Biomedical subjects

Emil Alexov

Publications and source records attributed to Emil Alexov.

14 recordsLinked to original sources

PubMed2007

Poisson-Boltzmann calculations of nonspecific salt effects on protein-protein binding free energies.

The salt dependence of the binding free energy of five protein-protein hetero-dimers and two homo-dimers/tetramers was calculated from numerical solutions to the Poisson-Boltzmann equation. Overall, the agreement with experimental values is very good. In all cases except one involving the highly charged lactoglobulin homo-dimer, increasing the salt concentration is found both experimentally and theoretically to decrease the binding affinity. To clarify the source of salt effects, the salt-dependent free energy of binding is partitioned into screening terms and to self-energy terms that involve the interaction of the charge distribution of a monomer with its own ion atmosphere. In six of the seven complexes studied, screening makes the largest contribution but self-energy effects can also be significant. The calculated salt effects are found to be insensitive to force-field parameters and to the internal dielectric constant assigned to the monomers. Nonlinearities due to high charge densities, which are extremely important in the binding of proteins to negatively charged membrane surfaces and to nucleic acids, make much smaller contributions to the protein-protein complexes studied here, with the exception of highly charged lactoglobulin dimers. Our results indicate that the Poisson-Boltzmann equation captures much of the physical basis of the nonspecific salt dependence of protein-protein complexation.

Binding Sites↗

PubMed2006

Calculation of pKas in RNA: on the structural origins and functional roles of protonated nucleotides.

pK(a) calculations based on the Poisson-Boltzmann equation have been widely used to study proteins and, more recently, DNA. However, much less attention has been paid to the calculation of pK(a) shifts in RNA. There is accumulating evidence that protonated nucleotides can stabilize RNA structure and participate in enzyme catalysis within ribozymes. Here, we calculate the pK(a) shifts of nucleotides in RNA structures using numerical solutions to the Poisson-Boltzmann equation. We find that significant shifts are predicted for several nucleotides in two catalytic RNAs, the hairpin ribozyme and the hepatitis delta virus ribozyme, and that the shifts are likely to be related to their functions. We explore how different structural environments shift the pK(a)s of nucleotides from their solution values. RNA structures appear to use two basic strategies to shift pK(a)s: (a) the formation of compact structural motifs with structurally-conserved, electrostatic interactions; and (b) the arrangement of the phosphodiester backbone to focus negative electrostatic potential in specific regions.

Catalysis↗

PubMed2006

PROTCOM: searchable database of protein complexes enhanced with domain-domain structures.

The database of protein complexes (PROTCOM) is a compilation of known 3D structures of protein-protein complexes enriched with artificially created domain-domain structures using the available entries in the Protein Data Bank. The domain-domain structures are generated by parsing single chain structures into loosely connected domains and are important features of the database. The database (http://www.ces.clemson.edu/compbio/protcom) could be used for benchmarking purposes of the docking and other algorithms for predicting 3D structures of protein-protein complexes. The database can be utilized as a template database in the homology or threading methods for modeling the 3D structures of unknown protein-protein complexes. PROTCOM provides the scientific community with an integrated set of tools for browsing, searching, visualizing and downloading a pool of protein complexes. The user is given the option to select a subset of entries using a combination of up to 10 different criteria. As on July 2006 the database contains 1770 entries, each of which consists of the known 3D structures and additional relevant information that can be displayed either in text-only or in visual mode.

Computer Graphics↗

PubMed2006

Predicting 3D structures of transient protein-protein complexes by homology.

The paper reports a homology based approach for predicting the 3D structures of full length hetero protein complexes. We have created a database of templates that includes structures of hetero protein-protein complexes as well as domain-domain structures (), which allowed us to expand the template pool up to 418 two-chain entries (at 40% sequence identity). Two protocols were tested-a protocol based on position specific Blast search (Protocol-I) and a protocol based on structural similarity of monomers (Protocol-II). All possible combinations of two monomers (350,284 pairs) in the ProtCom database were subjected to both protocols to predict if they form complexes. The predictions were benchmarked against the ProtCom database resulting to false-true positives ratios of approximately 5:1 and approximately 7:1 and recovery of 19% and 86%, respectively for protocols I and II. From 350,284 trials Protocol-I made only approximately 500 wrong predictions resulting to 0.5% error. In addition, though it was shown that artificially created domain-domain structures can in principle be good templates for modeling full length protein complexes, more sensitive methods are needed to detect homology relations. The quality of the models was assessed using two different criteria such as interfacial residues and overall RMSD. It was found that there is no correlation between these two measures. In many cases the interface residues were predicted correctly, but the overall RMSD was over 6 A and vice versa.

