Dynamics and Structures of Amyloid Aggregates under Fluid Flows.

In this work, we investigate how fluid flows impact the aggregation mechanisms of Aß40 proteins and Aß16-22 peptides and mechanically perturb their (pre)fibrillar aggregates. We exploit the OPEP coarse-grained model for proteins and the Lattice Boltzmann Molecular Dynamics technique. We show that beyond a critical shear rate, amyloid aggregation speeds up in Couette flow because of the shorter collisions times between aggregates, following a transition from diffusion limited to advection dominated dynamics. We also characterize the mechanical deformation of (pre)fibrillar states due to the fluid flows (Couette and Poiseuille), confirming the capability of (pre)fibrils to form pathological loop-like structures as detected in experiments. Our findings can be of relevance for microfluidic applications and for understanding aggregation in the interstitial brain space.

Structure and Elasticity of Mitochondrial Membranes: A Molecular Dynamics Simulation Study.

Mitochondria are known as the powerhouse of the cell because they produce energy in the form of adenosine triphosphate. They also have other crucial functions such as regulating apoptosis, calcium homeostasis, and reactive oxygen species production. To perform these diverse functions, mitochondria adopt specific structures and frequently undergo dynamic shape changes, indicating that their mechanical properties play an essential role in their functions. To gain a detailed understanding at the molecular level of the structure and mechanical properties of mitochondria, we carry out atomistic molecular dynamics simulations for three inner mitochondrial membranes and three outer mitochondrial membrane models. These models take into account variations in cardiolipin and cholesterol concentrations as well as the symmetry/asymmetry between the two leaflets. Our simulations allow us to calculate various structural quantities and the bending, twisting, and tilting elastic moduli of the membrane models. Our results indicate that the structures of the inner and outer mitochondrial membranes are quite similar and do not depend much on the variation in lipid compositions. However, the bending modulus of the membranes increases with increasing concentrations of cardiolipin or cholesterol but decreases with a membrane asymmetry. Notably, we found that the dipole potential of the membrane increases with an increasing cardiolipin concentration. Finally, possible roles of cardiolipin in regulating ion and proton currents and maintaining the cristate are discussed in some details.

Recent Computational Advances Regarding Amyloid-ß and Tau Membrane Interactions in Alzheimer's Disease.

The interactions of amyloid proteins with membranes have been subject to many experimental and computational studies, as these interactions contribute in part to neurodegenerative diseases. In this review, we report on recent simulations that have focused on the adsorption and insertion modes of amyloid-ß and tau proteins in membranes. The atomistic-resolution characterization of the conformational changes of these amyloid proteins upon lipid cell membrane and free lipid interactions is of interest to rationally design drugs targeting transient oligomers in Alzheimer's disease.

Multistep molecular mechanisms of Aß16-22 fibril formation revealed by lattice Monte Carlo simulations.

As a model of self-assembly from disordered monomers to fibrils, the amyloid-ß fragment Aß16-22 was subject to past numerous experimental and computational studies. Because dynamics information between milliseconds and seconds cannot be assessed by both studies, we lack a full understanding of its oligomerization. Lattice simulations are particularly well suited to capture pathways to fibrils. In this study, we explored the aggregation of 10 Aß16-22 peptides using 65 lattice Monte Carlo simulations, each simulation consisting of 3 × 109 steps. Based on a total of 24 and 41 simulations that converge and do not converge to the fibril state, respectively, we are able to reveal the diversity of the pathways leading to fibril structure and the conformational traps slowing down the fibril formation.

PEP-FOLD4: a pH-dependent force field for peptide structure prediction in aqueous solution.

Accurate and fast structure prediction of peptides of less 40 amino acids in aqueous solution has many biological applications, but their conformations are pH- and salt concentration-dependent. In this work, we present PEP-FOLD4 which goes one step beyond many machine-learning approaches, such as AlphaFold2, TrRosetta and RaptorX. Adding the Debye-Hueckel formalism for charged-charged side chain interactions to a Mie formalism for all intramolecular (backbone and side chain) interactions, PEP-FOLD4, based on a coarse-grained representation of the peptides, performs as well as machine-learning methods on well-structured peptides, but displays significant improvements for poly-charged peptides. PEP-FOLD4 is available at http://bioserv.rpbs.univ-paris-diderot.fr/services/PEP-FOLD4. This server is free and there is no login requirement.

Structure and elasticity of healthy and Alzheimer's disease cell membranes revealed by molecular dynamics simulations.

Interactions of amyloid-ß (Aß) peptides with neuronal membrane are associated with the development of Alzheimer's disease (AD). Ganglioside monosialotetrahexosylganglioside (GM1) lipids have been shown to form clusters that induce the structural conversion of Aß and promote the incorporation of Aß into the membrane via the membrane surface electrical potential. Prior to the onset of AD symptoms, GM1 clusters may not have formed but the concentration of GM1 may have already changed, and our question is whether this early concentration modification affects the structure and mechanical properties of the membrane. Using one model for healthy cell membranes and three models for AD cell membranes, we carry out 2 µs all-atom molecular dynamics simulations for each model to compare the structure and elasticity of the two membrane types. The simulations show that at the physiological concentration, 1%-3%, GM1 does not form clusters. The reduction of the GM1 lipid does not significantly alter the area per lipid, the membrane thickness, and the lipid order parameters of the AD membranes. However, the dipole potential, the bending, and twist moduli are decreased for the AD membranes. We suggest that these changes in the AD membranes are factors that could trigger the interaction and incorporation of Aß to the membranes. Finally, we show that changes in the sphingomyelin lipid concentrations do not affect the membrane structure and elasticity.

Evolution of large Aß16-22 aggregates at atomic details and potential of mean force associated to peptide unbinding and fragmentation events.

