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F E Cohen

Publications and source records attributed to F E Cohen.

At least 127 records · Page 7Linked to original sources
PubMed1995

Protein conformational landscapes: energy minimization and clustering of a long molecular dynamics trajectory.

Using energy minimization and cluster analysis, we have analyzed a 1020 ps molecular dynamics trajectory of solvated bovine pancreatic trypsin inhibitor. Elucidation of conformational substates in this way both illustrates the degree of conformational convergence in the simulation and reduces the structural data to a tractable subset. The relative movement of structures upon energy minimization was used to estimate the sizes of features on the protein potential energy surface. The structures were analyzed using their pairwise root-mean-square C alpha deviations, which gave a global measure of conformational changes that would not be apparent by monitoring single degrees of freedom. At time scales of 0.1 ps, energy minimization detected sharp transitions between energy minima separated by 0.1 A rms deviation. Larger conformational clusters containing these smaller minima and separated by 0.25 A were seen at 1 ps time scales. Both of these small features of the conformational landscape were characterized by movements in loop regions associated with small, correlated backbone dihedral angle shifts. On a nanosecond time scale, the main features of the protein energy landscape were clusters separated by over 0.7 A rms deviation, with only seven of these substates visited over the 1 ns trajectory. These substrates, discernible both before and after energy minimization, differ mainly in a monotonic pivot of the loop residues 11-18 over the course of the simulation. This loop contains lysine 17, which specifically binds to trypsin in the active site. The trajectory did not return to previously visited clusters, indicating that this trajectory has not been shown to have completely sampled the conformational substates available to it. Because the apparent convergence to a single region of conformation space depends on both the time scale of observation and the size of the conformational features examined, convergence must be operationally defined within the context of the simulation.

Animals↗
PubMed1995

Evaluation of current techniques for ab initio protein structure prediction.

The results of a protein structure prediction contest are reviewed. Twelve different groups entered predictions on 14 proteins of known sequence whose structures had been determined but not yet disseminated to the scientific community. Thus, these represent true tests of the current state of structure prediction methodologies. From this work, it is clear that accurate tertiary structure prediction is not yet possible. However, protein fold and motif prediction are possible when the motif is recognizable similar to another known structure. Internal symmetry and the information inherent in an aligned family of homologous sequences facilitate predictive efforts. Novel folds remain a major challenge for prediction efforts.

Amino Acid Sequence↗
PubMed1995

Theoretical studies of sequence effects on the conformational properties of a fragment of the prion protein: implications for scrapie formation.

BACKGROUND: Prion diseases are neurodegenerative disorders that appear to be due to a conformational change, involving the conversion of alpha-helices in the normal, cellular isoform of the prion protein (PrPC) to beta-structure in the infectious scrapie form (PrPSc). One form of Gerstmann-Sträussler-Scheinker syndrome (GSS), an inherited prion disease, is caused by mutation of Ala117 of PrPC to Val. We therefore set out to evaluate the effects of this mutation on the stability of the PrPC form. RESULTS: We have performed molecular dynamics simulations of a portion of the PrPC sequence (residues 109-122, termed H1) that is proposed to figure prominently in the conversion of PrPC to PrPSc. In particular, beginning with H1 in the alpha-helical state, the conformational consequences of sequence changes at position 117 were investigated for six hydrophobic mutations. Of these, only the Val mutation was helix-destabilizing. Portions of this mutant peptide adopted and retained an extended conformation during a 2 ns simulation of the peptide in water. CONCLUSIONS: The conformational transitions and structures observed in the simulation of the mutant peptide with Val at position 117 provide insight into the possible early steps in the conversion of PrPC to PrPSc.

Animals↗
PubMed1995

Proposed structure of the A domains of factor VIII by homology modelling.

We have predicted a structure for the three A domains of blood coagulation factor VIII by virtue of their homology to blue copper-binding proteins. This structure, consisting of six beta-barrels, is arranged in a triangular configuration with a single type II copper-binding site linking the A1 and A3 domains.

Amino Acid Sequence↗
PubMed1994

The origins of protein secondary structure. Effects of packing density and hydrogen bonding studied by a fast conformational search.

