Paul Emsley

Features and development of Coot

Coot is a molecular-graphics program designed to assist in the building of protein and other macromolecular models. The current state of development and available features are presented.

Overview of the CCP4 suite and current developments

An overview of the CCP4 software suite for macromolecular crystallography is given.

Developments in the CCP4 molecular-graphics project

Progress towards structure determination that is both high-throughput and high-value is dependent on the development of integrated and automatic tools for electron-density map interpretation and for the analysis of the resulting atomic models. Advances in map-interpretation algorithms are extending the resolution regime in which fully automatic tools can work reliably, but at present human intervention is required to interpret poor regions of macromolecular electron density, particularly where crystallographic data is only available to modest resolution [for example, I/sigma(I) < 2.0 for minimum resolution 2.5 A]. In such cases, a set of manual and semi-manual model-building molecular-graphics tools is needed. At the same time, converting the knowledge encapsulated in a molecular structure into understanding is dependent upon visualization tools, which must be able to communicate that understanding to others by means of both static and dynamic representations. CCP4 mg is a program designed to meet these needs in a way that is closely integrated with the ongoing development of CCP4 as a program suite suitable for both low- and high-intervention computational structural biology. As well as providing a carefully designed user interface to advanced algorithms of model building and analysis, CCP4 mg is intended to present a graphical toolkit to developers of novel algorithms in these fields.

A New Generation of Crystallographic Validation Tools for the Protein Data Bank

Summary This report presents the conclusions of the X-ray Validation Task Force of the worldwide Protein Data Bank (PDB). The PDB has expanded massively since current criteria for validation of deposited structures were adopted, allowing a much more sophisticated understanding of all the components of macromolecular crystals. The size of the PDB creates new opportunities to validate structures by comparison with the existing database, and the now-mandatory deposition of structure factors creates new opportunities to validate the underlying diffraction data. These developments highlighted the need for a new assessment of validation criteria. The Task Force recommends that a small set of validation data be presented in an easily understood format, relative to both the full PDB and the applicable resolution class, with greater detail available to interested users. Most importantly, we recommend that referees and editors judging the quality of structural experiments have access to a concise summary of well-established quality indicators.

Tools for macromolecular model building and refinement into electron cryo-microscopy reconstructions.

A description is given of new tools to facilitate model building and refinement into electron cryo-microscopy reconstructions.

Handling ligands with Coot

Coot is a molecular-graphics program designed to assist in the building of protein and other macromolecular models. The current state of ligand tools is presented.

The new CCP4 Coordinate Library as a toolkit for the design of coordinate-related applications in protein crystallography

The new CCP4 Coordinate Library is a development aiming to provide a common layer of coordinate-related functionality to the existing applications in the CCP4 suite, as well as a variety of tools that can simplify the design of new applications where they relate to atomic coordinates. The Library comprises a wide spectrum of useful functions, ranging from parsing coordinate formats and elementary editing operations on the coordinate hierarchy of biomolecules, to high-level functionality such as calculation of secondary structure, interatomic bonds, atomic contacts, symmetry transformations, structure superposition and many others. Most of the functions are available in a C++ object interface; however, a Fortran interface is provided for compatibility with older CCP4 applications. The paper describes the general principles of the Library design and the most important functionality. The Library, together with documentation, is available under the LGPL license from the CCP4 suite version 5.0 and higher.

Stereochemistry of carbon monoxide binding to normal human adult and Cowtown haemoglobins.

The structures of carbonmonoxyhaemoglobins A and Cowtown (His146 beta----Leu) have been refined at 2.2 A (1 A = 0.1 nm) and 2.3 A resolution, respectively. The least squares fit to the Fe-C-O line makes an angle to the haem normal of about 6 degrees. The Fe-C-O group is bent from linearity by about 7 degrees. The porphyrins in the CO liganded haemoglobins are ruffled. This deformation of the haem and the distortion of the Fe-C-O group may explain the low CO affinity of haemoglobin. The electron density for the C-terminal residues is low but sufficient to distinguish the histidyl and leucyl residues clearly. The similarity between these two structures, apart from 146 beta, means that the reduced alkaline Bohr effect is due solely to the replacement of histidine by a leucine.

From crystal to structure with CCP4

An introduction to the proceedings of the CCP4 study weekend is given.

Autofix for backward-fit sidechains: using MolProbity and real-space refinement to put misfits in their place

Misfit sidechains in protein crystal structures are a stumbling block in using those structures to direct further scientific inference. Problems due to surface disorder and poor electron density are very difficult to address, but a large class of systematic errors are quite common even in well-ordered regions, resulting in sidechains fit backwards into local density in predictable ways. The MolProbity web site is effective at diagnosing such errors, and can perform reliable automated correction of a few special cases such as 180° flips of Asn or Gln sidechain amides, using all-atom contacts and H-bond networks. However, most at-risk residues involve tetrahedral geometry, and their valid correction requires rigorous evaluation of sidechain movement and sometimes backbone shift. The current work extends the benefits of robust automated correction to more sidechain types. The Autofix method identifies candidate systematic, flipped-over errors in Leu, Thr, Val, and Arg using MolProbity quality statistics, proposes a corrected position using real-space refinement with rotamer selection in Coot, and accepts or rejects the correction based on improvement in MolProbity criteria and on χ angle change. Criteria are chosen conservatively, after examining many individual results, to ensure valid correction. To test this method, Autofix was run and analyzed for 945 representative PDB files and on the 50S ribosomal subunit of file 1YHQ. Over 40% of Leu, Val, and Thr outliers and 15% of Arg outliers were successfully corrected, resulting in a total of 3,679 corrected sidechains, or 4 per structure on average. Summary Sentences: A common class of misfit sidechains in protein crystal structures is due to systematic errors that place the sidechain backwards into the local electron density. A fully automated method called “Autofix” identifies such errors for Leu, Val, Thr, and Arg and corrects over one third of them, using MolProbity validation criteria and Coot real-space refinement of rotamers.