondence

                Paul Emsley, MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge CB2 0QH, UK.

                Email: pemsley@ 123456mrc-lmb.cam.ac.uk

                Author information
                https://orcid.org/0000-0002-0334-0591
                https://orcid.org/0000-0002-5646-2090
                https://orcid.org/0000-0002-0138-5227
                Article
                PRO3791
                10.1002/pro.3791
                7096722
                31730249
                72692cd7-c153-48c3-bf86-4c872c01608d
                © 2019 The Authors. Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

                This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

                History
                : 03 October 2019
                : 13 November 2019
                : 14 November 2019
                Page count
                Figures: 3, Tables: 0, Pages: 10, Words: 7019
                Funding
                Funded by: European Molecular Biology Organization , open-funder-registry 10.13039/100004410;
                Award ID: ALTF66‐2015
                Funded by: FP7 People: Marie‐Curie Actions , open-funder-registry 10.13039/100011264;
                Award ID: ERC 725685
                Award ID: GA‐2013‐609409
                Funded by: Medical Research Council , open-funder-registry 10.13039/501100007155;
                Award ID: MC_U105184330
                Award ID: MC_U105192715
                Award ID: MC_UP_A025_1012
                Funded by: Röntgen‐Ångström Cluster , open-funder-registry 10.13039/501100011750;
                Award ID: 349‐2013‐597
                Funded by: Röntgen Ångström Cluster , open-funder-registry 10.13039/501100011750;
                Funded by: UK Research and Innovation , open-funder-registry 10.13039/100014013;
                Funded by: European Union's Horizon 2020 research and innovation programme
                Funded by: European Commission , open-funder-registry 10.13039/501100000780;
                Categories
                Tools for Protein Science
                Tools for Protein Science
                Custom metadata
                2.0
                April 2020
                Converter:WILEY_ML3GV2_TO_JATSPMC version:5.7.8 mode:remove_FC converted:26.03.2020

                Biochemistry
                ligands,macromolecular model building,molecular biophysics,real space refinement,rotamers,validation

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