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J M Thornton

Publications and source records attributed to J M Thornton.

At least 73 records · Page 4Linked to original sources
PubMed1998

Structural studies of Impatiens balsamina antimicrobial protein (Ib-AMP1).

Structural studies of Ib-AMP1, a small antimicrobial peptide derived from the seeds of Impatiens balsamina have been performed using circular dichroism (CD) and two-dimensional proton nuclear magnetic resonance (1H NMR). This 20-residue peptide is highly basic with five arginine residues and contains four cysteines which form two intramolecular disulfide bonds. CD results reveal that the peptide may include a beta-turn but do not show evidence for either helical or beta-sheet structure over a range of temperature and pH. Structural information from NMR was obtained in the form of proton-proton internuclear distances inferred from NOEs and dihedral angle restraints from spin-spin coupling constants, which were used for distance geometry calculations. Owing to the difficulty in obtaining the correct disulfide connectivities by chemical methods, three separate calculations were performed; with no disulfides and with the two possible alternate disulfide connectivities. Results from distance geometry calculations reveal that although the peptide is small, the cysteines constrain part of it to adopt a well-defined main chain conformation. From residue 6 to 20, the backbone is well defined, whilst the N-terminal region, residues 1-5, has very few constraints and appears to be very flexible. In the defined core region, there are three beta-turns at residues 9-12, 10-13, and 12-15. The side chains show no strong interactions in the NMR spectra and are therefore thought to adopt multiple conformations. Superposition of the structures generated shows that the peptide has two hydrophilic patches which are at opposite ends of the molecule separated by a large hydrophobic patch. Little is known about the mode of action of this protein, but it is thought to interact with a membrane-bound receptor, and possible sites of interaction are discussed. The structures determined are compared with those of the alpha-conotoxins, which are also highly basic proteins with similar disulfide connectivities.

Amino Acid Sequence↗
PubMed1998

Contemporary approaches to protein structure classification.

In a similar manner to sequence database searching, it is also possible to compare three-dimensional protein structure. Such methods can be extremely useful because a structural similarity may represent a distant evolutionary relationship that is undetectable by sequence analysis. In this review, we summarise the most popular structure comparison methods, show how they can be used for database searching, and then describe some of the most advanced attempts to develop comprehensive protein structure classifications. With such data, it is possible to identify distant evolutionary relationships, provide libraries of unique folds for structure prediction, estimate the total number of folds that exist, and investigate the preference for certain types of structures over others.

Databases, Factual↗
PubMed1998

Domain assignment for protein structures using a consensus approach: characterization and analysis.

A consensus approach for the assignment of structural domains in proteins is presented. The approach combines a number of previously published algorithms, and takes advantage of the elevated accuracy obtained when assignments from the individual algorithms are in agreement. The consensus approach is tested on a data set of 55 protein chains, for which domain assignments from four automated methods were known, and for which crystallographers assignments had been reported in the literature. Accuracy was found to increase in this test from 72% using individual algorithms to 100% when all four methods were in agreement. However a consensus prediction using all four methods was only possible for 52% of the dataset. The consensus approach [using three publicly available domain assignment algorithms (PUU, DETECTIVE, DOMAK)] was then used to make domain assignments for a data set of 787 protein chains from the Protein Data Bank. Analysis of the assignments showed 55.7% of assignments could be made automatically, and of these, 13.5% were multi-domain proteins. Of the remaining 44.3% that could not be assigned by the consensus procedure 90.4% had their domain boundaries assigned correctly by at least one of the algorithms. Once identified, these domains were analyzed for trends in their size and secondary structure class. In addition, the discontinuity of each domain along the protein chain was considered.

Algorithms↗
PubMed1998

Determinants of strand register in antiparallel beta-sheets of proteins.

Antiparallel beta-sheets present two distinct environments to inter-strand residue pairs: beta(A,HB) sites have two backbone hydrogen bonds; whereas at beta(A,NHB) positions backbone hydrogen bonding is precluded. We used statistical methods to compare the frequencies of amino acid pairs at each site. Only approximately 10% of the 210 possible pairs showed occupancies that differed significantly between the two sites. Trends were clear in the preferred pairs, and these could be explained using stereochemical arguments. Cys-Cys, Aromatic-Pro, Thr-Thr, and Val-Val pairs all preferred the beta(A,NHB) site. In each case, the residues usually adopted sterically favored chi1 conformations, which facilitated intra-pair interactions: Cys-Cys pairs formed disulfide bonds; Thr-Thr pairs made hydrogen bonds; Aromatic-Pro and Val-Val pairs formed close van der Waals contacts. In contrast, to make intimate interactions at a beta(A,HB) site, one or both residues had to adopt less favored chi1 geometries. Nonetheless, pairs containing glycine and/or aromatic residues were favored at this site. Where glycine and aromatic side chains combined, the aromatic residue usually adopted the gauche conformation, which promoted novel aromatic ring-peptide interactions. This work provides rules that link protein sequence and tertiary structure, which will be useful in protein modeling, redesign, and de novo design. Our findings are discussed in light of previous analyses and experimental studies.

