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Results for “Homology modelling”

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At least 37 records · Page 2Linked to original sources

PubMed2000

Estimating local backbone structural deviation in homology models.

After the atomic coordinates themselves, the most important data in a homology model are the spatial reliability estimates associated with each of the atoms (atom annotation). Recent blind homology modeling predictions have demonstrated that principally correct sequence-structure alignments are achievable to sequence identities as low as 25% [Martin, A.C., MacArthur, M.W., Thornton, J.M., 1997. Assessment of comparative modeling in CASP2. Proteins Suppl(1), 14-28]. The locations and extent of spatial deviations in the backbone between correctly aligned homologous protein structures remained very poorly estimated however, and these errors were the cause of errant loop predictions [Abagyan, R., Batalov, S., Cardozo, T., Totrov, M., Webber, J., Zhou, Y., 1997. Homology modeling with internal coordinate mechanics: deformation zone mapping and improvements of models via conformational search. Proteins Suppl(1), 29-37]. In order to derive accurate measures for local backbone deviations, we made a systematic study of static local backbone deviations between homologous pairs of protein structures. We found that 'through space' proximity to gaps and chain termini, local three-dimensional 'density', three-dimensional environment conservation, and B-factor of the template contribute to local deviations in the backbone in addition to local sequence identity. Based on these finding, we have identified the meaningful ranges of values within which each of these parameters correlates with static local backbone deviation and produced a combined scoring function to greatly improve the estimation of local backbone deviations. The optimized function has more than twice the accuracy of local sequence identity or B-factor alone and was validated in a recent blind structure prediction experiment. This method may be used to evaluate the utility of a preliminary homology model for a particular biological investigation (e.g. drug design) or to provide an improved starting point for molecular mechanics loop prediction methods.

Computer Simulation↗

PubMed2002

Homology modeling and molecular dynamics study of NAD-dependent glycerol-3-phosphate dehydrogenase from Trypanosoma brucei rhodesiense, a potential target enzyme for anti-sleeping sickness drug development.

Sleeping sickness and Chagas disease are among the most severe diseases in Africa as well as Latin America. These two diseases are caused by Trypanosoma spp. Recently, an enzyme of a glycolytic pathway, NAD-dependent glycerol-3-phosphate dehydrogenase, of Leishmania mexicana was crystallized and its structure determined by x-ray crystallography. This structure has offered an excellent template for modeling of the homologous enzymes from another Trypanosoma species. Here, a homology model of the T. brucei enzyme based on the x-ray structure of LmGPDH has been generated. This model was used as the starting point for molecular dynamics simulation in a water box. The analysis of the molecular dynamics trajectory indicates that the functionally important motifs have both a very stable secondary structure and tertiary arrangement.

Amino Acid Sequence↗

PubMed1994

Homology modelling of the catalytic domain of human furin. A model for the eukaryotic subtilisin-like proprotein convertases.

A model is presented for the three-dimensional structure of the catalytic domain of the human serine proteinase furin and its interaction with model substrates. This homology model is based on the crystal structures of subtilisin BPN' and thermitase in complex with the inhibitor eglin, and it also applies to other members of the eukaryotic subtilisin-like proprotein convertases. Predictions are made of the general protein fold, inserted loops, disulfide bonds, Ca(2+)-binding sites and salt bridges. A detailed prediction of the substrate-binding region attempts to explain the basis of specificity for multiple basic residues preceding the cleavage site. Specific acidic residues in the S1, S2 and S4 subsites of the substrate-binding region of furin are identified which appear to be of particular importance, while residues of the S2', S3, S5 and S6 subsites may also contribute to substrate binding. Based on this model, protein engineering can be employed not only to test the predicted enzyme-substrate interactions, as demonstrated for human furin, but, equally importantly, to design proprotein convertases with a desired specificity, or to design novel substrates or inhibitors.

Amino Acid Sequence↗

PubMed2003

Ligand-supported homology modelling of protein binding-sites using knowledge-based potentials.

