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M Karplus

Publications and source records attributed to M Karplus.

At least 55 records · Page 3Linked to original sources
PubMed1998

Specific amino acid recognition by aspartyl-tRNA synthetase studied by free energy simulations.

Specific amino acid binding by aminoacyl-tRNA synthetases is necessary for correct translation of the genetic code. To obtain insight into the origin of the specificity, the binding to aspartyl-tRNA synthetase (AspRS) of the negatively charged substrate aspartic acid and the neutral analogue asparagine are compared by use of molecular dynamics and free energy simulations. Simulations of the Asn-AspRS complex show that although Asn cannot bind in the same position as Asp, several possible positions exist 1.5 to 2 A away from the Asp site. The binding free energy of Asn in three of these positions was compared to that of Asp through alchemical free energy simulations, in which Asp is gradually mutated ito Asn in the complex with the enzyme. To correctly account for the electrostatic interactions in the system (including bulk solvent), a recently developed hybrid approach was used, in which the region of the mutation site is treated microscopically, whereas distant protein and solvent are treated by continuum electrostatics. Seven free energy simulations were performed in the protein and two in solution. The various Asn positions and orientations sampled at the Asn endpoints of the protein simulations yielded very similar free energy differences. The calculated Asp-->Asn free energy change is 79.8(+/-1.5) kcal/mol in solution and 95.1(+/-2.8) kcal/mol in the complex with the protein. Thus, the substrate Asp is predicted to bind much more strongly than Asn, with a binding free energy difference of 15.3 kcal/mol. This implies that erroneous binding of Asn by AspRS is highly improbable, and cannot account for any errors in the translation of the genetic code. Almost all of the protein contributions to the Asp versus Asn binding free energy difference arise from an arginine and a lysine residue that hydrogen bond to the substrate carboxylate group and an Asp and a Glu that hydrogen bond to these; all four amino acid residues are completely conserved in AspRSs. The protein effectively "solvates" the Asp side-chain more strongly than water does. The simulations are analyzed to determine the interactions that Asn is able to make in the binding pocket, and which sequence differences between AspRS and the highly homologous AsnRS are important for modifying the amino acid specificity. A double or triple mutation of AspRS that could make it specific for Asn is proposed, and supported by preliminary simulations of a mutant complex.

Amino Acid Sequence↗
PubMed1998

Titration calculations of foot-and-mouth disease virus capsids and their stabilities as a function of pH.

Foot-and-mouth disease virus (FMDV), a non-enveloped picornavirus, is sensitive to acidic conditions. At pH values below 7 the icosahedral virus capsid, formed from 60 copies of a protomer containing four polypeptides (VP1 to 4), dissociates into 12 pentamers, releasing the viral RNA. Evidence suggests that this acid lability may assist FMDV cell entry via an endosomal pathway. Calculations of titration curves and pH-stability profiles are presented for three different strains of FMDV, O1BFS, A10(61) and A22 Iraq, and compared with experimental data for complete virions and empty capsids (which lack RNA). The finite difference Poisson-Boltzmann method was used for the calculation of electrostatic free energies with the solvent treated as a dielectric continuum. The inter-pentamer interface in the virus is formed by two protomers related by 2-fold icosahedral symmetry. As a simple model for inter-pentamer interactions, a dimer and two separate protomers were compared. The association free energy was computed by integrating the difference between the titration curves of the two species. The calculations reproduced the observed decrease in capsid stability at acidic pH but not the difference in pH sensitivities of the two type A viruses. It is shown that only residues within 15 A of the interface play a significant role in determining acid lability. For the experimentally studied pH range (5 to 7.6), histidine residues were found to dominate the pH-dependence of the stability. Two histidine residues in VP3, H142 and H145, are shown to have the greatest effect by virtue of their interactions with many polar residues across the inter-pentamer interface; the interaction of H142 with an alpha-helix in the opposite pentamer contributes only a small proportion of the destabilization energy.

