Privateer: software for the conformational validation of carbohydrate structures

833 from the start and may require rebuilding (Fig. 1a). At lower resolutions, higher-energy conformations may appear as a consequence of underparameterized refinement, in spite of starting from a correct input model. These can be corrected during both realand reciprocalspace refinement (Fig. 1b), provided that enough restraints are introduced to balance the parameter-to-observation ratio. It is such examples that emphasize the problems of modeling sugars where the density is poor. Both published structures have high-energy conformations that cannot be deduced from the electron density, although for different reasons. In the first case, holds the conformation as determined by the Cremer-Pople algorithm4, as well as stereochemical and geometric information, for use as a reference upon validation. Additionally, puckering amplitudes4 of pyranoses are compared to those registered during ab initio metadynamics simulations of the conformational free-energy landscape of cyclohexane5. Privateer is able to feed results into Coot6 through its Python or Scheme scripting interface, loading models and maps automatically and flagging issues visually. Most of the conformational outliers detected at higher resolutions (<1.6 Å) are typically wrongly modeled Privateer (http://www.ccp4.ac.uk/html/ privateer.html) is a new software package aimed at the detection and prevention of conformational, regiochemical and stereochemical anomalies in cyclic monosaccharide structures. Carbohydrates, including Oand N-glycans attached to protein and lipid structures, are increasingly being studied in cellular biology. Crystallographic refinement of sugars is, however, poorly performed, thus leading to thousands of incorrect structures having been deposited in the Protein Data Bank (PDB)1,2. Although nomenclature validation has become possible in the past decade, with the introduction of tools such as PDB carbohydrate residue check (pdb-care)3, inappropriate refinement protocols at resolutions lower than 1.6 Å can still force a correct sugar into a highly improbable ring conformation, let alone distort one with chemical errors1. High-energy conformations are very infrequent in nature—perhaps even out of the question in N-glycans—and must always be backed by clear electron density. Otherwise, such conformations should be treated as outliers. Privateer identifies incorrect regiochemistry and stereochemistry and unlikely conformations. A real-space correlation coefficient against omit mFo – DFc electron density is also calculated as a quality-of-fit indicator. Bias-minimized map coefficients are exported automatically and can be subsequently used to assess identification of the sugars. Using this information, Privateer produces a visual checklist for rapid correction of the errors in real space and ensures that conformational preferences are accounted for during subsequent rebuilding and refinement. The software holds a manually curated database of supported monosaccharides based on the PDB Chemical Component Dictionary, whose entries contain coordinates for an energyminimized conformer that the PDB calculates upon new ligand depositions, by using Corina (Molecular Networks) or Omega (OpenEye). For each of these supported sugars, the database Privateer: software for the conformational validation of carbohydrate structures