A new approach, MOBILE, is presented that models protein binding-sites including bound ligand molecules as restraints. Initially generated, homology models of the target protein are refined iteratively by including information about bioactive ligands as spatial restraints and optimising the mutual interactions between the ligands and the binding-sites. Thus optimised models can be used for structure-based drug design and virtual screening. In a first step, ligands are docked into an averaged ensemble of crude homology models of the target protein. In the next step, improved homology models are generated, considering explicitly the previously placed ligands by defining restraints between protein and ligand atoms. These restraints are expressed in terms of knowledge-based distance-dependent pair potentials, which were compiled from crystallographically determined protein-ligand complexes. Subsequently, the most favourable models are selected by ranking the interactions between the ligands and the generated pockets using these potentials. Final models are obtained by selecting the best-ranked side-chain conformers from various models, followed by an energy optimisation of the entire complex using a common force-field. Application of the knowledge-based pair potentials proved efficient to restrain the homology modelling process and to score and optimise the modelled protein-ligand complexes. For a test set of 46 protein-ligand complexes, taken from the Protein Data Bank (PDB), the success rate of producing near-native binding-site geometries (rmsd<2.0A) with MODELLER is 70% when the ligand restrains the homology modelling process in its native orientation. Scoring these complexes with the knowledge-based potentials, in 66% of the cases a pose with rmsd <2.0A is found on rank 1. Finally, MOBILE has been applied to two case studies modelling factor Xa based on trypsin and aldose reductase based on aldehyde reductase.

Binding Sites↗

PubMed2004

Homology modelling and molecular dynamics simulations: comparative studies of human aquaporin-1.

The structures of the mammalian water transport protein Aqp1 and of its bacterial homologue GlpF enables us to test whether homology models can be used to explore relationships between structure, dynamics and function in mammalian transport proteins. Molecular dynamics simulations (totalling almost 40 ns) were performed starting from: the X-ray structure of Aqp1; a homology model of Aqp1 based on the GlpF structure; and intermediate resolution structures of Aqp1 derived from electron microscopy. Comparisons of protein RMSDs vs. time suggest that the homology models are of comparable conformational stability to the X-ray structure, whereas the intermediate resolution structures exhibit significant conformation drift. For simulations based on the X-ray structure and on homology models, the flexibility profile vs. residue number correlates well with the crystallographic B-values for each residue. In the simulations based on intermediate resolution structures, mobility of the highly conserved NPA loops is substantially higher than in the simulations based on the X-ray structure or the homology models. Pore radius profiles remained relatively constant in the X-ray and homology model simulations but showed substantial fluctuations (reflecting the higher NPA loop mobility) in the intermediate resolution simulations. The orientation of the dipoles of water molecules within the pore is of key importance in maintaining low proton permeability through Aqp1. This property seems to be quite robust to the starting model used in the simulation. These simulations suggest that homology models based on bacterial homologues may be used to derive functionally relevant information on the structural dynamics of mammalian transport proteins.

Aquaporin 1↗

PubMed2005

Structure-based drug discovery using GPCR homology modeling: successful virtual screening for antagonists of the alpha1A adrenergic receptor.

In this paper, we describe homology modeling of the alpha1A receptor based on the X-ray structure of bovine rhodopsin. The protein model has been generated by applying ligand-supported homology modeling, using mutational and ligand SAR data to guide the protein modeling procedure. We performed a virtual screening of the company's compound collection to test how well this model is suited to identify alpha1A antagonists. We applied a hierarchical virtual screening procedure guided by 2D filters and three-dimensional pharmacophore models. The ca. 23,000 filtered compounds were docked into the alpha1A homology model with GOLD and scored with PMF. From the top-ranked compounds, 80 diverse compounds were tested in a radioligand displacement assay. 37 compounds revealed K(i) values better than 10 microM; the most active compound binds with 1.4 nM to the alpha1A receptor. Our findings suggest that rhodopsin-based homology models may be used as the structural basis for GPCR lead finding and compound optimization.

Adrenergic alpha-1 Receptor Antagonists↗

PubMed2006

Homology modeling of G-protein-coupled receptors and implications in drug design.

G-protein-coupled receptors (GPCRs) are considered therapeutically important due to their involvement in a variety of processes governing several cellular functions, and their tractability as drug targets. A large percentage of drugs on the market, and in development stages, target the super family of the GPCRs. The enormous interest in GPCR drug design is, however, limited by the scarcity of structural information. The only GPCR for which a three dimensional (3D) structure is reported is bovine rhodopsin and it belongs to class A of the GPCR family. As a result, there has been considerable interest in alternative techniques, for example, homology modeling of GPCRs, in order to derive useful three dimensional models of other proteins for use in structure-based drug design. However, homology modeling of GPCRs is not straightforward, and encounters several problems, owing to the availability of a single structural template, as well as the low degree of sequence homology between the template and target sequences. There are several key issues which need to be considered during every stage of GPCR homology modeling, in order to derive reasonable 3D models. Homology modeling of GPCRs has been utilized increasingly in the past few years and has been successful, not only in furthering the understanding of ligand-protein interactions, but also in the identification of new and potent ligands. Thus, with the lessons learned from past experiences and new developments, homology modeling in case of GPCRs can be harnessed for developing more reliable three dimensional models. This, in turn, will provide better tools to use in structure-based drug design leading to the identification of novel and potent GPCR ligands for several therapeutic indications.