Aphthovirus↗
PubMed1998

Locally accessible conformations of proteins: multiple molecular dynamics simulations of crambin.

Multiple molecular dynamics (MD) simulations of crambin with different initial atomic velocities are used to sample conformations in the vicinity of the native structure. Individual trajectories of length up to 5 ns sample only a fraction of the conformational distribution generated by ten independent 120 ps trajectories at 300 K. The backbone atom conformational space distribution is analyzed using principal components analysis (PCA). Four different major conformational regions are found. In general, a trajectory samples only one region and few transitions between the regions are observed. Consequently, the averages of structural and dynamic properties over the ten trajectories differ significantly from those obtained from individual trajectories. The nature of the conformational sampling has important consequences for the utilization of MD simulations for a wide range of problems, such as comparisons with X-ray or NMR data. The overall average structure is significantly closer to the X-ray structure than any of the individual trajectory average structures. The high frequency (less than 10 ps) atomic fluctuations from the ten trajectories tend to be similar, but the lower frequency (100 ps) motions are different. To improve conformational sampling in molecular dynamics simulations of proteins, as in nucleic acids, multiple trajectories with different initial conditions should be used rather than a single long trajectory.

Computer Simulation↗
PubMed1998

Protein sidechain conformer prediction: a test of the energy function.

BACKGROUND: Homology modeling is an important technique for making use of the rapidly increasing number of protein sequences in the absence of structural information. The major problems in such modeling, once the alignment has been made, concern the positions of loops and the orientations of sidechains. Although progress has been made in recent years for sidechain prediction, current methods appear to have a limit on the order of 70% in their accuracy. It is important to have an understanding of this limitation, which for energy-based methods could arise from inaccuracies of the potential function. RESULTS: A test of the CHARMM function for sidechain prediction was performed. To eliminate the multiple-residue search problem, the minimum energy positions of individual sidechains in ten proteins were calculated in the presence of all other sidechains in their crystal orientations. This test provides a necessary condition that any energy function useful for sidechain placement must satisfy. For chi1 x chi2 rotations, the accuracies were 77.4% and 89.5%, respectively, and in the presence of crystal waters were 86.5% and 94.9%, respectively. If there was an error, the crystal structure usually corresponded to an alternative local minimum on the calculated energy map. Prediction accuracy correlated with the size of the energy gap between primary and secondary minima. CONCLUSIONS: The results indicate that the errors in current sidechain prediction schemes cannot be attributed to the potential energy function per se. The test used here establishes a necessary condition that any proposed energy-based sidechain prediction method, as well as many statistically based methods, must satisfy.

Crystallography, X-Ray↗
PubMed1998

A metastable state in folding simulations of a protein model.

The native state of a protein is generally believed to be the global free energy minimum. However, there is increasing evidence that kinetically selected states play a role in the biological function of some proteins. In a recent folding study of a 125-residue heteropolymer model, one of 200 sequences was found to fold repeatedly to a particular local minimum that did not interconvert to the global minimum. The kinetic preference for this 'metastable' state is shown to derive from an entropic barrier associated with inserting a tail segment into the protein interior of the serpin-like global minimum structure. The relation of the present results to the role of metastable states in functioning and pathogenic proteins is discussed.

Computer Simulation↗
PubMed1997

Folding thermodynamics of a model three-helix-bundle protein.

The calculated folding thermodynamics of a simple off-lattice three-helix-bundle protein model under equilibrium conditions shows the experimentally observed protein transitions: a collapse transition, a disordered-to-ordered globule transition, a globule to native-state transition, and the transition from the active native state to a frozen inactive state. The cooperativity and physical origin of the various transitions are explored with a single "optimization" parameter and characterized with the Lindemann criterion for liquid versus solid-state dynamics. Below the folding temperature, the model has a simple free energy surface with a single basin near the native state; the surface is similar to that calculated from a simulation of the same three-helix-bundle protein with an all-atom representation [Boczko, E. M. & Brooks III, C. L. (1995) Science 269, 393-396].