Globular proteins fold to create compact structures rich in alpha-helices and beta-sheets. While studies of cubic lattice models of simplified polypeptide chains have concluded that secondary structure is a necessary consequence of chain compactness, different conclusions have been reached from studies of off-lattice models of simplified chains. In an attempt to resolve this controversy, we study an all-atom off-lattice model of a protein subject to a variety of simplified energy functions. A Monte Carlo simulated annealing algorithm is used to search conformational space quickly. The algorithm uses pivot-type moves in which a residue is selected at random and the values of its main-chain dihedral angles are changed. The energy function used to accept or reject moves is taken to be either a term proportional to the volume occupied by a structure (to mimic the hydrophobic effect), a term proportional to the energy of main-chain hydrogen bonding, or a combination of these two terms. Secondary structure content is evaluated using several different definitions. For all the definitions used, compactness alone produces a 10% increase in secondary structure content. However, this is a small fraction of the secondary structure observed in native protein structures. Structures produced by minimizing the hydrogen bond energy have extensive secondary structure but are not densely packed. Structures having both the high density of native structures and extensive secondary structure are produced by minimizing combinations of the volume and hydrogen bond energy terms. Our results emphasize the close relationship between secondary structure and the geometry of main-chain hydrogen bonding. The results are consistent with a description of protein folding in which the hydrophobic effect favors dense packing while hydrogen bonding determines the specific local geometry which generates secondary structure. To make an analogy with lattice studies of packing density and secondary structure, it seems that hydrophobicity provides the packing density while hydrogen bonding provides the lattice.

Algorithms↗
PubMed1994

Characterization of a Plasmodium vivax cysteine proteinase gene identifies uniquely conserved amino acids that may mediate the substrate specificity of malarial hemoglobinases.

The gene encoding a cysteine proteinase of the human malaria parasite Plasmodium vivax has been identified and characterized. The sequence predicted by the proteinase gene shares several unique features with the sequences of two recently characterized cysteine proteinases of other malarial species. These features include the conservation of a number of amino acids that are predicted, based on a recently devised model for the related Plasmodium falciparum cystine proteinase, to be located near the enzyme's active site. We hypothesize that these residues have been conserved to maintain optimal proteolytic specificity in the hydrolysis of globin by malaria parasites.

Amino Acid Sequence↗
PubMed1994

Proposed three-dimensional structure for the cellular prion protein.

Prion diseases are a group of neurodegenerative disorders in humans and animals that seem to result from a conformational change in the prion protein (PrP). Utilizing data obtained by circular dichroism and infrared spectroscopy, computational studies predicted the three-dimensional structure of the cellular form of PrP (PrPc). A heuristic approach consisting of the prediction of secondary structures and of an evaluation of the packing of secondary elements was used to search for plausible tertiary structures. After a series of experimental and theoretical constraints were applied, four structural models of four-helix bundles emerged. A group of amino acids within the four predicted helices were identified as important for tertiary interactions between helices. These amino acids could be essential for maintaining a stable tertiary structure of PrPc. Among four plausible structural models for PrPc, the X-bundle model seemed to correlate best with 5 of 11 known point mutations that segregate with the inherited prion diseases. These 5 mutations cluster around a central hydrophobic core in the X-bundle structure. Furthermore, these mutations occur at or near those amino acids which are predicted to be important for helix-helix interactions. The three-dimensional structure of PrPc proposed here may not only provide a basis for rationalizing mutations of the PrP gene in the inherited prion diseases but also guide design of genetically engineered PrP molecules for further experimental studies.

Amino Acid Sequence↗
PubMed1994

Spectroscopic characterization of conformational differences between PrPC and PrPSc: an alpha-helix to beta-sheet transition.

Although no chemical modifications have been found to distinguish the cellular prion protein PrPC from its infectious analogue PrPSc, spectroscopic methods such as Fourier transform infrared (FTIR) spectroscopy reveal a major conformational difference. PrPC is rich in alpha-helix but is devoid of beta-sheet, whereas PrPSc is high in beta-sheet. N-terminal truncation of PrPSc by limited proteolysis does not destroy infectivity but it increases the beta-sheet content and shifts the FTIR absorption to lower frequencies, typical of the cross beta-pleated sheets of amyloids. Thus the formation of PrPSc from PrPC involves a conformational transition in which one or more alpha-helical regions of the protein is converted to beta-sheet. This transition is mimicked by synthetic peptides, allowing predictions of domains of PrP involved in prion diseases.

Amino Acid Sequence↗
PubMed1994

Four helix bundle diversity in globular proteins.