Amino Acid Sequence↗

PubMed2006

Secondary structure and 3D homology modeling of grass carp (Ctenopharyngodon idellus) major histocompatibility complex class I molecules.

No information to date is available on the structure of fish major histocompatibility complex (MHC) class I and beta2-microglobulin (beta2m) proteins. In the present study, grass carp (Ctenopharyngodon idellus) MHC class I (Ctid-MHC I) and beta(2)-microglobulin (Ctid-beta2m) genes were expressed as soluble maltose binding protein (MBP)-proteins and purified in a pMAL-p2X/Escherichia coli TB1 system. The expressed proteins were purified on amylase affinity columns followed by DEAE-Sepharose. The purified products were identified by Western blotting with anti-MBP polyclonal antibodies. The MBP-Ctid-MHC I and MBP-Ctid-beta2m were cleaved separately with Factor Xa, mixed together and purified on DEAE-Sepharose. The secondary structures were analyzed by circular dichroism (CD) spectrophotometry. The three-dimensional (3D) structure of their peptide-binding domain (PBD) was modeled based sequence homology. The sequence lengths of the alpha-helix, beta-sheet, turn, and random coil in the Ctid-MHC I protein were 79aa, 75aa, 20aa, and 99aa, respectively. In the 97aa of Ctid-beta2m, the contents of the alpha-helix, beta-sheet, turn, and random coil were 0aa, 41aa, 12aa, and 44aa, respectively. The Ctid-beta2m protein displayed a typical beta-sheet. Homology modeling of the Ctid-MHC I and Ctid-beta2m proteins demonstrated similarities with the structure of human MHC class I proteins.

Animals↗

PubMedDate not supplied

A homology model for rat mu class glutathione S-transferase 4-4.

Glutathione S-transferases (GSTs) are an important class of phase II (de)toxifying enzymes, catalyzing the conjugation of glutathione (GSH) to electrophilic species. Recently, a number of cytosolic GSTs was crystallized. In the present study, molecular modeling techniques have been used to derive a three-dimensional homology model for rat GST 4-4 based upon the crystal structure of rat GST 3-3, both members of the mu class. GST 3-3 and GST 4-4 isoenzymes share a sequence homology of 88%. GST 4-4 distinguishes itself from GST 3-3 in being much more efficient and stereoselective in the nucleophilic addition of GSH to epoxides and alpha,beta-unsaturated ketones. GST 3-3, however, is much more efficient in catalyzing nucleophilic aromatic substitution reactions. In this study, several known substrates of GST 4-4 were selected and their GSH conjugates docked into the active site of GST 4-4. GSH conjugates of phenanthrene 9(S),10(R)-oxide and 4,5-diazaphenanthrene 9(S),10(R)-oxide were docked into the active site of both GST 3-3 and GST 4-4. From these homology modeling and docking data, the difference in stereoselectivity between GST 3-3 and GST 4-4 for the R- and S-configured carbons of the oxirane moiety could be rationalized. The data acquired from a recently derived small molecule model for GST 4-4 substrates were compared with the results of the present protein homology model of GST 4-4. The energy optimized positions of the conjugates in the protein model agreed very well with the original relative positions of the substrates within the substrate model, confirming the usefulness of small molecule models in the absence of structural protein data. The protein homology model, together with the substrate model, will be useful to further rationalize the substrate selectivity of GST 4-4, and to identify new potential GST 4-4 substrates.

Amino Acid Sequence↗

PubMed2000

Sequence annotation of nuclear receptor ligand-binding domains by automated homology modeling.

The quality of three-dimensional homology models derived from protein sequences provides an independent measure of the suitability of a protein sequence for a certain fold. We have used automated homology modeling and model assessment tools to identify putative nuclear hormone receptor ligand-binding domains in the genome of Caenorhabditis elegans. Our results indicate that the availability of multiple crystal structures is crucial to obtaining useful models in this receptor family. The majority of annotated mammalian nuclear hormone receptors could be assigned to a ligand-binding domain fold by using the best model derived from any of four template structures. This strategy also assigned the ligand-binding domain fold to a number of C.elegans. sequences without prior annotation. Interestingly, the retinoic acid receptor crystal structure contributed most to the number of sequences that could be assigned to a ligand-binding domain fold. Several causes for this can be suggested, including the high quality of this protein structure in terms of our assessment tools, similarity between the biological function or ligand of this receptor and the modeled genes and gene duplication in C.elegans.