Animals↗
PubMed1997

Three-dimensional quantitative structure-activity relationships from molecular similarity matrices and genetic neural networks. 1. Method and validations.

The utility of genetic neural network (GNN) to obtain quantitative structure-activity relationships (QSAR) from molecular similarity matrices is described. In this application, the corticosteroid-binding globulin (CBG) binding affinity of the well-known steroid data set is examined. Excellent predictivity can be obtained through the use of either electrostatic or shape properties alone. Statistical validation using a standard randomization test indicates that the results are not due to chance correlations. Application of GNN on the combined electrostatic and shape matrix produces a six-descriptor model with a cross-validated r2 value of 0.94. The model is superior to those obtained from partial least-squares and genetic regressions, and it also compares favorably with the results for the same data set from other established 3D QSAR methods. The theoretical basis for the use of molecular similarity in QSAR is discussed.

Algorithms↗
PubMed1997

Three-dimensional quantitative structure-activity relationships from molecular similarity matrices and genetic neural networks. 2. Applications.

Validation of a method that uses a genetic neural network with electrostatic and steric similarity matrices (SM/GNN) to obtain quantitative structure-activity relationships (QSARs) is performed with eight data sets. Biological and physicochemical properties from a broad range of chemical classes are correlated and predicted using this technique. Quantitatively the results compare favorably with the benchmarks obtained by a number of well-established QSAR methods; qualitatively the models are consistent with the published descriptions on the relative contribution of steric and electrostatic factors. The results demonstrate the general utility of this method in deriving QSARs. The implication of the importance of molecular alignment and possible methodological improvements are discussed.

Cholinesterase Inhibitors↗
PubMed1997

"New view" of protein folding reconciled with the old through multiple unfolding simulations.

Twenty-four molecular dynamics trajectories of chymotrypsin inhibitor 2 provide a direct demonstration of the diversity of unfolding pathways. Comparison with experiments suggests that the transition state region for folding and unfolding occurs early with only 25 percent of the native contacts and that the root-mean-square deviations between contributing structures can be as large as 15 angstroms. Nevertheless, a statistically preferred unfolding pathway emerges from the simulations; disruption of tertiary interactions between the helix and a two-stranded portion of the beta sheet is the primary unfolding event. The results suggest a synthesis of the "new" and the classical view of protein folding with a preferred pathway on a funnel-like average energy surface.

Computer Simulation↗
PubMed1997

Ligand-induced conformational changes in ras p21: a normal mode and energy minimization analysis.

A normal mode and energy minimization of ras p21 is used to determine the flexibility of the protein and the origin of the conformational differences between GTP and GDP-bound forms. To preserve the integrity of the structures, a hydration shell of water molecules was included as part of the system. Certain low-frequency modes were found to have high involvement coefficients with the conformational transition between the GTP and GDP-bound structures; the involvement coefficients of some of the modes increase when the gamma-phosphate group is removed. Two unstable modes that appear in the GTP-bound structure upon deletion of the gamma-phosphate group were determined and shown to have dominant contributions in the regions of switch I and switch II; there was also a significant displacement of loop 1. The initial motion in these regions is predicted by the modes to be approximately perpendicular to the direction of the transition from the GTP-bound state to the GDP-bound state. The overall conformational change in the switch I and II regions involves rearrangements of the protein backbone within these regions, rather than rigid body motion. Differences in the low-frequency modes of the GTP and GDP-bound forms appear to play a role in ligand binding. A coupling between the helix alpha3 position and the deletion of the gamma-phosphate group may be involved in the interaction with GAP. The oncogenic mutation G12D leads to a global increase in the rigidity of the protein. Thus, the mutant is likely to have a higher barrier for the conformational change to the inactive form; this would slow the transition and could be related to its oncogenic properties.

Binding Sites↗
PubMed1997

Molecular switch in signal transduction: reaction paths of the conformational changes in ras p21.

