research papers 12 doi:10.1107/S0907444909042073 Acta Cryst. (2010). D66, 12–21 Acta Crystallographica Section D Biological Crystallography ISSN 0907-4449 MolProbity: all-atom structure validation for macromolecular crystallography Vincent B. Chen, W. Bryan Arendall III, Jeffrey J. Headd, Daniel A. Keedy, Robert M. Immormino, Gary J. Kapral, Laura W. Murray, Jane S. Richardson and David C. Richardson* Department of Biochemistry, Duke University, Durham, NC 27710, USA Correspondence e-mail: dcr@kinemage.biochem.duke.edu MolProbity is a structure-validation web service that provides broad-spectrum solidly based evaluation of model quality at both the global and local levels for both proteins and nucleic acids. It relies heavily on the power and sensitivity provided by optimized hydrogen placement and all-atom contact analysis, complemented by updated versions of covalent-geometry and torsion-angle criteria. Some of the local corrections can be performed automatically in MolProbity and all of the diagnostics are presented in chart and graphical forms that help guide manual rebuilding. X-ray crystallography provides a wealth of biologically important molecular data in the form of atomic three-dimensional structures of proteins, nucleic acids and increasingly large complexes in multiple forms and states. Advances in automation, in everything from crystal- lization to data collection to phasing to model building to refinement, have made solving a structure using crystallo- graphy easier than ever. However, despite these improve- ments, local errors that can affect biological interpretation are widespread at low resolution and even high-resolution structures nearly all contain at least a few local errors such as Ramachandran outliers, flipped branched protein side chains and incorrect sugar puckers. It is critical both for the crystallographer and for the end user that there are easy and reliable methods to diagnose and correct these sorts of errors in structures. MolProbity is the authors’ contribution to helping solve this problem and this article reviews its general capabilities, reports on recent enhancements and usage, and presents evidence that the resulting improvements are now beneficially affecting the global database. Received 21 August 2009 Accepted 13 October 2009 A version of this paper will be published as a chapter in the new edition of Volume F of International Tables for Crystallography. 1. Summary of MolProbity flow and user interactions The usual interaction with MolProbity (Davis et al. , 2007) is through the internet at http://molprobity.biochem.duke.edu or as a main menu item on our general laboratory website at http://kinemage.biochem.duke.edu. [For bulk users, it is also possible to set up your own local MolProbity server or to use the individual programs in command-line mode.] Tutorial exercises for the whole process of diagnosing and fixing errors can be found on the kinemage site under Teaching/ MolProbity. A typical MolProbity session starts with the user uploading a coordinate file of their own or fetching one from the PDB or NDB databases (Berman et al., 1992, 2000) in new or old PDB format or in mmCIF format. After checking the thumbnail image and listed characteristics of the input file and editing or reloading if needed, H atoms are added and optimized, with