A salt-bridge switch in the molecular recognition between RS receptor and RGD ligand from the ABEEM σπ molecular dynamics simulations

Abstract

How the receptor and ligand recognise each other is a challenging subject in explaining the mechanism of recognition at the molecular level. As a starting point, here, a synthesised RS receptor and its RGD ligand were investigated as a proper model to simulate their recognition process in terms of ABEEMσπ/MM polarisable force field. It is found that a switch of forming up a salt bridge in the ligand triggers the recognition of the receptor and ligand. Through the salt-bridge switch that undergoes several cycles from on-state with parallel hydrogen bonds to off-state with bifurcated hydrogen bonds, the active site of ligand can flex easily to interact with the active site of the receptor. In addition, the water molecules form a decisive bridge connecting the active sites of the bound system. The salt-bridge switch and water-mediated movement are cooperative as the important factors for the receptor-ligand recognition. In addition, the properties, such as binding free energy, conformational flexibility and solvent accessible surface area have been calculated to provide adequate evidence for the whole recognition process. According to the simulation, a detailed mechanism was derived involving diffusion, a switch triggered cooperative water-mediated movement, and conformational folding, for the flexible recognition.     

Insights on the acetylated NF-κB transcription factor complex with DNA from molecular dynamics simulations

The nuclear factor-κB (NF-κB) is a DNA sequence-specific regulator of many important biological processes, whose activity is modulated by enzymatic acetylation. In one of the best functionally characterized NF-κB complexes, the p50/p65 heterodimer, acetylation of K221 at p65 causes a decrease of DNA dissociation rate, whilst the acetylation of K122 and K123, also at p65, markedly decreases the binding affinity for DNA. By means of molecular dynamics simulations based on the X-ray structure of the p50/p65 complex with DNA, we provide insights on the structural determinants of the acetylated complexes in aqueous solution. Lysine acetylation involves the loss of favorable electrostatic interactions between DNA and NF-κB, which is partially compensated by the reduction of the desolvation free-energy of the two binding partners. Acetylation at both positions K122 and K123 is associated with a decrease of the electrostatic potential at the p65/DNA interface, which is only partially counterbalanced by an increase of the local Na(+) concentration. It induces the disruption of base-specific and nonspecific interactions between DNA and NF-κB and it is consistent with the observed decrease of binding affinity. In contrast, acetylation at position K221 results in the loss of nonspecific protein-DNA interactions, but the DNA recognition sites are not affected. In addition, the loss of protein-DNA interactions is likely to be counterbalanced by an increase of the configurational entropy of the complex, which provides, at a speculative level, a justification for the observed decrease of NF-κB/DNA dissociation rate.     

Structure and dynamics of two β-peptides in solution from molecular dynamics simulations validated against experiment

We have studied two different beta-peptides in methanol using explicit solvent molecular dynamics simulations and the GROMOS 53A6 force field: a heptapeptide (peptide 1) expected to form a left-handed 3(14)-helix, and a hexapeptide (peptide 2) expected to form a beta-hairpin in solution. Our analysis has focused on identifying and analyzing the stability of the dominant secondary structure conformations adopted by the peptides, as well as on comparing the experimental NOE distance upper bounds and 3J-coupling values with their counterparts calculated on the basis of the simulated ensembles. Moreover, we have critically compared the present results with the analogous results obtained with the GROMOS 45A3 (peptide 1) and 43A1 (peptide 2) force fields. We conclude that within the limits of conformational sampling employed here, the GROMOS 53A6 force field satisfactorily reproduces experimental findings regarding the behavior of short beta-peptides, with accuracy that is comparable to but not exceeding that of the previous versions of the force field. GCE legend Conformational clustering analysis of the simulated ensemble of a ss-hexapeptide with two different simulation setups (a and b). The central members of all of the clusters populating more than 5% of all of the structures are shown, together with the most dominant hydrogen bonds and the corresponding percentages of cluster members containing them.     

