Malcolm W. MacArthur

PROCHECK: a program to check the stereochemical quality of protein structures

The PROCHECK suite of programs provides a detailed check on the stereochemistry of a protein structure. Its outputs comprise a number of plots in PostScript format and a comprehensive residue-by-residue listing. These give an assessment of the overall quality of the structure as compared with well refined structures of the same resolution and also highlight regions that may need further investigation. The PROCHECK programs are useful for assessing the quality not only of protein structures in the process of being solved but also of existing structures and of those being modelled on known structures.

AQUA and PROCHECK-NMR: Programs for checking the quality of protein structures solved by NMR*

SummaryThe AQUA and PROCHECK-NMR programs provide a means of validating the geometry and restraint violations of an ensemble of protein structures solved by solution NMR. The outputs include a detailed breakdown of the restraint violations, a number of plots in PostScript format and summary statistics. These various analyses indicate both the degree of agreement of the model structures with the experimental data, and the quality of their geometrical properties. They are intended to be of use both to support ongoing NMR structure determination and in the validation of the final results.

Influence of proline residues on protein conformation

To study the influence of proline residues on three-dimensional structure, an analysis has been made of all proline residues and their local conformations extracted from the Brookhaven Protein Data bank. We have considered the conformation of the proline itself, the relative occurrence of cis and trans peptides preceding proline residues, the influence of proline on the conformation of the preceding residue and the conformations of various proline patterns (Pro-Pro, Pro-X-Pro, etc.). The results highlight the unique role of proline in determining local conformation.

Validation of protein models derived from experiment

The growing number of protein structures solved at atomic resolution holds the promise of further improvements in geometry-based validation parameters. Additionally, the estimated standard uncertainties of the atomic coordinates have been computed for a number of X-ray structures, providing a measure of the coordinate precision. In NMR spectroscopy, a measure analogous to the crystallographic R-factor has been developed.

Assessment of comparative modeling in CASP2

An assessment is presented for all submissions to the comparative modeling challenge in the 1996 Critical Assessment of Structure Prediction (CASP2). Of the original 12 target structures, 9 were solved prior to the meeting: 8 by X‐ray crystallography and 1 by NMR spectroscopy. These targets varied over a large range of difficulty, as assessed by the percentage sequence identity with the principal parent structure, which ranged from 20% up to 85%. The overall quality of the models reflected the similarity of the principal parent. As expected, when the sequence alignment was correct, the core was accurately modeled, with the largest deviations occurring in the loops. Models were built which gave Cα root‐mean‐square deviations (RMSDs) compared with the observed structure of <1 Å for targets with high parental similarity; even at 26% sequence identity, the best model structures had Cα deviations of only 2.2 Å. Overall, these deviations are comparable with those observed between the parent structure and the target, but locally there are several examples where the model approaches closer to the target than does the parent. There were three targets below 25% sequence identity, and the models generated for these targets were, in general, significantly less accurate. This principally reflects errors in the alignment which, if systematically shifted, can generate Cα RMSDs <18 Å. Compared with CASP1, the geometry of the models was significantly improved with no D‐amino acids. By far the major contribution to RMSD error was the alignment accuracy, which varied from 100% down to 7% over the range of targets. In the structurally variable regions, global shifts, caused by hinge bending, were the major source of error, giving significantly lower local RMSDs than global RMSDs. In over 50% of these noncore regions, the difference between global and local RMSDs was more than 3 Å, and was as high as 10 Å for one structurally variable region. For the side chains, the χ1 RMSDs are strongly correlated with the Cα RMSDs. For models with Cα deviations less than 1 Å, on average 78.5% of side chains are placed in the correct rotamer, although the χ1 RMSDs, though clearly better than random, were disappointing at around 46°. As the backbone deviations increased, the side chain placement became less accurate, with an average χ1 RMSD of 75° on a 1.5–2.5 Å Cα backbone (average 51.4% correct rotamer). Refinement by energy minimization or molecular dynamics made only minor adjustments to improve local geometry and generally made small, but not significant, improvements to the RMSD. In total, 19 groups submitted 62 models (89 coordinate sets) that could be assessed. Most modelers used manual adjustments to sequence alignments and, in general, good alignments were obtained down to 25% sequence identity. The modeling methods ranged from “classical” modeling, involving core building followed by loop and side chain addition, to more sophisticated approaches based on probability distributions, Monte Carlo sampling or distance geometry. For each target, several groups produced equally good models, given the expected errors in the structures (about 0.5 Å). No one method came out as clearly superior, although the approaches that inherit directly from the parents generally performed better than the more radical techniques. However, for each target there were some poor models, usually reflecting a poor sequence alignment, and the range of accuracy for each target is therefore large. Fully automated methods are able to perform very well for “easy” targets (85% sequence identity with parent), but when modeling using a distantly related parent, care and expertise, especially in performing the alignment, still appear to be important factors in generating accurate models. Proteins, Suppl. 1:14–28, 1997.