Conformational changes in ras p21 triggered by the hydrolysis of GTP play an essential role in the signal transduction pathway. The path for the conformational change is determined by molecular dynamics simulation with a holonomic constraint directing the system from the known GTP-bound structure (with the gamma-phosphate removed) to the GDP-bound structure. The simulation is done with a shell of water molecules surrounding the protein. In the switch I region, the side chain of Tyr-32, which undergoes a large displacement, moves through the space between loop 2 and the rest of the protein, rather than on the outside of the protein. As a result, the charged residues Glu-31 and Asp-33, which interact with Raf in the homologous RafRBD-Raps complex, remain exposed during the transition. In the switch II region, the conformational changes of alpha2 and loop 4 are strongly coupled. A transient hydrogen bonding complex between Arg-68 and Tyr-71 in the switch II region and Glu-37 in switch I region stabilizes the intermediate conformation of alpha2 and facilitates the unwinding of a helical turn of alpha2 (residues 66-69), which in turn permits the larger scale motion of loop 4. Hydrogen bond exchange between the protein and solvent molecules is found to be important in the transition. Possible functional implications of the results are discussed.

Computer Simulation↗
PubMed1997

PDB-based protein loop prediction: parameters for selection and methods for optimization.

An approach to loop prediction that starts with a database search is presented and analyzed. To obtain meaningful statistics, 130 loops from 21 proteins were studied. The correlation between the internal conformation of the loop and the conformation of the neighboring stem residues was examined. Distances between C(alpha) and C(beta) of the immediate neighbor residues at each end select template loops as well as more complex (e.g. three residues on either side) matching criteria. To have a high probability that the best possible loop candidate in the database is included in the set, relatively large cutoffs for matching the interatomic distances of the stem residues have to be used in the template loop selection procedure; for loops of length 5, this results in an average of 1000 loops and for loops of length 9, the number is about 1500. The required number increases only slowly with loop length, in contrast to the exponential time increase involved in direct searches of the conformational space. The best loops among the large number of candidates can be determined by ranking them with the standard CHARMM non-bonded energy function (without electrostatics) applied to the backbone and C(beta) atoms. The same representation (backbone plus C(beta)) can be used to optimize the loop orientations relative to the rest of the protein by constrained energy minimization. Target loops that have many non-bonded contacts with the protein yield better results so that analysis of the non-bonded contacts of the selected template loops is useful in determining the expected accuracy of a prediction. The method for loop selection and optimization predicted eight (out of 18) loops of up to nine residues to an RMSD better than 1.07 A relative to the crystal structure; for 17 of the 18 loops, one of the three lowest energy template loops had an RMSD of less than 1.79 A. The prediction of antibody loops from a database search is more effective than that for non-antibody loops. Provided that they belong to one of the canonical classes, very similar antibody loops are certain to exist in the database. Superposition of the stem residues for antibody loops also results in a better orientation than with arbitrary target loops because the neighboring residues tend to have a more similar beta-strand structure. Two H3 loops (for which no canonical structures have been proposed) were predicted with reasonable accuracy (RMSD of 0.49 A and 1.07 A) even though no corresponding antibody loops were in the database.

Algorithms↗
PubMed1997

Use of the multiple copy simultaneous search (MCSS) method to design a new class of picornavirus capsid binding drugs.

A combinatorial ligand design approach based on the multiple copy simultaneous search (MCSS) method and a simple scheme for joining MCSS functional group sites was applied to the binding pocket of P3/Sabin poliovirus and rhinovirus 14. The MCSS method determines where specific functional (chemical) groups have local potential energy minima in the binding site. Before the virus application, test calculations were run to determine the optimal set of input parameters to be used in evaluating the MCSS results. The MCSS minima are analysed and selected minima are connected with (CH2)n linkers to form candidate ligands, whose structures are optimized in the binding site. Estimates of the binding strength were made for the ligands and compared with those for known drugs. The results indicate that the proposed ligands should bind to P3/Sabin poliovirus at least as well as the best of the existing drugs, and that they should also bind to P1/Mahoney poliovirus and rhinovirus 14. A detailed comparison of the poliovirus and rhinovirus binding pockets and an analysis of drug binding specificity is presented.