Study of the inhibition of cyclin-dependent kinases with roscovitine and indirubin-3′-oxime from molecular dynamics simulations

Molecular dynamics simulations were performed to elucidate the interactions of CDK2 and CDK5 complexes with three inhibitors: R-roscovitine, S-roscovitine, and indirubin-3′-oxime. The preference of the two complexes for R-roscovitine over the S enantiomer, as reported by the experiment, was also found by the simulations. More importantly, the simulations showed that the cause of the stronger affinity for the R enantiomer is the presence of an important hydrogen bond between R-roscovitine and the kinases not found with S-roscovitine. The simulations also showed two amino acid mutations in the active site of CDK5/R-roscovitine that favor binding-enhanced electrostatic contributions, making the inhibitor more effective for CDK5 than for CDK2. This suggests that the effectiveness of roscovitine-like inhibitors can be improved by enhancing their electrostatic interaction with the kinases. Finally, molecular mechanics–Possion–Boltzmann/surface area calculations of the CDK5/indirubin-3′-oxime system in both water-excluded and water-included environments gave significantly different electrostatic contributions to the binding. The simulations detected the displacement of a water molecule in the active site of the water-included CDK/indirubin-3′-oxime system. This resulted in a more conserved binding pattern than the water-excluded structure. Hence, in the design of new indirubin-like inhibitors, it is important to include the water molecule in the analysis. Figure Hydrogen bonding networks at the active sites of both CDK5/R-roscotivine (light grey) and CDK2/R-roscovitine (black).     

Influence of the SNPs on the structural stability of CBS protein： Insight from molecular dynamics simulations

Cystathionine β-synthase is an essential enzyme of the trans-suifuration pathway that condenses serine with homocysteine to form cystathionine. Missense mutations in CBS are the major cause of inherited homocystinuria, and the detailed effect of disease associated amino acid substitutions on the structure and stability of human CBS is yet unknown. Here, we apply a unique approach in combining in silico tools and molecular dynamics simulation to provide structural and functional insight into the effect of SNP on the stability and activity of mutant CBS. In addition, principal component analysis and free energy landscape were used to predict the collective motions, thermodynamic stabilities and essential subspace relevant to CBS function. The obtained results indicate that C109R, E176K and D376N mutations have the diverse effect on dynamic behavior of CBS protein. We found that highly conserved D376N mutation, which is present in the active pocket, affects the protein folding mechanism. Our strategy may provide a way in near future to understand and study effects of functional nsSNPs and their role in causing homocystinuria.     

Crankshaft motions of the polypeptide backbone in molecular dynamics simulations of human type-α transforming growth factor

Summary Order parameters for the backbone N–H and C–H bond vectors have been calculated from a 150 ps molecular dynamics (MD) simulation of human type- transforming growth factor in H₂O solvent. Two kinds of crankshaft motions of the polypeptide backbone are observed in this MD trajectory. The first involves small-amplitude rocking of the rigid peptide bond due to correlated changes in the backbone dihedral angles _i–1 and _i. These high-frequency librational crankshaft motions are correlated with systematically smaller values of motional order parameters for backbone N–H bond vectors compared to C–H bond vectors. In addition, infrequent crankshaft flips of the peptide bond from one local minimum to another are observed for several amino acid residues. These MD simulations demonstrate that comparisons of N–H and C–H order parameters provide a useful approach for identifying crank-shaft librational motions in proteins.     

