Biomedical subjects

David S Moss

Publications and source records attributed to David S Moss.

9 recordsLinked to original sources

PubMed2006

Benchmarking pK(a) prediction.

BACKGROUND: pKa values are a measure of the protonation of ionizable groups in proteins. Ionizable groups are involved in intra-protein, protein-solvent and protein-ligand interactions as well as solubility, protein folding and catalytic activity. The pKa shift of a group from its intrinsic value is determined by the perturbation of the residue by the environment and can be calculated from three-dimensional structural data. RESULTS: Here we use a large dataset of experimentally-determined pKas to analyse the performance of different prediction techniques. Our work provides a benchmark of available software implementations: MCCE, MEAD, PROPKA and UHBD. Combinatorial and regression analysis is also used in an attempt to find a consensus approach towards pKa prediction. The tendency of individual programs to over- or underpredict the pKa value is related to the underlying methodology of the individual programs. CONCLUSION: Overall, PROPKA is more accurate than the other three programs. Key to developing accurate predictive software will be a complete sampling of conformations accessible to protein structures.

Catalysis↗

PubMed2006

Statistical deconvolution of enthalpic energetic contributions to MHC-peptide binding affinity.

BACKGROUND: MHC Class I molecules present antigenic peptides to cytotoxic T cells, which forms an integral part of the adaptive immune response. Peptides are bound within a groove formed by the MHC heavy chain. Previous approaches to MHC Class I-peptide binding prediction have largely concentrated on the peptide anchor residues located at the P2 and C-terminus positions. RESULTS: A large dataset comprising MHC-peptide structural complexes was created by re-modelling pre-determined x-ray crystallographic structures. Static energetic analysis, following energy minimisation, was performed on the dataset in order to characterise interactions between bound peptides and the MHC Class I molecule, partitioning the interactions within the groove into van der Waals, electrostatic and total non-bonded energy contributions. CONCLUSION: The QSAR techniques of Genetic Function Approximation (GFA) and Genetic Partial Least Squares (G/PLS) algorithms were used to identify key interactions between the two molecules by comparing the calculated energy values with experimentally-determined BL50 data. Although the peptide termini binding interactions help ensure the stability of the MHC Class I-peptide complex, the central region of the peptide is also important in defining the specificity of the interaction. As thermodynamic studies indicate that peptide association and dissociation may be driven entropically, it may be necessary to incorporate entropic contributions into future calculations.

Binding Sites↗

PubMed2004

Clostridium perfringens epsilon-toxin shows structural similarity to the pore-forming toxin aerolysin.

Epsilon-toxin from Clostridium perfringens is a lethal toxin. Recent studies suggest that the toxin acts via an unusually potent pore-forming mechanism. Here we report the crystal structure of epsilon-toxin, which reveals structural similarity to aerolysin from Aeromonas hydrophila. Pore-forming toxins can change conformation between soluble and transmembrane states. By comparing the two toxins, we have identified regions important for this transformation.

Aeromonas↗

PubMed2004

A Fugu-Human Genome Synteny Viewer: web software for graphical display and annotation reports of synteny between Fugu genomic sequence and human genes.

A web server has been developed to access annotation and graphical reports of synteny and gene order between the Fugu genome and human genes. In this system, the assembled Fugu genomic sequences (also known as scaffolds) are annotated. The annotations for each Fugu scaffold are computed, stored and made publicly available. The annotations describe matches to human homologous genes. For each significant human gene match on the Fugu scaffold, the corresponding human chromosome map and measures of the significance of each match are given. The web-based server provides public access to these annotations and graphical displays of the results. The user is provided with a selection of views including a chromosome-colour-coded image and a table containing the details of the matches. The Fugu-Human Genome Synteny Viewer has been tested by comparing results with examples from a paper that includes a study of transcription factors, Fos and Jun encoding regions. The Fugu-human genome synteny views are available for each Fugu scaffold through the clonesearch web page located at the Fugu Genomics website (http://fugu.rfcgr.mrc.ac.uk/).

Animals↗

PubMed2003

Clostridium absonum alpha-toxin: new insights into clostridial phospholipase C substrate binding and specificity.

Clostridium absonum phospholipase C (Caa) is a 42.7 kDa protein, which shows 60% amino acid sequence identity with the Clostridium perfringens phospholipase C, or alpha-toxin (Cpa), and has been isolated from patients suffering from gas gangrene. We report the cloning and sequencing, purification, characterisation and crystal structure of the Caa enzyme. Caa had twice the phospholipid-hydrolysing (lecithinase) activity, 1.5 times the haemolytic activity and over seven times the activity towards phosphatidylcholine-based liposomes when compared with Cpa. However, the Caa enzyme had a lower activity than Cpa to the free (i.e. not in lipid bilayer) substrate para-nitrophenylphosphorylcholine, towards sphingomyelin-based liposomes and showed half the cytotoxicity. The lethal dose (LD(50)) of Caa in mice was approximately twice that of Cpa. The crystal structure of Caa shows that the 72-93 residue loop is in a conformation different from those of previously determined open-form alpha-toxin structures. This conformational change suggests a role for W84 in membrane binding and a possible route of entry into the active site along a hydrophobic channel created by the re-arrangement of this loop. Overall, the properties of Caa are compatible with a role as a virulence-determinant in gas gangrene caused by C.absonum.