Algorithms↗

PubMed2006

Electrostatic properties of protein-protein complexes.

Statistical electrostatic analysis of 37 protein-protein complexes extracted from the previously developed database of protein complexes (ProtCom, http://www.ces.clemson.edu/compbio/protcom) is presented. It is shown that small interfaces have a higher content of charged and polar groups compared to large interfaces. In a vast majority of the cases the average pKa shifts for acidic residues induced by the complex formation are negative, indicating that complex formation stabilizes their ionizable states, whereas the histidines are predicted to destabilize the complex. The individual pKa shifts show the same tendency since 80% of the interfacial acidic groups were found to lower their pKas, whereas only 25% of histidines raise their pKa upon the complex formation. The interfacial groups have been divided into three sets according to the mechanism of their pKa shift, and statistical analysis of each set was performed. It was shown that the optimum pH values (pH of maximal stability) of the complex tend to be the same as the optimum pH values of the complex components. This finding can be used in the homology-based prediction of the 3D structures of protein complexes, especially when one needs to evaluate and rank putative models. It is more likely for a model to be correct if both components of the model complex and the entire complex have the same or at least similar values of the optimum pH.

Binding Sites↗

PubMed2006

Coating single-walled carbon nanotubes with phospholipids.

Single-walled carbon nanotubes (SWNTs), being hydrophobic by nature, aggregate in water to form large bundles. However, isolated SWNTs possess unique physical and chemical properties that are desirable for sensing and biological applications. Conventionally isolated SWNTs can be obtained by wrapping the tubes with biopolymers or surfactants. The binding modes proposed for these solubilization schemes, however, are less than comprehensive. Here we characterize the efficacies of solubilizing SWNTs through various types of phospholipids and other amphiphilic surfactants. Specifically, we demonstrate that lysophospholipids, or single-chained phospholipids offer unprecedented solubility for SWNTs, while double-chained phospholipids are ineffective in rendering SWNTs soluble. Using transmission electron microscopy (TEM) we show that lysophospholipids wrap SWNTs as striations whose size and regularity are affected by the polarity of the lysophospholipids. We further show that wrapping is only observed when SWNTs are in the lipid phase and not the vacuum phase, suggesting that the environment has a pertinent role in the binding process. Our findings shed light on the debate over the binding mechanism of amphiphilic polymers and cylindrical nanostructures and have implications on the design of novel supramolecular complexes and nanodevices.

Binding Sites↗

PubMed2005

Characterizing a partially folded intermediate of the villin headpiece domain under non-denaturing conditions: contribution of His41 to the pH-dependent stability of the N-terminal subdomain.

The contribution of interactions involving the imidazole ring of His41 to the pH-dependent stability of the villin headpiece (HP67) N-terminal subdomain has been investigated by nuclear magnetic resonance (NMR) spin relaxation. NMR-derived backbone N-H order parameters (S2) for wild-type (WT) HP67 and H41Y HP67 indicate that reduced conformational flexibility of the N-terminal subdomain in WT HP67 is due to intramolecular interactions with the His41 imidazole ring. These interactions, together with desolvation effects, contribute to significantly depress the pKa of the buried imidazole ring in the native state. 15N R1rho relaxation dispersion data indicate that WT HP67 populates a partially folded intermediate state that is 10.9 kJ mol(-1) higher in free energy than the native state under non-denaturing conditions at neutral pH. The partially folded intermediate is characterized as having an unfolded N-terminal subdomain while the C-terminal subdomain retains a native-like fold. Although the majority of the residues in the N-terminal subdomain sample a random-coil distribution of conformations, deviations of backbone amide 1H and 15N chemical shifts from canonical random-coil values for residues within 5A of the His41 imidazole ring indicate that a significant degree of residual structure is maintained in the partially folded ensemble. The pH-dependence of exchange broadening is consistent with a linear three-state exchange model whereby unfolding of the N-terminal subdomain is coupled to titration of His41 in the partially folded intermediate with a pKa,I=5.69+/-0.07. Although maintenance of residual interactions with the imidazole ring in the unfolded N-terminal subdomain appears to reduce pKa,I compared to model histidine compounds, protonation of His41 disrupts these interactions and reduces the difference in free energy between the native state and partially folded intermediate under acidic conditions. In addition, chemical shift changes for residues Lys70-Phe76 in the C-terminal subdomain suggest that the HP67 actin binding site is disrupted upon unfolding of the N-terminal subdomain, providing a potential mechanism for regulating the villin-dependent bundling of actin filaments.

Animals↗

PubMed2005

Comparative study of generalized born models: Born radii and peptide folding.

