21 research outputs found
An in silico study of the molecular basis of B-RAF activation and conformational stability
<p>Abstract</p> <p>Background</p> <p>B-RAF kinase plays an important role both in tumour induction and maintenance in several cancers and it is an attractive new drug target. However, the structural basis of the B-RAF activation is still not well understood.</p> <p>Results</p> <p>In this study we suggest a novel molecular basis of B-RAF activation based on molecular dynamics (MD) simulations of B-RAF<sup>WT </sup>and the B-RAF<sup>V600E</sup>, B-RAF<sup>K601E </sup>and B-RAF<sup>D594V </sup>mutants. A strong hydrogen bond network was identified in B-RAF<sup>WT </sup>in which the interactions between Lys601 and the well known catalytic residues Lys483, Glu501 and Asp594 play an important role. It was found that several mutations, which directly or indirectly destabilized the interactions between these residues within this network, contributed to the changes in B-RAF activity.</p> <p>Conclusion</p> <p>Our results showed that the above mechanisms lead to the disruption of the electrostatic interactions between the A-loop and the αC-helix in the activating mutants, which presumably contribute to the flipping of the activation segment to an active form. Conversely, in the B-RAF<sup>D594V </sup>mutant that has impaired kinase activity, and in B-RAF<sup>WT </sup>these interactions were strong and stabilized the kinase inactive form.</p
PPARγ non-covalent antagonists exhibit mutable binding modes with a similar free energy of binding: a case study
<p>The structural and dynamical properties of PPARγ receptor in a complex with either partial or full agonists have been intensively studied but little is known about the receptor antagonistic conformation. A composition of microsecond accelerated molecular dynamics (aMD) simulation show that like partial agonists a non-covalent PPARγ full antagonist can bind in different modes of similar population size and free energies of binding. Four different and periodically exchanging ligand conformations are detected and described. The studied antagonist interacts with different receptor substructures and affects both the co-activator and the Cdk5 phosphorylation sites and, presumably, the natural complex with the DNA. However, no significant changes in the conformational states of the activation helix 12, and in particular an antagonist orientation, have been recorded. Finally, our results show also that the aMD approach can be successfully used in recovering the possible binding modes, considering fully the receptor flexibility, and is not dependent on the starting conformation.</p
An Improved Free Energy Perturbation FEP+ Sampling Protocol for Flexible Ligand-Binding Domains
Recent improvements to free energy perturbation (FEP) calculations, especiallyFEP+, established their utility for pharmaceutical lead optimization. However, to dateFEP has typically been helpful only when (1) high-quality X-ray data is available and(2) the target protein does not undergo significant conformational changes. Also, alack of systematic studies on determining an adequate sampling time is often one ofthe primary limitations of FEP calculations. Herein, we propose a modified versionof the FEP/REST (i.e., replica exchange with solute tempering) sampling protocol,based on systematic studies on several targets by probing a large number of permutations with different sampling schemes. Improved FEP+ binding affinity predictions for regular flexible-loop (F-loop) motions and considerable structural changes can be obtained by extending the pre-REST sampling time from 0.24 ns to 5 ns/λand 2×10 ns/λ, respectively. We obtained much more precise ∆∆G calculations of the individual perturbations, including the sign of the transformations and less error. We extended the REST simulations from 5 ns to 8 ns to achieve reasonable free energy convergence.Implementing REST to the entire ligand as opposed to solely the perturbed region, and also some important flexible protein residues (pREST region) in ligand binding domain (LBD) , also considerably improved the FEP+ results in most of the studied cases. Preliminary molecular dynamics (MD) runs were useful for establishing the correct binding mode of the compounds and thus precise alignment for FEP+.<br /
3D-QSAR and Molecular Mechanics Study for the Differences in the Azole Activity against Yeastlike and Filamentous Fungi and Their Relation to P450DM Inhibition. 1. 3-Substituted-4(3H)-quinazolinones
An Improved Free Energy Perturbation FEP+ Sampling Protocol for Flexible Ligand-Binding Domains