Amino Acid Sequence↗

PubMed2003

Estimation of weights and validation: a marginal likelihood approach.

The estimation of weights is quite an important aspect of the restrained refinement of macromolecular structures and related procedures such as the estimation of coordinate errors and structure validation using geometrical criteria. In principle, the method of maximum likelihood can be used for estimation of both atomic and weighting parameters. However, the low observation-to-parameter ratio in macromolecular refinement makes this kind of estimate of weighting parameters seriously biased; thus, the weighting parameters have traditionally been estimated separately from atomic parameters using a special technique, such as minimizing the free R factor. However, the variance of the latter estimate is large, as only a small portion of all data is used. In this work, an estimator of weights is proposed which is based on an approximation of a marginal likelihood function of the weighting parameters and which uses all the X-ray data. There is a known true value for the overall scaling coefficient for distance variances in restrained refinement and therefore the (maximum-likelihood) estimator for this coefficient may be used as a validation statistic.

Crystallography, X-Ray↗

PubMed2002

Crystal structure of the C. perfringens alpha-toxin with the active site closed by a flexible loop region.

Clostridium perfringens biotype A strains are the causative agents of gas-gangrene in man and are also implicated as etiological agents in sudden death syndrome in young domestic livestock. The main virulence factor produced by these strains is a zinc-dependent, phosphatidylcholine-preferring phospholipase C (alpha-toxin). The crystal structure of alpha-toxin, at pH 7.5, with the active site open and therefore accessible to substrate has previously been reported, as has calcium-binding to the C-terminal domain of the enzyme at pH 4.7. Here we focus on conformation changes in the N-terminal domain of alpha-toxin in crystals grown at acidic pH. These changes result in both the obscuring of the toxin active site and the loss of one of three zinc ions from it. Additionally, this "closed" form contains a small alpha helix, not present in the open structure, which hydrogen bonds to both the N and C-terminal domains. In conjunction with the previously reported findings that alpha-toxin can exist in active and inactive forms and that Thr74Ile and Phe69Cys substitutions markedly reduced the haemolytic activity of the enzyme, our work suggests that these loop conformations play a critical role in the activity of the toxin.

Binding Sites↗

PubMed2002

The first strain of Clostridium perfringens isolated from an avian source has an alpha-toxin with divergent structural and kinetic properties.

Clostridium perfringens alpha-toxin is a 370-residue, zinc-dependent, phospholipase C that is the key virulence determinant in gas gangrene. It is also implicated in the pathogenesis of sudden death syndrome in young animals and necrotic enteritis in chickens. Previously characterized alpha-toxins from different strains of C. perfringens are almost identical in sequence and biochemical properties. We describe the cloning, nucleotide sequencing, expression, characterization, and crystal structure of alpha-toxin from an avian strain, SWan C. perfringens (SWCP), which has a large degree of sequence variation and altered substrate specificity compared to these strains. The structure of alpha-toxin from strain CER89L43 has been previously reported in open (active site accessible to substrate) and closed (active site obscured by loop movements) conformations. The SWCP structure is in an open-form conformation, with three zinc ions in the active site. This is the first example of an open form of alpha-toxin crystallizing without the addition of divalent cations to the crystallization buffer, indicating that the protein can retain three zinc ions bound in the active site. The topology of the calcium binding site formed by residues 269, 271, 336, and 337, which is essential for membrane binding, is significantly altered in comparison with both the open and closed alpha-toxin structures. We are able to relate these structural changes to the different substrate specificity and membrane binding properties of this divergent alpha-toxin. This will provide essential information when developing an effective vaccine that will protect against C. perfringens infection in a wide range of domestic livestock.

Amino Acid Sequence↗

PubMedDate not supplied

A novel predictive technique for the MHC class II peptide-binding interaction.

Antigenic peptide is presented to a T-cell receptor through the formation of a stable complex with a Major Histocompatibility Complex (MHC) molecule. Various predictive algorithms have been developed to estimate a peptide's capacity to form a stable complex with a given MHC Class II allele, a technique integral to the strategy of vaccine design. These have previously incorporated such computational techniques as quantitative matrices and neural networks. We have developed a novel predictive technique that uses molecular modeling of predetermined crystal structures to estimate the stability of an MHC Class II peptide complex. This is the 1st structure-based technique, as previous methods have been based on binding data. ROC curves are used to quantify the accuracy of the molecular modeling technique. The novel predictive technique is found to be comparable with the best predictive software currently available.

Alleles↗