Atomistic MD Simulations Research Articles

Analysis of the Atomistic MD Simulations of CI+CIII2 Interactions and Associated Lipid Binding

Analysis of the Atomistic MD Simulations of CI+CIII2 Interactions and Associated Lipid Binding

Molecular dynamics simulations in pre-polymerization mixtures for peptide recognition.

Molecularly imprinted polymers (MIPs) have promising applications as synthetic antibodies for protein and peptide recognition. A critical aspect of MIP design is the selection of functional monomers and their adequate proportions to achieve materials with high recognition capacity toward their targets. To contribute to this goal, we calibrated a molecular dynamics protocol to reproduce the experimental trends in peptide recognition of 13 pre-polymerization mixtures reported in the literature for the peptide toxin melittin. Three simulation conditions were tested for each mixture by changing the box size and the number of monomers and cross-linkers surrounding the template in a solvent-explicit environment. Fully atomistic MD simulations of 350 ns were conducted with the AMBER20 software, with ff19SB parameters for the peptide, gaff2 parameters for the monomers and cross-linkers, and the OPC water model. Template-monomer interaction energies under the LIE approach showed significant differences between high-affinity and low-affinity mixtures. Simulation systems containing 100 monomers plus cross-linkers in a cubic box of 90 Å3 successfully ranked the mixtures according to their experimental performance. Systems with higher monomer densities resulted in non-specific intermolecular contacts that could not account for the experimental trends in melittin recognition. The mixture with the best recognition capacity showed preferential binding to the 13-26-α-helix, suggesting a relevant role for this segment in melittin imprinting and recognition. Our findings provide insightful information to assist the computational design of molecularly imprinted materials with a validated protocol that can be easily extended to other templates.

CGeNArate: a sequence-dependent coarse-grained model of DNA for accurate atomistic MD simulations of kb-long duplexes.

We present CGeNArate, a new model for molecular dynamics simulations of very long segments of B-DNA in the context of biotechnological or chromatin studies. The developed method uses a coarse-grained Hamiltonian with trajectories that are back-mapped to the atomistic resolution level with extreme accuracy by means of Machine Learning Approaches. The method is sequence-dependent and reproduces very well not only local, but also global physical properties of DNA. The efficiency of the method allows us to recover with a reduced computational effort high-quality atomic-resolution ensembles of segments containing many kilobases of DNA, entering into the gene range or even the entire DNA of certain cellular organelles.

Comparative analysis of structural dynamics and allosteric mechanisms of RecA/Rad51 family proteins: Integrated atomistic MD simulation and network-based analysis

Homologous recombination plays a key role in double-strand break repair, stalled replication fork repair, and meiosis. The RecA/Rad51 family recombinases catalyze the DNA strand invasion reaction that occurs during homologous recombination. However, the high sequence differences between homologous groups have hindered the thoroughly studies of this ancient protein family. The dynamic mechanisms of the family, particularly at the residual level, remain poorly understood. In this work, five representative RecA/Rad51 recombinase family members from all major kingdoms of living organisms: prokaryotes, eukaryotes, archaea, and viruses, were selected to explore the molecular mechanisms behind their conserved biological significance. A variety of techniques, including all-atom molecular dynamics simulation, perturbation response scanning, and protein structure network analysis, were used to examine the flexibility and correlation of protein domains, distribution of sensors and effectors and conserved hub residues. Furthermore, the potential communication routes between the ATP-binding region and the DNA-binding region of each recombinase were identified. Our results demonstrate the conserved molecular dynamics of these recombinases in the early stage of homologous recombination, including cooperative motions between regions, conserved sensing and effecting functional residue distribution, and conserved hub residues. Meanwhile, the unique ATP-DNA communication routes of each recombinase was also revealed. These results provide new insights into the mechanism of RecA/Rad51 family proteins, and provide new theoretical guidance for the development of allosteric inhibitors and the application of RecA/Rad51 family proteins.