Molecular basis of inherited diseases: a structural perspective

Mutations in human genes can change the sequence and structure of a protein, impair its function, and could lead to disease. The increasing number of human protein structures provides an opportunity to explore further the molecular basis of many diseases. By studying inherited disease genes and analysing three-dimensional information, we can often explain why different phenotypes originate from mutations in the same gene. Although interpreting the effects of mutations in a protein structure can be difficult, it can provide more detailed information about the environment and role of a mutated residue than the protein sequence alone. Nevertheless, protein sequence information and evolutionary sequence conservation still remain powerful indicators of which mutations will impair the function of a gene product.

Knowledge-based validation of protein structure coordinates derived by X-ray crystallography and NMR spectroscopy

Abstract Over the past few years, several protein structures solved by X-ray crystallography have been found to contain serious errors. This has prompted the development of methods for validating protein structures, in both the X-ray and NMR fields. A number of computational tools have recently been developed for assessing structures independently of the experimental data, thereby providing an additional, complementary check on their correctness. In effect, the tests assess the normality of a protein and are both local, checking the correctness of the local geometry, and global, verifying the overall fold. In structure determination by NMR the problems of validation are compounded by multiple solutions for the experimental data and lack of agreement over means of assessing the fit of the models to the data.

Protein side-chain conformation: a systematic variation of χ1 mean values with resolution – a consequence of multiple rotameric states?

A systematic variation with resolution of the mean values of the gauche-, trans and gauche+ chi1 rotamers in protein structures determined by X-ray crystallography has been observed. Further analysis revealed that these correlations differ considerably between residue types, being highly significant for some residue types (e.g. Ser, Thr, Leu, Lys) and absent for others (e.g. aromatics). For the individual residue types which exhibited the trend most strongly, these changes were accompanied by corresponding systematic variations in the percentage relative populations in the three energy wells. Examination of a uniformly sized subset of monomers showed that this effect, while attenuated, was still present, and was thus not entirely a consequence of the change in size and surface area which also correlates with resolution. An analysis of B values in the disfavoured high-energy barrier region between the rotameric wells showed a pronounced tendency towards larger than average values. As a plausible hypothesis, it is suggested here that these observations can be accounted for by the presence of multiple rotameric states. The averaged electron density produced by dual occupancy at low resolution giving an averaged conformation is resolved at high resolution into its individual components.

NMR and crystallography : complementary approaches to structure determination

A knowledge of the three-dimensional structure of a protein is essential to understand how a protein performs its functions. It is also a prime requirement for the rational design of novel sequences with specific structural, chemical or catalytic properties. Until recently, such information could only be obtained from X-ray diffraction studies on protein crystals. In the past few years, however, nuclear magnetic resonance (NMR) spectroscopy in solution has rapidly become established as an effective alternative method. This review briefly examines the two techniques and their relevance to protein engineering and design.

Evolution of binding sites for zinc and calcium ions playing structural roles.

The geometry of metal coordination by proteins is well understood, but the evolution of metal binding sites has been less studied. Here we present a study on a small number of well‐documented structural calcium and zinc binding sites, concerning how the geometry diverges between relatives, how often nonrelatives converge towards the same structure, and how often these metal binding sites are lost in the course of evolution. Both calcium and zinc binding site structure is observed to be conserved; structural differences between those atoms directly involved in metal binding in related proteins are typically less than 0.5 Å root mean square deviation, even in distant relatives. Structural templates representing these conserved calcium and zinc binding sites were used to search the Protein Data Bank for cases where unrelated proteins have converged upon the same residue selection and geometry for metal binding. This allowed us to identify six “archetypal” metal binding site structures: two archetypal zinc binding sites, both of which had independently evolved on a large number of occasions, and four diverse archetypal calcium binding sites, where each had evolved independently on only a handful of occasions. We found that it was common for distant relatives of metal‐binding proteins to lack metal‐binding capacity. This occurred for 13 of the 18 metal binding sites we studied, even though in some of these cases the original metal had been classified as “essential for protein folding.” For most of the calcium binding sites studied (seven out of eleven cases), the lack of metal binding in relatives was due to point mutation of the metal‐binding residues, whilst for zinc binding sites, lack of metal binding in relatives always involved more extensive changes, with loss of secondary structural elements or loops around the binding site. Proteins 2008.