Binding Sites↗
PubMed1998

Characterization of atypical Aeromonas salmonicida by different methods.

Fifty two isolates of atypical Aeromonas salmonicida, recovered from a wide range of hosts and geographical locations, were heterogeneous in terms of molecular and phenotypic characteristics, and represented taxa which could not be accommodated by the current classification of four subspecies. Generally, there was incongruence between the molecular (PCR, RAPD and ribotyping) and phenotypic methods in terms of cluster membership. By PCR, 6 groups were described of which Group 1 encompassed 12 isolates including the type strain of A. salmonicida subsp. smithia. Group 2 accommodated 23 isolates including the reference cultures of subspecies achromogenes and masoucida. The named culture of Haemophilus piscium was recovered in Group 6. By ribotyping and RAPD, the reference cultures were recovered in separate groups. All methods pointed to the uniqueness of subspecies smithia. Most isolates contained 2-6 plasmids, of 2.3 to 150 kb in length. Nevertheless, all isolates possessed certain key characteristics, including Gram-negativity, and the absence of motility.

Aeromonas↗
PubMed1998

Protein structure prediction.

Genome sequencing projects continue to provide a flood of new protein sequences, and prediction methods remain an important means of adding structural information. Recently, there have been advances in secondary structure prediction, which feed, in turn, into improved fold recognition algorithms. Finally, there have been technical improvements in comparative modelling, and studies of the expected accuracy of three-dimensional structural models built by this method.

Journal Article↗
PubMed1998

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.

Crystallography, X-Ray↗
PubMed1998

Dihydrofolate reductase: a potential drug target in trypanosomes and leishmania.

Dihydrofolate reductase has successfully been used as a drug target in the area of anti-cancer, anti-bacterial and anti-malarial chemotherapy. Little has been done to evaluate it as a drug target for treatment of the trypanosomiases and leishmaniasis. A crystal structure of Leishmania major dihydrofolate reductase has been published. In this paper, we describe the modelling of Trypanosoma cruzi and Trypanosoma brucei dihydrofolate reductases based on this crystal structure. These structures and models have been used in the comparison of protozoan, bacterial and human enzymes in order to highlight the different features that can be used in the design of selective anti-protozoan agents. Comparison has been made between residues present in the active site, the accessibility of these residues, charge distribution in the active site, and the shape and size of the active sites. Whilst there is a high degree of similarity between protozoan, human and bacterial dihydrofolate reductase active sites, there are differences that provide potential for selective drug design. In particular, we have identified a set of residues which may be important for selective drug design and identified a larger binding pocket in the protozoan than the human and bacterial enzymes.

Amino Acid Sequence↗
PubMed1998

Sequences annotated by structure: a tool to facilitate the use of structural information in sequence analysis.

With the aim of bridging the gap between protein sequence and structural analyses, we have developed a tool to aid the identification of new protein sequences by recognizing distant homologues using structural information. The tool generates sequence annotated by structure (SAS) files, applying structural information derived from structural analyses to a given protein sequence. A World Wide Web interface allows a given sequence to be submitted either for structural annotation or, where its structure is unknown, for search and alignment against sequences of known structure. In both cases, SAS will colour residues in the sequence of known structure according to a selection of properties, including secondary structure, interatomic contacts and active site information. SAS can also be used to inspect properties of a single structure.

Amino Acid Sequence↗
PubMed1998

Assessment of conformational parameters as predictors of limited proteolytic sites in native protein structures.

Despite the importance of limited proteolysis in biological systems it is often difficult to rationalize why a proteinase hydrolyses a particular bond, given a simple sequence specificity alone. Understanding of the structural properties limiting the proteolysis represents a first step on the pathway to control and manipulation of this phenomena. An expanded set of nick-sites in proteins of known tertiary structure, cut by both narrow and broad specificity proteinases, has been generated yielding a robust data set of strictly limited sites. A critical evaluation of an expanded set of conformational parameters revealed a strong correlation with limited proteolytic sites, although they are only modest predictors in isolation. The overall predictive power is significantly improved when the conformational parameters are combined in a weighted predictive scheme that permits their relative importance to be compared via a Metropolis search protocol. A subset of the parameters performs equally well demonstrating the key determinants of susceptibility. The derived predictive algorithm has been made available via the internet. Its utility for predicting other surface-correlated features is also discussed.