Origin of the central pit in hemimicelles of semifluorinated alkanes: How molecular dipoles and substrate deformation can determine supra-molecular morphology

HypothesisSemifluorinated alkanes amphiphiles spontaneously form highly monodispersed hemimicelles at the surface of water. The origin of the formation and complex structure of these surprising supramolecular aggregates were only recently clarified using molecular dynamics simulations (MD). The existence of a pit at the center of these aggregates made up of almost 3000 molecules was indeed reproduced by the MD simulations, but not understood. MethodA careful strategy of atomistic MD simulations comparing non-electrostatic force fields with force fields that include electrostatic forces, thus bearing an implicit or explicit dipole, allowed demonstrating the roles of dipolar interactions and interactions with the liquid subphase on the morphology of the aggregates. FindingsThe simulation results clearly show that within the hemimicelles the strong molecular dipoles located at the CH2-CF2 junctions tend to align, leading to a collective shift of the PFAA molecules relatively to each other. This shift is responsible for the curvature of the hemimicelles and originates the central pit, provided the possibility of deforming the surface of the water sub-phase. Comparisons with non-electrostatic force field results further contribute to understand the origin of the self-assembling process. The results directly connect for the first time a molecular property with a mesoscopic structural feature. Given the molecular simplicity of these “primitive” amphiphiles compared to the common hydrophilic/hydrophobic surfactants, the results contribute to understand self-assembly in general.

Investigating the mechanical performance of graphene reinforced polymer nanocomposites via atomistic and continuum simulation approaches

Graphene, having exceptional mechanical properties, is expected to improve the overall mechanical performance of polymer nanocomposites when inserted as a nanofiller. In this work, the mechanical properties of graphene-based cis-1,4-polybutadiene glassy polymer nanocomposites are investigated via a hierarchical multiscale approach, combining atomistic simulations and continuum mechanics. The global mechanical properties of the graphene-based polymer nanocomposites are computed via a rich enough set of remote applied deformations. On a localized level, the effective Young's modulus and Poisson's ratio are calculated in both the interphase and polymer matrix regions where the former shows greater rigidity. Finally, the properties of the different phases forming the heterogeneous polymer nanocomposite are used to develop a continuum three-phase micromechanical finite element model for predicting the overall mechanical behavior of the structure, which are found to be in very good agreement with the results from the atomistic MD simulations.

Heterogeneous Glass Transition Behavior of Poly(Ethylene oxide)/Silica Nanocomposites via Atomistic MD Simulations

Heterogeneous Glass Transition Behavior of Poly(Ethylene oxide)/Silica Nanocomposites via Atomistic MD Simulations

Autophagosome membrane expansion is mediated by the N-terminus and cis-membrane association of human ATG8s.

Autophagy is an essential catabolic pathway which sequesters and engulfs cytosolic substrates via autophagosomes, unique double-membraned structures. ATG8 proteins are ubiquitin-like proteins recruited to autophagosome membranes by lipidation at the C-terminus. ATG8s recruit substrates, such as p62, and play an important role in mediating autophagosome membrane expansion. However, the precise function of lipidated ATG8 in expansion remains obscure. Using a real-time in vitro lipidation assay, we revealed that the N-termini of lipidated human ATG8s (LC3B and GABARAP) are highly dynamic and interact with the membrane. Moreover, atomistic MD simulation and FRET assays indicate that N-termini of LC3B and GABARAP associate in cis on the membrane. By using non-tagged GABARAPs, we show that GABARAP N-terminus and its cis-membrane insertion are crucial to regulate the size of autophagosomes in cells irrespectively of p62 degradation. Our study provides fundamental molecular insights into autophagosome membrane expansion, revealing the critical and unique function of lipidated ATG8.

Molecular Biophysics of Class A G Protein Coupled Receptors-Lipids Interactome at a Glance-Highlights from the A2A Adenosine Receptor.

G protein-coupled receptors (GPCRs) are embedded in phospholipid membrane bilayers with cholesterol representing 34% of the total lipid content in mammalian plasma membranes. Membrane lipids interact with GPCRs structures and modulate their function and drug-stimulated signaling through conformational selection. It has been shown that anionic phospholipids form strong interactions between positively charged residues in the G protein and the TM5-TM6-TM 7 cytoplasmic interface of class A GPCRs stabilizing the signaling GPCR-G complex. Cholesterol with a high content in plasma membranes can be identified in more specific sites in the transmembrane region of GPCRs, such as the Cholesterol Consensus Motif (CCM) and Cholesterol Recognition Amino Acid Consensus (CRAC) motifs and other receptor dependent and receptor state dependent sites. Experimental biophysical methods, atomistic (AA) MD simulations and coarse-grained (CG) molecular dynamics simulations have been applied to investigate these interactions. We emphasized here the impact of phosphatidyl inositol-4,5-bisphosphate (PtdIns(4,5)P2 or PIP2), a minor phospholipid component and of cholesterol on the function-related conformational equilibria of the human A2A adenosine receptor (A2AR), a representative receptor in class A GPCR. Several GPCRs of class A interacted with PIP2 and cholesterol and in many cases the mechanism of the modulation of their function remains unknown. This review provides a helpful comprehensive overview for biophysics that enter the field of GPCRs-lipid systems.