© 2019, The Author(s). Recent improvements to the free energy perturbation (FEP) calculations, especially FEP+ , established their utility for pharmaceutical lead optimization. Herein, we propose a modified version of the FEP/REST (i.e., replica exchange with solute tempering) sampling protocol, based on detail studies on several targets by probing a large number of perturbations with different sampling schemes. Improved FEP+ binding affinity predictions for regular flexible-loop motions and considerable structural changes can be obtained by extending the prior to REST (pre-REST) sampling time from 0.24 ns/λ to 5 ns/λ and 2 × 10 ns/λ, respectively. With this new protocol, much more precise ∆∆G values of the individual perturbations, including the sign of the transformations and decreased error were obtained. We extended the REST simulations from 5 ns to 8 ns to achieve reasonable free energy convergence. Implementing REST to the entire ligand as opposed to solely the perturbed region, and also some important flexible protein residues (pREST region) in the ligand binding domain (LBD) has considerably improved the FEP+ results in most of the studied cases. Preliminary molecular dynamics (MD) runs were useful for establishing the correct binding mode of the compounds and thus precise alignment for FEP+. Our improved protocol may further increase the FEP+ accuracy
An Improved Free Energy Perturbation FEP+ Sampling Protocol for Flexible Ligand-Binding Domains
Recent improvements to free energy perturbation (FEP) calculations, especiallyFEP+, established their utility for pharmaceutical lead optimization. However, to dateFEP has typically been helpful only when (1) high-quality X-ray data is available and(2) the target protein does not undergo significant conformational changes. Also, alack of systematic studies on determining an adequate sampling time is often one ofthe primary limitations of FEP calculations. Herein, we propose a modified versionof the FEP/REST (i.e., replica exchange with solute tempering) sampling protocol,based on systematic studies on several targets by probing a large number of permutations with different sampling schemes. Improved FEP+ binding affinity predictions for regular flexible-loop (F-loop) motions and considerable structural changes can be obtained by extending the pre-REST sampling time from 0.24 ns to 5 ns/λand 2×10 ns/λ, respectively. We obtained much more precise ∆∆G calculations of the individual perturbations, including the sign of the transformations and less error. We extended the REST simulations from 5 ns to 8 ns to achieve reasonable free energy convergence.Implementing REST to the entire ligand as opposed to solely the perturbed region, and also some important flexible protein residues (pREST region) in ligand binding domain (LBD) , also considerably improved the FEP+ results in most of the studied cases. Preliminary molecular dynamics (MD) runs were useful for establishing the correct binding mode of the compounds and thus precise alignment for FEP+.<br
Discovery of New AKT1 Inhibitors by Combination of In silico Structure Based Virtual Screening Approaches and Biological Evaluations
AKT1 is emerging as a useful target for treating cancer. Herein, we discovered a new set of ligands that inhibit the AKT1, as shown by in vitro binding and cell line studies, using a newly designed virtual screening protocol that combines structure-based pharmacophore and docking screens. Taking together with the biological data, the combination of structure based pharamcophore and docking methods demonstrated reasonable success rate in identifying new inhibitors (60-70%) proving the success of aforementioned approach. A detail analysis of the ligand-protein interactions was performed explaining observed activities.<br /
Prediction of Accurate Binding Modes Using Combination of Classical and Accelerated Molecular Dynamics and Free-Energy Perturbation Calculations: An Application to Toxicity Studies
Estimating the correct
binding modes of ligands in protein–ligand
complexes is crucial not only in the drug discovery process but also
for elucidating potential toxicity mechanisms. In the current paper,
we propose a computational modeling workflow using the combination
of docking, classical molecular dynamics (cMD), accelerated molecular
dynamics (aMD) and free-energy perturbation (FEP+ protocol) for identification
of possible ligand binding modes. It was applied for investigation
of selected perfluorocarboxyl acids (PFCAs) in the PPARγ nuclear
receptor. Although both regular and induced fit docking failed to
reproduce the experimentally determined binding mode of the ligands
when docked into a non-native X-ray structure, cMD and aMD simulations
successfully identified the most probable binding conformations. Moreover,
multiple binding modes were identified for all of these compounds
and the shorter-chain PFCAs continuously moved between a few energetically