Characterization of the internal dynamics and conformational space of zinc-bound amyloid β peptides by replica-exchange molecular dynamics simulations

Amyloid β (Aβ) peptides and metal ions have been associated with the pathogenesis of Alzheimer’s disease. The conformational space of Aβ fragments of different length with and without binding of metal ions has been extensively investigated by replica-exchange molecular dynamics (REMD) simulation. However, only trajectories extracted at relatively low temperatures have been used for this analysis. The capability of REMD simulations to characterize the internal dynamics of such intrinsically disordered proteins (IDPs) as Aβ has been overlooked. In this work, we use an approach recently developed by Xue and Skrynnikov (J Am Chem Soc 133:14614–14628, 2011) to calculate NMR observables, including ¹⁵N relaxation rates and ¹⁵N–¹H nuclear Overhauser enhancement (NOE), from the high-temperature trajectory of REMD simulations for zinc-bound Aβ peptides. The time axis of the trajectory was rescaled to correct for the effect of the high temperature (408 K) compared with the experimental temperature (278 K). Near-quantitative agreement between simulated values and experimental results was obtained. When the structural properties and free-energy surfaces of zinc-bound Aβ_(1–40) and Aβ_(1–42) were compared at the physiological temperature 310 K it was found that zinc-bound Aβ_(1–42) was more rigid than Aβ_(1–40) at the C terminus, and its conformational transitions were also more preferred. The self-consistent results derived from trajectories at high and low temperatures demonstrate the capability of REMD simulations to capture the internal dynamics of IDPs.     

Do new reconstructions clarify the relationships between the Phanerozoic diversity dynamics of marine invertebrates and long-term eustatic trends?

A relationship between global sea levels and the diversity of marine invertebrates throughout the Phanerozoic remains an urgent matter for debates. Its recognition depends on a proper selection of diversity and eustatic curves. A comparison of changes in the revised sample-standardized generic diversity and long-term global sea-level changes provides a weaker evidence for a direct covarying relationship than established earlier, although the eustatic control on diversity dynamics of marine invertebrates was important during ∼74% of the Phanerozoic. Multiple causation of biodiversity changes, data bias, erroneous reconstructions, and conceptual misinterpretations are likely explanations of observed difference between the new biodiversity and eustatic curves.     

Unfolding of the amyloid β-peptide central helix: mechanistic insights from molecular dynamics simulations

Alzheimer's disease (AD) pathogenesis is associated with formation of amyloid fibrils caused by polymerization of the amyloid β-peptide (Aβ), which is a process that requires unfolding of the native helical structure of Aβ. According to recent experimental studies, stabilization of the Aβ central helix is effective in preventing Aβ polymerization into toxic assemblies. To uncover the fundamental mechanism of unfolding of the Aβ central helix, we performed molecular dynamics simulations for wild-type (WT), V18A/F19A/F20A mutant (MA), and V18L/F19L/F20L mutant (ML) models of the Aβ central helix. It was quantitatively demonstrated that the stability of the α-helical conformation of both MA and ML is higher than that of WT, indicating that the α-helical propensity of the three nonpolar residues (18, 19, and 20) is the main factor for the stability of the whole Aβ central helix and that their hydrophobicity plays a secondary role. WT was found to completely unfold by a three-step mechanism: 1) loss of α-helical backbone hydrogen bonds, 2) strong interactions between nonpolar sidechains, and 3) strong interactions between polar sidechains. WT did not completely unfold in cases when any of the three steps was omitted. MA and ML did not completely unfold mainly due to the lack of the first step. This suggests that disturbances in any of the three steps would be effective in inhibiting the unfolding of the Aβ central helix. Our findings would pave the way for design of new drugs to prevent or retard AD.     

Structural insights into the two sequential folding transition states of the PB1 domain of NBR1 from Φ value analysis and biased molecular dynamics simulations

The PB1 domain of NBR1 folds via a single pathway mechanism involving two sequential energy barriers separated by a high-energy intermediate. The structural ensemble representing each of the two transition states (TS1 and TS2) has been calculated using experimental Φ values and biased molecular dynamics simulations. Both TS1 and TS2 represent compact states (β(TS1) = 0.71, and β(TS2) = 0.93) but are defined by quite different distributions of Φ values, degrees of structural heterogeneity, and nativelike secondary structure. TS1 forms a heterogeneous ensemble of dynamic structures, representing a global collapse of the polypeptide chain around a set of weak nativelike contacts. In contrast, TS2 has a high proportion of nativelike secondary structure, which is reflected in an extensive distribution of high Φ values. Two snapshots along the folding pathway of the PB1 domain reveal insights into the malleability, the solvent accessibility, and the timing of nativelike core packing that stabilizes the folded state.     