Inferring the transport stoichiometry of the proton/zinc transporter YiiP from microscale thermophoresis and simulations.

The transport stoichiometry of the Zn2+/H+ antiporter YiiP remains an open question. We used experimental microscale thermophoresis (MST) zinc binding data in combination with a microscopic thermodynamic model and constant pH simulation in order to infer the likely microstates of H+ and Zn2+ binding. The pH dependence of Zn2+ binding in different binding sites was measured with MST. For these sites, we constructed microscopic, thermodynamically consistent models based on the pKas of the binding residues and zinc binding affinities. Such a model relates the macroscopically measured MST binding curves to microscopic protonation states and zinc binding in a concentration and pH-dependent manner. We inferred microscopic model parameters from the experimental data with a Monte Carlo approach. We compared the MST-inferred microscopic parameters to results from atomistic constant pH MD simulations in the absence of Zn2+. For zinc binding site B on a solvent-exposed loop between transmembrane helices 2 and 3, MST and simulation data broadly agree with each other. For the transport site A in the transmembrane domain, the CpHMD simulations indicate that the binding site His residue is protonated before any Asp. Using the CpHMD results as a starting point for the MST inference approach, we find that the microstate distribution that best fits the experimental data indicates competitive binding between either one Zn2+ ion or two or three protons bound, with the one-proton state suppressed, possibly due to high cooperativity between the Asp and His residues. The microscopic model indicates a transport stoichiometry of 1:2, or possibly 1:3 at lower pH.

An integrative and multiscale approach to study dengue virus-like particle dynamics.

Dengue virus is an enveloped virus which infects millions of people worldwide. The only available vaccine has low efficacy and is recommended only for individuals with prior dengue infection. Virus-like particles (VLPs) are promising platforms as vaccine candidates due to their non-infectious and highly immunogenic nature. These particles lack a genomic core and are highly unstable and heterogeneous in size. This motivates the characterization of the biophysical and structural properties of VLPs to facilitate the rational production of stable and homogenous particles. Dengue VLPs elicits neutralizing antibodies similar to infectious virions. Dengue VLPs are enveloped particles in which sixty envelope (E) and membrane (M) proteins are embedded in a lipid vesicle. Similarly, to the dengue virus, the VLPs undergo a maturation process, involving large conformational changes, leading to exposure of epitopes that subsequently determine the immune response. In this work, the structure and intrinsic dynamics of VLPs and their conformational transitions were investigated using integrative modeling and molecular simulations. To gain atomistic insights into the maturation process and to facilitate the development of stable enveloped VLPs, we employed an integrative approach combining multiscale modeling and simulations with experimental data from cryo-electron microscopy, lipidomics, and secretion assays. Coarse-grained simulations of mature and immature VLPs revealed the effective number of lipids that result in optimally stable and spherical particles. Furthermore, these models were used as a platform to simulate the transition between conformational states in order to understand the maturation process in unprecedented detail. Finally, atomistic MD simulations performed on native dengue and dengue-Japanese encephalitis virus chimeric E/M proteins and their mutants highlighted the role of protein-lipid interactions in governing VLP stability. This integrative approach aims towards the future development of stable and immunogenic next-generation vaccines.

The single residue K12 governs highly sensitive voltage gating of the voltage-dependent anion channel.

The voltage-dependent anion channel (VDAC) has emerged as a key player in mitochondrial bioenergetics. As the main pathway for calcium and mitochondrial metabolites, such as ATP and ADP, to cross the mitochondrial outer membrane, VDAC is central to maintenance of mitochondrial homeostasis. It is thus unsurprising that VDAC is implicated in nearly every mitochondria-related pathology, including cardiovascular disease, cancers, and neurodegeneration. Despite decades of research, the relatively uncomplicated structure of VDAC—a β-barrel with an internal broken α-helical belt—has hampered efforts to elucidate its functional mechanisms. Unlike other β-barrel channels, such as bacterial porins and toxins, VDAC transitions to low-conducting states, or “gates”, at relatively low voltages, less than 30 mV. In VDAC's gated state, metabolite transport is suppressed and calcium transport enhanced; thus, gating is one means by which VDAC may regulate fluxes across the mitochondrial membrane. In this talk, I will describe computational and experimental efforts that have led to identification of K12 as a key residue that is responsible for the exquisite sensitivity to voltage of its gating process [JACS 2022, 144(32) 14564-14577, DOI:10.1021/jacs.2c03316]. K12 is bistable; its motions between two widely separated positions along the pore axis enhance the fluctuations of the β-barrel and augment the likelihood of gating. These results are confirmed by single channel electrophysiology of various K12 mutants, which show a dramatic reduction of the frequency of voltage-induced gating transitions. The crystallographic structure of the K12E mutant is similar to the wild type; however, 60 µs of atomistic MD simulations using the K12E mutant structure show restricted motion of the mutated residue 12, due to enhanced connectivity with neighboring residues, and diminished amplitude of barrel motions. We conclude that β-barrel fluctuations, governed particularly by residue K12, drive VDAC gating transitions.