favorable binding conformations. On the basis of MD predictions of
binding conformations, we applied the default and also redesigned
FEP+ sampling protocols, which accurately reproduced experimental
differences in the binding energies. Thus, the preliminary MD simulations
can also provide helpful information about correct setup of the FEP+
calculations. These results show that the PFCA binding modes were
accurately predicted and that the FEP+ protocol can be used to estimate
free energies of binding of flexible ligands that are not typical
druglike compounds. Our in silico workflow revealed the specific ligand–residue
interactions within the ligand binding domain and the main characteristics
of the PFCAs, and it was concluded that these compounds are week PPARγ
partial agonists. This work also suggests a common pipeline for identification
of ligand binding modes, ligand–protein dynamics description,
and relative free-energy calculations
Prediction of Accurate Binding Modes Using Combination of Classical and Accelerated Molecular Dynamics and Free-Energy Perturbation Calculations: An Application to Toxicity Studies
Estimating the correct
binding modes of ligands in protein–ligand
complexes is crucial not only in the drug discovery process but also
for elucidating potential toxicity mechanisms. In the current paper,
we propose a computational modeling workflow using the combination
of docking, classical molecular dynamics (cMD), accelerated molecular
dynamics (aMD) and free-energy perturbation (FEP+ protocol) for identification
of possible ligand binding modes. It was applied for investigation
of selected perfluorocarboxyl acids (PFCAs) in the PPARγ nuclear
receptor. Although both regular and induced fit docking failed to
reproduce the experimentally determined binding mode of the ligands
when docked into a non-native X-ray structure, cMD and aMD simulations
successfully identified the most probable binding conformations. Moreover,
multiple binding modes were identified for all of these compounds
and the shorter-chain PFCAs continuously moved between a few energetically
favorable binding conformations. On the basis of MD predictions of
binding conformations, we applied the default and also redesigned
FEP+ sampling protocols, which accurately reproduced experimental
differences in the binding energies. Thus, the preliminary MD simulations
can also provide helpful information about correct setup of the FEP+
calculations. These results show that the PFCA binding modes were
accurately predicted and that the FEP+ protocol can be used to estimate
free energies of binding of flexible ligands that are not typical
druglike compounds. Our in silico workflow revealed the specific ligand–residue
interactions within the ligand binding domain and the main characteristics
of the PFCAs, and it was concluded that these compounds are week PPARγ
partial agonists. This work also suggests a common pipeline for identification
of ligand binding modes, ligand–protein dynamics description,
and relative free-energy calculations
Combination of Genetic Screening and Molecular Dynamics as a Useful Tool for Identification of Disease-Related Mutations: ZASP PDZ Domain G54S Mutation Case
Cypher/ZASP
(<i>LDB3</i> gene) is known to interact with
a network of proteins. It binds to α-actinin and the calcium
voltage channels (LTCC) via its PDZ domain. Here we report the identification
of a highly conserved ZASP G54S mutation classified as a variant of
unknown significance in a sample of an adult with hypertrophic cardiomyopathy
(HCM). The initial bioinformatics calculations strongly evaluated
G54S as damaging. Furthermore, we employed accelerated and classical
molecular dynamics and free energy calculations to study the structural
impact of this mutation on the ZASP apo form and to address the question
of whether it can be linked to HCM. Seventeen independent MD runs
and simulations of 2.5 μs total were performed and showed that
G54S perturbs the α2 helix position via destabilization of the
adjacent loop linked to the β5 sheet. This also leads to the
formation of a strong H-bond between peptide target residues Leu17
and Gln66, thus restricting both the α-actinin2 and LTCC C-terminal
peptides to access their natural binding site and reducing in this
way their binding capacity. On the basis of these observations and
the adult’s clinical data, we propose that ZASP<sup>G54S</sup> and presumably other ZASP PDZ domain mutations can cause HCM. To
the best of our knowledge, this is the first reported ZASP PDZ domain
mutation that might be linked to HCM. The integrated workflow used
in this study can be applied for the identification and description
of other mutations that might be related to particular diseases