Animals↗

PubMed2000

Homology model directed alignment selection for comparative molecular field analysis: application to photosystem II inhibitors.

The use of a computational docking protocol in conjunction with a protein homology model to derive molecular alignments for Comparative Molecular Field Analysis (CoMFA) was examined. In particular, the DOCK program and a model of the herbicidal target site, photosystem II (PSII), was used to derive alignments for two PSII inhibitor training sets, a set of benzo- and napthoquinones and a set of butenanilides. The protein design software in the QUANTA molecular modeling package was used to develop a homology model of spinach PSII based on the reported amino acid sequence and the X-ray crystal structure of the purple bacterium reaction center. The model is very similar to other reported PSII protein homology models. DOCK was then used to derive alignments for CoMFA modeling by docking the inhibitors in the PSII binding pocket. The molecular alignments produced from docking yielded highly predictive CoMFA models. As a comparison, the more traditional atom-atom alignments of the same two training sets failed to produce predictive CoMFA models. The general utilities of this application for homology model refinement and as an alternative scoring method are discussed.

Amino Acid Sequence↗

PubMed1998

Homology modeling, model and software evaluation: three related resources.

MOTIVATION: Homology modeling is rapidly becoming the method of choice for obtaining three-dimensional coordinates for proteins because genome projects produce sequences at a much higher rate than NMR and X-ray laboratories can solve the three-dimensional structures. The quality of protein models will not be immediately clear to novices and support with the evaluation seems to be needed. Expert users are sometimes interested in evaluating the quality of modeling programs rather than the quality of the models themselves. RESULTS: Three servers have been made available to the scientific community: a homology modeling server, a model quality evaluation server and a server that evaluates models built of proteins for which the structure is already known, thereby implicitly evaluating the quality of the modeling program. AVAILABILITY: The modeling-related servers and several structure analysis servers are freely available at http://swift.embl-heidelberg.de/servers/ CONTACT: gert.vriend@embl-heidelberg.de

Computational Biology↗

PubMed2006

Probing the catalytic center of porcine aminoacylase 1 by site-directed mutagenesis, homology modeling and substrate docking.

Three-dimensional structural models of porcine aminoacylase 1 (pACY1) were constructed by homology modeling and aligning the structures of members of the M20 peptidase family. After energy minimization and quality evaluation, the best model from the homology modeling was chosen for docking with the best substrate (N-acetyl-L-methionine). The most reasonable binding mode was found among a large number of predicted complexes by using clustering analysis and screening with expert knowledge. Structural analysis revealed that the zinc ion is not likely to bind to the substrate, and that Arg348 and Glu146 play vital roles in binding and catalysis. In the site-directed mutagenesis experiments, mutation of His79, Asp112, Glu147, Arg348, and Glu146, resulted in significant reductions of specific activity, while the wild-type pACY1 overexpressed in Rosetta (DE3) had almost as high a specific activity as the native enzyme. On the basis of these observations, we proposed a revised catalytic mechanism for this metalloenzyme.

Amidohydrolases↗

PubMed2005

Access of the substrate to the active site of squalene and oxidosqualene cyclases: comparative inhibition, site-directed mutagenesis and homology-modelling studies.

Substrate access to the active-site cavity of squalene-hopene cyclase from Alicyclobacillus acidocaldarious and lanosterol synthase [OSC (oxidosqualene cyclase)] from Saccharomyces cerevisiae was studied by an inhibition, mutagenesis and homology-modelling approach. Crystal structure and homology modelling indicate that both enzymes possess a narrow constriction that separates an entrance lipophilic channel from the active-site cavity. The role of the constriction as a mobile gate that permits substrate passage was investigated by experiments in which critically located Cys residues, either present in native protein or inserted by site-directed mutagenesis, were labelled with specifically designed thiol-reacting molecules. Some amino acid residues of the yeast enzyme, selected on the basis of sequence alignment and a homology model, were individually replaced by residues bearing side chains of different lengths, charges or hydrophobicities. In some of these mutants, substitution severely reduced enzymatic activity and thermal stability. Homology modelling revealed that in these mutants some critical stabilizing interactions could no longer occur. The possible critical role of entrance channel and constriction in specific substrate recognition by eukaryotic OSC is discussed.

Amino Acid Sequence↗

PubMed2005

Homology model of the CDK1/cyclin B complex.