Algorithms↗
PubMed1997

NUCPLOT: a program to generate schematic diagrams of protein-nucleic acid interactions.

Proteins that bind to DNA are found in all areas of genetic activity within the cell. To help understand how these proteins perform their various functions, it is useful to analyse which residues are involved in binding to the DNA and how they interact with the bases and sugar-phosphate backbone of nucleic acids. Here we describe a program called NUCPLOT which can automatically identify these interactions from the 3D atomic coordinates of the complex from a PDB file and generate a plot that shows all the interactions in a schematic manner. The program produces a PostScript output file representing hydrogen, van der Waals and covalent bonds between the protein and the DNA. The resulting diagram is both clear and simple and allows immediate identification of important interactions within the structure. It also facilitates comparison of binding found in different structures. NUCPLOT is a completely automatic program, which can be used for any protein-DNA complex and will also work for certain protein-RNA structures.

Computer Graphics↗
PubMed1997

Prediction of protein-protein interaction sites using patch analysis.

A method for defining and analysing a series of residue patches on the surface of protein structures is used to predict the location of protein-protein interaction sites. Each residue patch is analysed for six parameters; solvation potential, residue interface propensity, hydrophobicity, planarity, protrusion and accessible surface area. The method involves the calculation of a relative combined score that gives the probability of a surface patch forming protein-protein interactions. Predictions are made for the known structures of protomers from 28 homo-dimers, large protomers from 11 hetero-complexes, small protomers from 14 hetero-complexes, and antigens from six antibody-antigen complexes. The predictions are successful for 66% (39/59) of the structures and the remainder can usually be rationalized in terms of additional interaction sites.

Antigens↗
PubMed1997

Analysis of protein-protein interaction sites using surface patches.

Protein-protein interaction sites in complexes of known structure are characterised using a series of parameters to evaluate what differentiates them from other sites on the protein surface. Surface patches are defined in protomers from a data set of 28 homo-dimers, 20 different hetero-complexes (segregated into large and small protomers), and antigens from six antibody-antigen complexes. Six parameters (solvation potential, residue interface propensity, hydrophobicity, planarity, protrusion and accessible surface area) are calculated for the observed interface patch and all other surface patches defined on each protein. A ranking of the observed interface, relative to all other possible patches, is calculated. With this approach it becomes possible to analyse the distribution of the rankings of all the observed patches, relative to all other surface patches, for each data set. For each type of complex, none of the parameters were definitive, but the majority showed trends for the observed interface to be distinguished from other surface patches.

Animals↗
PubMed1997

CATH--a hierarchic classification of protein domain structures.

BACKGROUND: Protein evolution gives rise to families of structurally related proteins, within which sequence identities can be extremely low. As a result, structure-based classifications can be effective at identifying unanticipated relationships in known structures and in optimal cases function can also be assigned. The ever increasing number of known protein structures is too large to classify all proteins manually, therefore, automatic methods are needed for fast evaluation of protein structures. RESULTS: We present a semi-automatic procedure for deriving a novel hierarchical classification of protein domain structures (CATH). The four main levels of our classification are protein class (C), architecture (A), topology (T) and homologous superfamily (H). Class is the simplest level, and it essentially describes the secondary structure composition of each domain. In contrast, architecture summarises the shape revealed by the orientations of the secondary structure units, such as barrels and sandwiches. At the topology level, sequential connectivity is considered, such that members of the same architecture might have quite different topologies. When structures belonging to the same T-level have suitably high similarities combined with similar functions, the proteins are assumed to be evolutionarily related and put into the same homologous superfamily. CONCLUSIONS: Analysis of the structural families generated by CATH reveals the prominent features of protein structure space. We find that nearly a third of the homologous superfamilies (H-levels) belong to ten major T-levels, which we call superfolds, and furthermore that nearly two-thirds of these H-levels cluster into nine simple architectures. A database of well-characterised protein structure families, such as CATH, will facilitate the assignment of structure-function/evolution relationships to both known and newly determined protein structures.

Databases, Factual↗
PubMed1997

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 C alpha root-mean-square deviations (RMSDs) compared with the observed structure of < 1 A for targets with high parental similarity; even at 26% sequence identity, the best model structures had C alpha deviations of only 2.2 A. 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 C alpha RMSDs > 18 A. 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 A, and was as high as 10 A for one structurally variable region. For the side chains, the chi 1 RMSDs are strongly correlated with the C alpha RMSDs. For models with C alpha deviations less than 1 A, on average 78.5% of side chains are placed in the correct rotamer, although the chi 1 RMSDs, though clearly better than random, were disappointing at around 46 degrees. As the backbone deviations increased, the side chain placement became less accurate, with an average chi 1 RMSD of 75 degrees on a 1.5-2.5 A C alpha 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 A). 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.

Models, Molecular↗