Towards a multiscale model of P-glycoprotein efflux.

The blood-brain barrier (BBB) remains a major roadblock to the delivery of drugs to the brain. While in vitro and in vivo measurements of permeability are widely used to predict brain penetration, very little is known about the mechanisms of passive and active transport. Detailed insight into interactions between solutes and cell membranes could provide new insight into drug design and screening. Multidrug resistance (MDR) is a problem that affects treatment of many diseases, and is often associated with P-glycoprotein (P-gp, ABCB1 or MDR1), one of the ABC family of transporters. These efflux pumps transport substrates that have crossed the blood-brain barrier back out into circulation, thereby limiting bioavailability in the local tissue. The mechanism of how P-gp is inhibited is not well understood, in particular how the ligand reaches its binding site. Here, we build a multiscale model of transport across the BBB. Firstly, we demonstrate our BBB membrane model of the brain microvascular endothelial cell (hBMEC) membrane. We perform simulate unbiased atomistic MD simulations to visualize translocation of a library of 23 solutes across a lipid bilayer representative of brain microvascular endothelial cells (hBMEC). Secondly, we perform simulations of the active transport of P-glycoprotein (P-gp) efflux out of the cell for a substrate and a potential third-generation inhibitor. Then, we demonstrate results using a novel coarse-grain model of P-gp with MARTINI 3.0, which enables us to access much longer timescales.

Effect of Nanoconfinement on NMR Relaxation of Heptane in Kerogen from Molecular Simulations and Measurements

Kerogen-rich shale reservoirs will play a key role during the energy transition, yet the effects of nanoconfinement on the NMR relaxation of hydrocarbons in kerogen are poorly understood. We use atomistic MD simulations to investigate the effects of nanoconfinement on the 1H NMR relaxation times T1 and T2 of heptane in kerogen. In the case of T1, we discover the important role of confinement in reducing T1 by ∼3 orders of magnitude from that of bulk heptane, in agreement with measurements of heptane dissolved in kerogen from the Kimmeridge Shale, without any models or free parameters. In the case of T2, we discover that confinement breaks spatial isotropy and gives rise to residual dipolar coupling which reduces T2 by ∼5 orders of magnitude from the value for bulk heptane. We use the simulated T2 to calibrate the surface relaxivity and thence predict the pore-size distribution of the organic nanopores in kerogen, without additional experimental data.

Mechanisms by which small molecules of diverse chemotypes arrest Sec14 lipid transfer activity

Phosphatidylinositol (PtdIns) transfer proteins (PITPs) enhance the activities of PtdIns 4-OH kinases that generate signaling pools of PtdIns-4-phosphate. In that capacity, PITPs serve as key regulators of lipid signaling in eukaryotic cells. Although the PITP phospholipid exchange cycle is the engine that stimulates PtdIns 4-OH kinase activities, the underlying mechanism is not understood. Herein, we apply an integrative structural biology approach to investigate interactions of the yeast PITP Sec14 with small-molecule inhibitors (SMIs) of its phospholipid exchange cycle. Using a combination of X-ray crystallography, solution NMR spectroscopy, and atomistic MD simulations, we dissect how SMIs compete with native Sec14 phospholipid ligands and arrest phospholipid exchange. Moreover, as Sec14 PITPs represent new targets for the development of next-generation antifungal drugs, the structures of Sec14 bound to SMIs of diverse chemotypes reported in this study will provide critical information required for future structure-based design of next-generation lead compounds directed against Sec14 PITPs of virulent fungi.

The impact of bilayer composition on the dimerization properties of the Slg1 stress sensor TMD from a multiscale analysis.