In this study, we have implemented four analytical generalized Born (GB) models and investigated their performance in conjunction with the GROMOS96 force field. The four models include that of Still and co-workers, the HCT model of Cramer, Truhlar, and co-workers, a modified form of the AGB model of Levy and co-workers, and the GBMV2 model of Brooks and co-workers. The models were coded independently and implemented in the GROMOS software package and in TINKER. They were compared in terms of their ability to reproduce the results of Poisson-Boltzmann (PB) calculations and in their performance in the ab initio peptide folding of two peptides, one that forms a beta-hairpin in solution and one that forms an alpha-helix. In agreement with previous work, the GBMV2 model is most successful in reproducing PB results while the other models tend to underestimate the effective Born radii of buried atoms. In contrast, stochastic dynamics simulations on the folding of the two peptides, the C-terminus beta-hairpin of the B1 domain of protein G and the alanine-based alpha-helical peptide 3K(I), suggest that the simpler GB models are more effective in sampling conformational space. Indeed, the Still model used in conjunction with the GROMOS96 force field is able to fold the hairpin peptide to a native-like structure without the benefit of enhanced sampling techniques. This is due in part to the properties of the united-atom GROMOS96 force field which appears to be more flexible, and hence to sample more efficiently, than force fields such as OPLSAA. Our results suggest a general strategy which involves using different combinations of force fields and solvent models in different applications, for example, using GROMOS96 and a simple GB model in sampling and OPLSAA and a more accurate GB model in refinement. The fact that various methods have been implemented in a unified way should facilitate the testing and subsequent use of different methods to evaluate conformational free energies in different applications. Our results also bear on some general issues involved in peptide folding and structure prediction which are addressed in the Discussion.

Alanine↗

PubMed2004

Calculating proton uptake/release and binding free energy taking into account ionization and conformation changes induced by protein-inhibitor association: application to plasmepsin, cathepsin D and endothiapepsin-pepstatin complexes.

The protein-inhibitor binding energies of enzymes are often pH dependent, and binding induces either proton uptake or proton release. The proton uptake/release and the binding energy for three complexes with available experimental data were numerically studied: pepstatin-cathepsin D, pepstatin-plasmepsin II and pepstatin-endothiapepsin. Very good agreement with the experimental data was achieved when conformational changes were taken into account. The role of the desolvation energy and the conformational changes was revealed by modeling the complex, the separated molecules in the complex conformation and the free molecules. It was shown that the conformational changes induced by the complex formation are as important for the proton transfer as the loss of solvation energy caused by the burial of interface residues. The residues responsible for the proton transfer were identified and their contribution to the proton uptake/release calculated. These residues were found to be scattered along the whole protein rather than being localized only at the active site. In the case of cathepsin D, these residues were found to be highly conserved among the cathepsin D sequences of other species. It was shown that conformation and ionization changes induced by the complex formation are critical for the correct calculation of the binding energy. Taking into account the electrostatics and the van der Waals (vdW) energies within the Boltzmann distribution of energies and allowing ionization and conformation changes to occur makes the calculated binding energy more realistic and closer to the experimental value. The interplay between electrostatic and vdW forces makes the pH dependence of the binding energy smoother, because the vdW force acts in reaction to the changes of the electrostatic energy. It was found that a small fraction of the ionizable groups remain uncharged in both the free and complexed molecules. The sequence and structural position of these groups aligns well within the three proteases, suggesting that these may have specific role.

Amino Acid Sequence↗

PubMed2004

Numerical calculations of the pH of maximal protein stability. The effect of the sequence composition and three-dimensional structure.

A large number of proteins, found experimentally to have different optimum pH of maximal stability, were studied to reveal the basic principles of their preference for a particular pH. The pH-dependent free energy of folding was modeled numerically as a function of pH as well as the net charge of the protein. The optimum pH was determined in the numerical calculations as the pH of the minimum free energy of folding. The experimental data for the pH of maximal stability (experimental optimum pH) was reproducible (rmsd = 0.73). It was shown that the optimum pH results from two factors - amino acid composition and the organization of the titratable groups with the 3D structure. It was demonstrated that the optimum pH and isoelectric point could be quite different. In many cases, the optimum pH was found at a pH corresponding to a large net charge of the protein. At the same time, there was a tendency for proteins having acidic optimum pHs to have a base/acid ratio smaller than one and vice versa. The correlation between the optimum pH and base/acid ratio is significant if only buried groups are taken into account. It was shown that a protein that provides a favorable electrostatic environment for acids and disfavors the bases tends to have high optimum pH and vice versa.