Ligand-escape pathways from the ligand-binding domain of PPARγ receptor as probed by molecular dynamics simulations

Conformational rearrangements of peroxysome proliferator activated receptor (PPARγ) ligand-binding domain (LBD) that accompany the release and binding of ligands are not well understood. To determine the major events associated with the escape of the partial agonist GW0072, molecular dynamic (MD) simulations were performed using two different methods: reversed targeted molecular dynamics (TMD⁻¹) and time-dependent distance restraints (TDR) using as restraints either the root mean square deviation from a reference structure (TMD⁻¹) or the distance between the geometrical centers of the binding pocket and of the ligand (TDR). Both methods do not assume any a priori route for ligand extraction. To avoid artifacts, different initial simulation conditions were used and particular attention was paid for giving time to the protein to relax during the extraction process by running 10–12 ns simulations within explicit water. Two distinct exit gates A and B were found, independently of initial conditions and method. During the exit process no interaction between GW0072 and the transactivation AF-2 helix was observed. Our results suggest that the ligand uses the intrinsic flexibility of the protein to move within the receptor. Paths A and B are very similar to those found for other nuclear receptors, suggesting that these routes are a common characteristics of nuclear receptors that are used by different kinds of ligands. Finally, the knowledge of entry/exit pathways of a receptor should be very useful in discriminating between different ligands that could have been favorably docked in the binding pocket by introducing docking along these pathways into computational drug design protocols.     

Exploration of the activation pathway of Δα-Chymotrypsin with molecular dynamics simulations and correlation with kinetic experiments

Correlating the experimentally observed kinetics of protein conformational changes with theoretical predictions is a formidable and challenging task, due to the multitude of degrees of freedom (>5,000) in a protein and the huge gap between the timescale of the kinetic event of interest (ms) and the typical timescale of computer simulations (ns). In this study we show that using the targeted molecular dynamics (TMD) method it is possible to simulate conformational changes of the ms time range and to correlate multiple simulations of single pathways with ensemble experiments on both the structural and energetic basis. As a model system we chose to study the conformational change of rat-Δα-chymotrypsin from its inactive to its active conformation. This activation process has been analyzed previously by experimental and theoretical methods, i.e. fluorescence stopped-flow spectroscopy (FSF), molecular dynamics (MD) and TMD. Inspired by the results of these studies on the wild type (WT) enzyme, several mutants were constructed to alter the conformational pathway and studied by FSF measurements. In the present work WT and mutant N18G were subjected to multiple MD and subsequent TMD simulations. We report the existence of two main activation pathways, a feature of chymotrypsin activation that has been hitherto unknown. A method to correlate the energetics of the different pathways calculated by TMD and the kinetic parameters observed by experimental methods such as FSF is presented. Our work is relevant for experimental single molecule studies of enzymes in general. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

A molecular modelling study of the interaction between β-cyclodextrin and the organophosphorothioate pesticide parathion

The interaction between parathion and -cyclodextrin was investigated by Molecular Dynamics. Several in vacuo trajectories were calculated for the system imposing a 1:1 stoichiometry. The influence of the solvent and temperature was considered. The results account for the formation of adducts which are stable at room temperature and involve mainly the nitrophenoxy group of the guest molecules which interacts with the hydrophobic cavity of the host by van der Waals forces.     

Tracking down the molecular substrates of stress: new roles for p38α MAPK and kappa-opioid receptors

In this issue, Bruchas et?al. (2011) uncover a novel stress-induced p38α MAPK signaling cascade within serotonergic neurons of the dorsal raphe nucleus that mediates depressive and drug-seeking behaviors. Their findings have potentially important implications for medication development.     