The impact of mutual interactions between the transmembrane domains of membrane proteins and lipids on bilayer properties has gained major attraction. Most simulation studies of membranes rely on the Martini force field, which has proven extremely helpful in providing molecular insights into realistic systems. Accordingly, an evaluation of the accuracy of the Martini force field is crucial to be able to correctly interpret the reported data. In this study, we combine atomistic and coarse-grained Martini simulations to investigate the properties of transmembrane domains (TMDs) in a model yeast membrane. The results show that the TMD binding state (monomeric and dimeric with positive or negative crossing angle) and the membrane composition significantly influence the properties around the TMDs and change TMD-TMD and TMD-lipid affinities. Furthermore, ergosterol (ERG) exhibits a strong affinity to TMD dimers. Importantly, the right-handed TMD dimer configuration is stabilized via TMD-TMD contacts by the addition of asymmetric anionic phosphatidylserine (PS). The coarse-grained simulations corroborate many of these findings, with two notable exceptions: a systematic overestimation of TMD-ERG interaction and lack of stabilization of the right-handed TMD dimers with the addition of PS.

Investigation of structure and properties of polymerizable deep eutectic solvent based on choline chloride and acrylic acid

Deep eutectic solvents (DESs) are multi-component solvents appearing in a broad range of applications. The next necessary step for the development of new DESs is understanding the molecular mechanisms of DES formation and the interactions that determine its structure and properties. In this work, we use multiscale simulations supported by experiments to investigate the detailed structure and properties of polymerizable DESs based on choline chloride and acrylic acid as a basis for creating inks for 3D printing. Thermodynamic and structural analyses show the physical mechanisms of DES formation in these materials: due to the significant size difference between the acrylic acid and choline ions, and favorable interactions between acrylic acid and the Cl- ions, the acrylic acid molecules are able to incorporate into the free spaces of the first coordination shells of the Cl- ions. As a consequence, the mixture has less volume than its individual components and this excess volume determines the negative value of the enthalpy of mixing. Structurally, the mixture is a network with the Cl- ions as nodes connecting the other DES components. This was confirmed by both the FTIR experiments and the atomistic MD simulations. The calculations show the necessity of correct accounting of excess enthalpy and entropy for determining DESs structures and other properties.

Probing Phase Transitions in Organic Crystals Using Atomistic MD Simulations

Probing Phase Transitions in Organic Crystals Using Atomistic MD Simulations

New insights into adsorption structure and hydration of polymer at oil-water interface obtained by molecular dynamics simulations: Isotactic poly(methacrylic acid)

We have performed atomistic MD simulations to investigate structure and hydration properties of isotactic Poly(methacrylic acid) (i-PMA) at CCl4–H2O interface at different concentration up to monolayer coverage. Three concentration regimes dilute, semi-dilute and concentrated are shown to occur at molecular level detail for the first time for polymer systems at oil-water interface. i-PMA chains at low interface concentration exists in a planar conformation along with an ordered arrangement of side groups and changes to a random coil structure at high concentration, which is observed from the variation of components of Rg and decrease in preferential orientation of groups. An abrupt change in chain conformation marks the end of the dilute regime and a rapid increase in thickness of adsorbed layer marks the beginning of the concentrated regime. This study provides new molecular insights into conformational changes and the associated changes in interactions among different chemical groups with variation of the interface concentration of polymer.

Probing effects of the SARS-CoV-2 E protein on membrane curvature and intracellular calcium

SARS-CoV-2 contains four structural proteins in its genome. These proteins aid in the assembly and budding of new virions at the ER-Golgi intermediate compartment (ERGIC). Current fundamental research efforts largely focus on one of these proteins – the spike (S) protein. Since successful antiviral therapies are likely to target multiple viral components, there is considerable interest in understanding the biophysical role of its other structural proteins, in particular structural membrane proteins. Here, we have focused our efforts on the characterization of the full-length envelope (E) protein from SARS-CoV-2, combining experimental and computational approaches. Recombinant expression of the full-length E protein from SARS-CoV-2 reveals that this membrane protein is capable of independent multimerization, possibly as a tetrameric or smaller species. Fluorescence microscopy shows that the protein localizes intracellularly, and coarse-grained MD simulations indicate it causes bending of the surrounding lipid bilayer, corroborating a potential role for the E protein in viral budding. Although we did not find robust electrophysiological evidence of ion-channel activity, cells transfected with the E protein exhibited reduced intracellular Ca2+, which may further promote viral replication. However, our atomistic MD simulations revealed that previous NMR structures are relatively unstable, and result in models incapable of ion conduction. Our study highlights the importance of using high-resolution structural data obtained from a full-length protein to gain detailed molecular insights, and eventually permitting virtual drug screening.