Aldose-Ketose Isomerases↗

PubMed2003

On the role of structural information in remote homology detection and sequence alignment: new methods using hybrid sequence profiles.

Structural alignments often reveal relationships between proteins that cannot be detected using sequence alignment alone. However, profile search methods based entirely on structural alignments alone have not been found to be effective in finding remote homologs. Here, we explore the role of structural information in remote homolog detection and sequence alignment. To this end, we develop a series of hybrid multidimensional alignment profiles that combine sequence, secondary and tertiary structure information into hybrid profiles. Sequence-based profiles are profiles whose position-specific scoring matrix is derived from sequence alignment alone; structure-based profiles are those derived from multiple structure alignments. We compare pure sequence-based profiles to pure structure-based profiles, as well as to hybrid profiles that use combined sequence-and-structure-based profiles, where sequence-based profiles are used in loop/motif regions and structural information is used in core structural regions. All of the hybrid methods offer significant improvement over simple profile-to-profile alignment. We demonstrate that both sequence-based and structure-based profiles contribute to remote homology detection and alignment accuracy, and that each contains some unique information. We discuss the implications of these results for further improvements in amino acid sequence and structural analysis.

Amino Acid Sequence↗

PubMed2003

Role of the protein side-chain fluctuations on the strength of pair-wise electrostatic interactions: comparing experimental with computed pK(a)s.

The effect of the protein side-chain fluctuations on the strength of electrostatic interactions was studied. The effect was modeled on 7 different crystal structures on the same enzyme as well as on 20 molecular dynamics snapshot structures. It was shown that the side-chain flexibility affects predominantly the magnitude of the strong pair-wise interactions, that is, the pair-wise interaction among ion pairs, and practically does not affect the interactions with the rest of the protein. This was used to suggest a correction function that should be applied to the original pair-wise electrostatic interaction to mimic the effects of the fluctuations. The procedure is applied on three ion pairs identified in lysozyme. It was shown that sampling different side-chain rotamers and modifying the strength of the pair-wise interaction energies makes calculated pK(a)s less sensitive to the fluctuations of the structure and improves the prediction accuracy.

Amino Acids↗

PubMed2003

Using multiple structure alignments, fast model building, and energetic analysis in fold recognition and homology modeling.

We participated in the fold recognition and homology sections of CASP5 using primarily in-house software. The central feature of our structure prediction strategy involved the ability to generate good sequence-to-structure alignments and to quickly transform them into models that could be evaluated both with energy-based methods and manually. The in-house tools we used include: a) HMAP (Hybrid Multidimensional Alignment Profile)-a profile-to-profile alignment method that is derived from sequence-enhanced multiple structure alignments in core regions, and sequence motifs in non-structurally conserved regions. b) NEST-a fast model building program that applies an "artificial evolution" algorithm to construct a model from a given template and alignment. c) GRASP2-a new structure and alignment visualization program incorporating multiple structure superposition and domain database scanning modules. These methods were combined with model evaluation based on all atom and simplified physical-chemical energy functions. All of these methods were under development during CASP5 and consequently a great deal of manual analysis was carried out at each stage of the prediction process. This interactive model building procedure has several advantages and suggests important ways in which our and other methods can be improved, examples of which are provided.

Algorithms↗

PubMed2002

Rapid grid-based construction of the molecular surface and the use of induced surface charge to calculate reaction field energies: applications to the molecular systems and geometric objects.

This article describes a number of algorithms that are designed to improve both the efficiency and accuracy of finite difference solutions to the Poisson-Boltzmann equation (the FDPB method) and to extend its range of application. The algorithms are incorporated in the DelPhi program. The first algorithm involves an efficient and accurate semianalytical method to map the molecular surface of a molecule onto a three-dimensional lattice. This method constitutes a significant improvement over existing methods in terms of its combination of speed and accuracy. The DelPhi program has also been expanded to allow the definition of geometrical objects such as spheres, cylinders, cones, and parallelepipeds, which can be used to describe a system that may also include a standard atomic level depiction of molecules. Each object can have a different dielectric constant and a different surface or volume charge distribution. The improved definition of the surface leads to increased precision in the numerical solutions of the PB equation that are obtained. A further improvement in the precision of solvation energy calculations is obtained from a procedure that calculates induced surface charges from the FDPB solutions and then uses these charges in the calculation of reaction field energies. The program allows for finite difference grids of large dimension; currently a maximum of 571(3) can be used on molecules containing several thousand atoms and charges. As described elsewhere, DelPhi can also treat mixed salt systems containing mono- and divalent ions and provide electrostatic free energies as defined by the nonlinear PB equation.

Journal Article↗