Partial activation of α7 nicotinic acetylcholine receptors: insights from molecular dynamics simulations

Nicotinic acetylcholine receptors (nAChRs) are drug targets for neuronal disorders and diseases. Partial agonists for nAChRs are currently being developed as drugs for the treatment of neurological diseases for their relative safety originated from reduced excessive stimulation. In the current study, molecular docking, molecular dynamics simulations and binding energy calculations were performed to theoretically investigate the interactions between the partial agonists, 4-OH-DMXBA and tropisetron with α7-nAChR. The results suggest that the partial agonists 4-OH-DMXBA and tropisetron bind with α7-nAChR in a binding mode similar to that with AChBP. The non-conserved residues in the binding sites contribute to the orientation deviation of these partial agonists from their orientation in AChBP. Energy calculation and decomposition using MM-GB/SA suggests that the van der Waals term (ΔE_VDW) is the main driving force for the binding of the partial agonists to α7-nAChR. The molecular dynamics simulations showed that the opening of the C-loop binding with the partial agonists is in-between the openings for the binding with the full agonist and in the apo state. This conformation difference for the C-loop sheds light on the partial agonism of nAChR.     

Can molecular dynamics simulations improve the structural accuracy and virtual screening performance of GPCR models?

The determination of G protein-coupled receptor (GPCR) structures at atomic resolution has improved understanding of cellular signaling and will accelerate the development of new drug candidates. However, experimental structures still remain unavailable for a majority of the GPCR family. GPCR structures and their interactions with ligands can also be modelled computationally, but such predictions have limited accuracy. In this work, we explored if molecular dynamics (MD) simulations could be used to refine the accuracy of in silico models of receptor-ligand complexes that were submitted to a community-wide assessment of GPCR structure prediction (GPCR Dock). Two simulation protocols were used to refine 30 models of the D₃ dopamine receptor (D₃R) in complex with an antagonist. Close to 60 μs of simulation time was generated and the resulting MD refined models were compared to a D₃R crystal structure. In the MD simulations, the receptor models generally drifted further away from the crystal structure conformation. However, MD refinement was able to improve the accuracy of the ligand binding mode. The best refinement protocol improved agreement with the experimentally observed ligand binding mode for a majority of the models. Receptor structures with improved virtual screening performance, which was assessed by molecular docking of ligands and decoys, could also be identified among the MD refined models. Application of weak restraints to the transmembrane helixes in the MD simulations further improved predictions of the ligand binding mode and second extracellular loop. These results provide guidelines for application of MD refinement in prediction of GPCR-ligand complexes and directions for further method development.     

Effects of ligands on unfolding of the amyloid β-peptide central helix: mechanistic insights from molecular dynamics simulations

Polymerization of the amyloid β-peptide (Aβ), a process which requires that the helical structure of Aβ unfolds beforehand, is suspected to cause neurodegeneration in Alzheimer's disease. According to recent experimental studies, stabilization of the Aβ central helix counteracts Aβ polymerization into toxic assemblies. The effects of two ligands (Dec-DETA and Pep1b), which were designed to bind to and stabilize the Aβ central helix, on unfolding of the Aβ central helix were investigated by molecular dynamics simulations. It was quantitatively demonstrated that the stability of the Aβ central helix is increased by both ligands, and more effectively by Pep1b than by Dec-DETA. In addition, it was shown that Dec-DETA forms parallel conformations with β-strand-like Aβ, whereas Pep1b does not and instead tends to bend unwound Aβ. The molecular dynamics results correlate well with previous experiments for these ligands, which suggest that the simulation method should be useful in predicting the effectiveness of novel ligands in stabilizing the Aβ central helix. Detailed Aβ structural changes upon loss of helicity in the presence of the ligands are also revealed, which gives further insight into which ligand may lead to which path subsequent to unwinding of the Aβ central helix.