Algorithms↗
PubMed1997

Dynamics and unfolding pathways of a hyperthermophilic and a mesophilic rubredoxin.

Molecular dynamics simulations in solution are performed for a rubredoxin from the hyperthermophilic archaeon Pyrococcus furiosus (RdPf) and one from the mesophilic organism Desulfovibrio vulgaris (RdDv). The two proteins are simulated at four temperatures: 300 K, 373 K, 473 K (two sets), and 500 K; the various simulations extended from 200 ps to 1,020 ps. At room temperature, the two proteins are stable, remain close to the crystal structure, and exhibit similar dynamic behavior; the RMS residue fluctuations are slightly smaller in the hyperthermophilic protein. An analysis of the average energy contributions in the two proteins is made; the results suggest that the intraprotein energy stabilizes RdPf relative to RdDv. At 373 K, the mesophilic protein unfolds rapidly (it begins to unfold at 300 ps), whereas the hyperthermophilic does not unfold over the simulation of 600 ps. This is in accord with the expected stability of the two proteins. At 473 K, where both proteins are expected to be unstable, unfolding behavior is observed within 200 ps and the mesophilic protein unfolds faster than the hyperthermophilic one. At 500 K, both proteins unfold; the hyperthermophilic protein does so faster than the mesophilic protein. The unfolding behavior for the two proteins is found to be very similar. Although the exact order of events differs from one trajectory to another, both proteins unfold first by opening of the loop region to expose the hydrophobic core. This is followed by unzipping of the beta-sheet. The results obtained in the simulation are discussed in terms of the factors involved in flexibility and thermostability.

Amino Acid Sequence↗
PubMed1997

Computer simulations of the OmpF porin from the outer membrane of Escherichia coli.

Molecular dynamics simulations were used to study the structure and dynamics of the Escherichia coli OmpF porin, which is composed of three identical 16-stranded beta-barrels. Simulations of the full trimer in the absence of water and the membrane led to significant contraction of the channel in the interior of each beta-barrel. With very weak harmonic constraints (0.005 kcal/mol A2/atom) applied to the main-chain C alpha atoms of the beta-barrel, the structure was stabilized without alteration of the average fluctuations. The resulting distribution of the fluctuations (small for beta-strands, large for loops and turns) is in good agreement with the x-ray B factors. Dynamic cross-correlation functions showed the importance of coupling between the loop motions and barrel flexibility. This was confirmed by the application of constraints corresponding to the observed temperature factors to the barrel C alpha atoms. With these constraints, the beta-barrel fluctuations were much smaller than the experimental values because of the intrinsic restrictions on the atomic motions, and the loop motions were reduced significantly. This result indicates that considerable care is required in introducing constraints to keep proteins close to the experimental structure during simulations, as has been done in several recent studies. Loop 3, which is thought to be important in gating the pore, undergoes a displacement that shifts it away from the x-ray structure. Analysis shows that this arises from the breakdown of a hydrogen bond network, which appears to result more from the absence of solvent that from the use of standard ionization states for the side chains of certain beta-barrel residues.

Amino Acid Sequence↗
PubMed1997

The Levinthal paradox: yesterday and today.

A change in the perception of the protein folding problem has taken place recently. The nature of the change is outlined and the reasons for it are presented. An essential element is the recognition that a bias toward the native state over much of the effective energy surface may govern the folding process. This has replaced the random search paradigm of Levinthal and suggests that there are many ways of reaching the native state in a reasonable time so that a specific pathway does not have to be postulated. The change in perception is due primarily to the application of statistical mechanical models and lattice simulations to protein folding. Examples of lattice model results on protein folding are presented. It is pointed out that the new optimism about the protein folding problem must be complemented by more detailed studies to determine the structural and energetic factors that introduce the biases which make possible the folding of real proteins.

Models, Chemical↗