We describe a refined homology model of a CDK1/cyclin B complex that was previously used for the structure-based optimization of the Paullone class of inhibitors. The preliminary model was formed from the homologous regions of the deposited CDK2/cyclin A crystal structure. Further refinement of the CDK1/cyclin B complex was accomplished using molecular mechanics and hydropathic analysis with a protocol of constraints and local geometry searches. For the most part, our CKD1/cyclin B homology model is very similar to the high resolution CDK2/cyclin A crystal structure regarding secondary and tertiary features. However, minor discrepancies between the two kinase structures suggest the possibility that ligand design may be specifically tuned for either CDK1 or CDK2. Our examination of the CDK1/cyclin B model includes a comparison with the CDK2/cyclin A crystal structure in the PSTAIRE interface region, connecting portions to the ATP binding domain, as well as the ATP binding site itself.

Adenosine Triphosphate↗

PubMed2006

On the accuracy of homology modeling and sequence alignment methods applied to membrane proteins.

In this study, we investigate the extent to which techniques for homology modeling that were developed for water-soluble proteins are appropriate for membrane proteins as well. To this end we present an assessment of current strategies for homology modeling of membrane proteins and introduce a benchmark data set of homologous membrane protein structures, called HOMEP. First, we use HOMEP to reveal the relationship between sequence identity and structural similarity in membrane proteins. This analysis indicates that homology modeling is at least as applicable to membrane proteins as it is to water-soluble proteins and that acceptable models (with C alpha-RMSD values to the native of 2 A or less in the transmembrane regions) may be obtained for template sequence identities of 30% or higher if an accurate alignment of the sequences is used. Second, we show that secondary-structure prediction algorithms that were developed for water-soluble proteins perform approximately as well for membrane proteins. Third, we provide a comparison of a set of commonly used sequence alignment algorithms as applied to membrane proteins. We find that high-accuracy alignments of membrane protein sequences can be obtained using state-of-the-art profile-to-profile methods that were developed for water-soluble proteins. Improvements are observed when weights derived from the secondary structure of the query and the template are used in the scoring of the alignment, a result which relies on the accuracy of the secondary-structure prediction of the query sequence. The most accurate alignments were obtained using template profiles constructed with the aid of structural alignments. In contrast, a simple sequence-to-sequence alignment algorithm, using a membrane protein-specific substitution matrix, shows no improvement in alignment accuracy. We suggest that profile-to-profile alignment methods should be adopted to maximize the accuracy of homology models of membrane proteins.

Algorithms↗

PubMed2004

Homology-modeled structure of the yeast mitochondrial citrate transport protein.

We have used homology modeling to construct a three-dimensional model of the yeast mitochondrial citrate transport protein (CTP), based on the recently published x-ray crystal structure of another mitochondrial transport protein, the ADP/ATP carrier. Superposition of the backbone traces of the homology-modeled CTP onto the crystallographically determined ADP carrier structure indicates that the CTP transmembrane domains are well modeled (i.e., root mean square deviation of 0.94 A), whereas the loops facing the intermembrane space and the mitochondrial matrix are less certain (i.e., root mean square deviation values of 0.72-2.06 A). The homology-modeled CTP is consistent with our earlier de novo models of the transporter's transmembrane domains, with respect to residues which face into the transport path. Importantly, the resulting model is consistent with our previous experimental data obtained from measuring reactivity of 34 single cysteine mutants in transmembrane domains 3 and 4 with methanethiosulfonate reagents. The model also points to a likely dimer interface region. In conclusion, our data help to define the substrate translocation pathway in both the modeled CTP structure and the crystallographic ADP carrier structure.

Amino Acid Sequence↗

PubMed2006

A comparative study of available software for high-accuracy homology modeling: from sequence alignments to structural models.

An open question in protein homology modeling is, how well do current modeling packages satisfy the dual criteria of quality of results and practical ease of use? To address this question objectively, we examined homology-built models of a variety of therapeutically relevant proteins. The sequence identities across these proteins range from 19% to 76%. A novel metric, the difference alignment index (DAI), is developed to aid in quantifying the quality of local sequence alignments. The DAI is also used to construct the relative sequence alignment (RSA), a new representation of global sequence alignment that facilitates comparison of sequence alignments from different methods. Comparisons of the sequence alignments in terms of the RSA and alignment methodologies are made to better understand the advantages and caveats of each method. All sequence alignments and corresponding 3D models are compared to their respective structure-based alignments and crystal structures. A variety of protein modeling software was used. We find that at sequence identities >40%, all packages give similar (and satisfactory) results; at lower sequence identities (<25%), the sequence alignments generated by Profit and Prime, which incorporate structural information in their sequence alignment, stand out from the rest. Moreover, the model generated by Prime in this low sequence identity region is noted to be superior to the rest. Additionally, we note that DSModeler and MOE, which generate reasonable models for sequence identities >25%, are significantly more functional and easier to use when compared with the other structure-building software.

Algorithms↗