Four helix bundles are a common structural motif that can be observed both independently and as components of larger folding units. We examined 221 globular proteins of known structure for possible four helix bundles. Previous computational studies of four helix bundles have placed arbitrary restrictions on interhelical packing angles. In this study we develop a geometric definition of four helix bundles based in part on solvent accessibility criteria that permits the removal of constraints on interhelical packing. Based on the observed pattern of interhelical angles, a bundle taxonomy is presented. This formalism should provide a useful categorization method for future structural studies of proteins rich in alpha-helices. The helix-helix interactions within bundles were studied in detail. Central residues, contact normals, and skew angles all were observed to have non-random distributions. A simple geometric model was developed for the helix-helix interface to explain these findings. Analysis of the helix-helix interaction data collected in this work confirms the importance of including skew angles in models of helix packing, and should improve the accuracy of combinatorial strategies for the prediction of the tertiary structure of all-helical proteins. Additionally, the geometric properties observed in globular proteins provide insight into the structural organization of membrane spanning proteins.

Animals↗
PubMed1994

Structure from function: screening structural models with functional data.

Structural constraints derived from different antibody epitopes on human growth hormone (hGH) were used to screen three-dimensional models of hGH that were generated by computer algorithms. Previously, alanine-scanning mutagenesis defined the residues that modulate binding to 21 different monoclonal antibodies to hGH. These functional epitopes were composed of 4-14 side chains whose alpha-carbons clustered within 4-23 A. Distance and topographic constraints for these functional epitopes were virtually the same as constraints derived from known x-ray structures of protein-antigen complexes. The constraints were used to evaluate about 1400 models of hGH that were computer-generated by a secondary-structure prediction and packing algorithm. On average each functional epitope reduced the number of models in the pool by a factor of 2, so that 8 monoclonal antibodies could reduce the number of possible models to < 10. The average root-mean-square deviation of alpha-carbon coordinates between the x-ray structure and either the pool of starting models or final models ranged from 13 to 16 A or 4 to 7 A, respectively, depending on the pool of starting models and the level of constraints imposed. All of the final models had the correct folding topography, and the best model was within 3.8 A root-mean-square deviation of the x-ray coordinates. This model was as close as it could have been because the models were built by using ideal helices and those in the x-ray structure are not. Our studies suggest that epitope mapping data can effectively screen structural models and, when coupled to predictive algorithms, can help to generate low-resolution models of a protein.

Amino Acid Sequence↗
PubMed1994

Conformation of CD4-derived cyclic hexapeptides by NMR and molecular dynamics.

Two cyclic hexapeptides, cyclo[Ala1-D-Ala2-Ser3-Phe4-Gly5-Ser6] and cyclo[Ala1-Gly2-Ser3-Phe4-Gly5-Ser6], derived from the loop portion of the C'C" ridge of CD4, were characterized by high-resolution nmr spectroscopy and simulated annealing studies. In DMSO-d6 both of these peptides display a single conformer on the nmr time scale with two intramolecular H-bond (1<--4) stabilized beta-turns at positions 2-3 and 5-6. The nmr derived distance constraints were used in simulated annealing calculations to generate the solution structures. These structures adopt energetically comparable conformational substates that are not resolvable on the nmr time scale. In aqueous solution, the H-bond stabilized beta-turn conformation for cyclo[Ala-D-Ala-Ser-Phe-Gly-Ser] is no longer the predominant structural form. Structures generated using molecular dynamics simulations with no experimental constraints were compared with those from nmr analysis. The correlation between these two sets of structures allows the use of molecular simulations as a predictive tool for the conformational analysis of small peptides.

Amino Acid Sequence↗
PubMed1994

GRAFTER: a computational aid for the design of novel proteins.

The GRAFTER suite of programs provides geometric search and evaluation functions that simplify and automate the process of identifying the best scaffolds for a particular structural motif. Three application of the GRAFTER suite are presented. Potential grafts between lambda repressor and 434 repressor were identified that should change the DNA binding specificity of these repressors. These results are compared with site-directed mutagenesis experiments that have been shown to alter repressor-DNA binding specificity. Next, 26 loops from antibody structures were grouped into families of similar structure. Grafts of antibody loops onto a pre-existing scaffold are an essential component of antibody humanization. Finally, interleukin (IL)-4 was searched as a scaffold that might accept the graft of a five residue epitope from human growth hormone (hGH). The existence of a crystal structure of the hGH-hGH receptor complex, extensive mutagenesis studies of the hGH residues that contribute to the energetics of ligand-receptor interactions and the gross structural homology between hGH and IL-4 make this an appealing computational target. The approach presented here could aid the development of novel enzymes and binding proteins.

