NMRe: a web server for NMR protein structure refinement with high-quality structure validation scores

State-of-the-art methods based on CNS and CYANA were used to recalculate the nuclear magnetic resonance (NMR) solution structures of 500+ proteins for which coordinates and NMR restraints are available from the Protein Data Bank. Curated restraints were obtained from the BioMagResBank FRED database. Although the original NMR structures were determined by various methods, they all were recalculated by CNS and CYANA and refined subsequently by restrained molecular dynamics (CNS) in a hydrated environment. We present an extensive analysis of the results, in terms of various quality indicators generated by PROCHECK and WHAT_CHECK. On average, the quality indicators for packing and Ramachandran appearance moved one standard deviation closer to the mean of the reference database. The structural quality of the recalculated structures is discussed in relation to various parameters, including number of restraints per residue, NOE completeness and positional root mean square deviation (RMSD). Correlations between pairs of these quality indicators were generally low; for example, there is a weak correlation between the number of restraints per residue and the Ramachandran appearance according to WHAT_CHECK (r = 0.31). The set of recalculated coordinates constitutes a unified database of protein structures in which potential user- and software-dependent biases have been kept as small as possible. The database can be used by the structural biology community for further development of calculation protocols, validation tools, structure-based statistical approaches and modeling. The RECOORD database of recalculated structures is publicly available from http://www.ebi.ac.uk/msd/recoord.

We have found that refinement of protein NMR structures using Rosetta with experimental NMR restraints yields more accurate protein NMR structures than those that have been deposited in the PDB using standard refinement protocols. Using 40 pairs of NMR and X-ray crystal structures determined by the Northeast Structural Genomics Consortium, for proteins ranging in size from 5-22 kDa, restrained Rosetta refined structures fit better to the raw experimental data, are in better agreement with their X-ray counterparts, and have better phasing power compared to conventionally determined NMR structures. For 37 proteins for which NMR ensembles were available and which had similar structures in solution and in the crystal, all of the restrained Rosetta refined NMR structures were sufficiently accurate to be used for solving the corresponding X-ray crystal structures by molecular replacement. The protocol for restrained refinement of protein NMR structures was also compared with restrained CS-Rosetta calculations. For proteins smaller than 10 kDa, restrained CS-Rosetta, starting from extended conformations, provides slightly more accurate structures, while for proteins in the size range of 10-25 kDa the less CPU intensive restrained Rosetta refinement protocols provided equally or more accurate structures. The restrained Rosetta protocols described here can improve the accuracy of protein NMR structures and should find broad and general for studies of protein structure and function.