Antibodies↗
PubMed1993

Computer-assisted drug discovery--a review.

A structure-based approach to new drug development is an attractive alternative to the traditional paradigm of drug discovery through screening. The elements of this approach are reviewed, with emphasis on the use of homology-built model structures. Two examples, proteases from the organisms that cause malaria and schistosomiasis, illustrate both the model-building process and the feasibility of using such models to computationally screen and identify compounds that inhibit their targets in the low micromolar range.

Computers↗
PubMed1993

Conversion of alpha-helices into beta-sheets features in the formation of the scrapie prion proteins.

Prions are composed largely, if not entirely, of prion protein (PrPSc in the case of scrapie). Although the formation of PrPSc from the cellular prion protein (PrPC) is a post-translational process, no candidate chemical modification was identified, suggesting that a conformational change features in PrPSc synthesis. To assess this possibility, we purified both PrPC and PrPSc by using nondenaturing procedures and determined the secondary structure of each. Fourier-transform infrared (FTIR) spectroscopy demonstrated that PrPC has a high alpha-helix content (42%) and no beta-sheet (3%), findings that were confirmed by circular dichroism measurements. In contrast, the beta-sheet content of PrPSc was 43% and the alpha-helix 30% as measured by FTIR. As determined in earlier studies, N-terminally truncated PrPSc derived by limited proteolysis, designated PrP 27-30, has an even higher beta-sheet content (54%) and a lower alpha-helix content (21%). Neither PrPC nor PrPSc formed aggregates detectable by electron microscopy, while PrP 27-30 polymerized into rod-shaped amyloids. While the foregoing findings argue that the conversion of alpha-helices into beta-sheets underlies the formation of PrPSc, we cannot eliminate the possibility that an undetected chemical modification of a small fraction of PrPSc initiates this process. Since PrPSc seems to be the only component of the "infectious" prion particle, it is likely that this conformational transition is a fundamental event in the propagation of prions.

Animals↗
PubMed1993

Erythropoietin structure-function relationships. Mutant proteins that test a model of tertiary structure.

On the basis of its primary sequence and the location of its disulfide bonds, we propose a structural model of the erythropoietic hormone erythropoietin (Epo) which predicts a four alpha-helical bundle motif, in common with other cytokines. In order to test this model, site-directed mutants were prepared by high level transient expression in Cos7 cells and analyzed by a radioimmuno assay and by bioassays utilizing mouse and human Epo-dependent cell lines. Deletions of 5 to 8 residues within predicted alpha-helices resulted in the failure of export of the mutant protein from the cell. In contrast, deletions at the NH2 terminus (delta 2-5), the COOH terminus (delta 163-166), or in predicted interhelical loops (AB: delta 32-36, delta 53-57; BC: delta 78-82; CD: delta 111-119) resulted in the export of immunologically detectable Epo muteins that were biologically active. The mutein delta 48-52 could be readily detected by radioimmunoassay but had markedly decreased biological activity. However, replacement of each of these deleted residues by serine resulted in Epo muteins with full biological activity. Replacement of Cys29 and Cys33 by tyrosine residues also resulted in the export of fully active Epo. Therefore, this small disulfide loop is not critical to Epo's stability or function. The properties of the muteins that we tested are consistent with our proposed model of tertiary structure.

Amino Acid Sequence↗
PubMed1993

Structure-based inhibitor design by using protein models for the development of antiparasitic agents.

The lack of an experimentally determined structure of a target protein frequently limits the application of structure-based drug design methods. In an effort to overcome this limitation, we have investigated the use of computer model-built structures for the identification of previously unknown inhibitors of enzymes from two major protease families, serine and cysteine proteases. We have successfully used our model-built structures to identify computationally and to confirm experimentally the activity of nonpeptidic inhibitors directed against important enzymes in the schistosome [2-(4-methoxybenzoyl)-1-naphthoic acid, Ki = 3 microM] and malaria (oxalic bis[(2-hydroxy-1-naphthylmethylene)hydrazide], IC50 = 6 microM) parasite life cycles.

Animals↗