Atomic characterization of large nonfibrillar aggregates of amyloid polypeptides cannot be determined by experimental means. Starting from ß-rich aggregates of Y and elongated topologies predicted by coarse-grained simulations and consisting of more than 100 Aß16-22 peptides, we performed atomistic molecular dynamics (MD), replica exchange with solute scaling (REST2), and umbrella sampling simulations using the CHARMM36m force field in explicit solvent. Here, we explored the dynamics within 3 µs, the free energy landscape, and the potential of mean force associated with either the unbinding of one single peptide in different configurations within the aggregate or fragmentation events of a large number of peptides. Within the time scale of MD and REST2, we find that the aggregates experience slow global conformational plasticity, and remain essentially random coil though we observe slow beta-strand structuring with a dominance of antiparallel beta-sheets over parallel beta-sheets. Enhanced REST2 simulation is able to capture fragmentation events, and the free energy of fragmentation of a large block of peptides is found to be similar to the free energy associated with fibril depolymerization by one chain for longer Aß sequences.

Metastable alpha-rich and beta-rich conformations of small Aß42 peptide oligomers.

Probing the structures of amyloid-ß (Aß) peptides in the early steps of aggregation is extremely difficult experimentally and computationally. Yet, this knowledge is extremely important as small oligomers are the most toxic species. Experiments and simulations on Aß42 monomer point to random coil conformations with either transient helical or ß-strand content. Our current conformational description of small Aß42 oligomers is funneled toward amorphous aggregates with some ß-sheet content and rare high energy states with well-ordered assemblies of ß-sheets. In this study, we emphasize another view based on metastable α-helix bundle oligomers spanning the C-terminal residues, which are predicted by the machine-learning AlphaFold2 method and supported indirectly by low-resolution experimental data on many amyloid polypeptides. This finding has consequences in developing novel chemical tools and to design potential therapies to reduce aggregation and toxicity.

Optimized OPEP Force Field for Simulation of Crowded Protein Solutions.

Macromolecular crowding has profound effects on the mobility of proteins, with strong implications on the rates of intracellular processes. To describe the dynamics of crowded environments, detailed molecular models are needed, capturing the structures and interactions arising in the crowded system. In this work, we present OPEPv7, which is a coarse-grained force field at amino-acid resolution, suited for rigid-body simulations of the structure and dynamics of crowded solutions formed by globular proteins. Using the OPEP protein model as a starting point, we have refined the intermolecular interactions to match the experimentally observed dynamical slowdown caused by crowding. The resulting force field successfully reproduces the diffusion slowdown in homogeneous and heterogeneous protein solutions at different crowding conditions. Coupled with the lattice Boltzmann technique, it allows the study of dynamical phenomena in protein assemblies and opens the way for the in silico rheology of protein solutions.

A refined pH-dependent coarse-grained model for peptide structure prediction in aqueous solution.

Introduction: Peptides carry out diverse biological functions and the knowledge of the conformational ensemble of polypeptides in various experimental conditions is important for biological applications. All fast dedicated softwares perform well in aqueous solution at neutral pH. Methods: In this study, we go one step beyond by combining the Debye-Hückel formalism for charged-charged amino acid interactions and a coarse-grained potential of the amino acids to treat pH and salt variations. Results: Using the PEP-FOLD framework, we show that our approach performs as well as the machine-leaning AlphaFold2 and TrRosetta methods for 15 well-structured sequences, but shows significant improvement in structure prediction of six poly-charged amino acids and two sequences that have no homologous in the Protein Data Bank, expanding the range of possibilities for the understanding of peptide biological roles and the design of candidate therapeutic peptides.

An S-Shaped Aß42 Cross-ß Hexamer Embedded into a Lipid Bilayer Reveals Membrane Disruption and Permeability.

The interactions of amyloid oligomers with membranes are known to contribute to cellular toxicity. Numerous in vitro experimental studies reported on the insertion of oligomers of different sizes that can induce cell membrane disruption, extract lipids, and form ion-permeable transmembrane pores. The current repertoire of amyloid-beta (Aß) membrane-inserted folds that was subject to high-resolution structure NMR spectroscopy and computer simulations is devoid of any cross-ß fibrillar structure. In this study, we explored the dynamics of an S-shaped Aß42 cross-ß hexamer model inserted into a lipid bilayer membrane by two atomistic molecular dynamics simulations. The initial model is characterized by the hydrophobic residues at the central hydrophobic core (residues 17-21, CHC) and the C-terminus (residues 30-42) embedded into the membrane. We observed major structural secondary, tertiary, and quaternary rearrangements leading to two distinct species, hexamer and two trimers, accompanied by membrane disruption and water permeation. The simulations show that some configurations, but not the majority, have the CHC and C-terminus hydrophobic residues exposed to the solvent. Overall, our computational results offer new perspectives to understand the relationship between Aß42 assemblies and membrane permeability.

Diffusive Dynamics of Bacterial Proteome as a Proxy of Cell Death.

Temperature variations have a big impact on bacterial metabolism and death, yet an exhaustive molecular picture of these processes is still missing. For instance, whether thermal death is determined by the deterioration of the whole or a specific part of the proteome is hotly debated. Here, by monitoring the proteome dynamics of E. coli, we clearly show that only a minor fraction of the proteome unfolds at the cell death. First, we prove that the dynamical state of the E. coli proteome is an excellent proxy for temperature-dependent bacterial metabolism and death. The proteome diffusive dynamics peaks at about the bacterial optimal growth temperature, then a dramatic dynamical slowdown is observed that starts just below the cell's death temperature. Next, we show that this slowdown is caused by the unfolding of just a small fraction of proteins that establish an entangling interprotein network, dominated by hydrophobic interactions, across the cytoplasm. Finally, the deduced progress of the proteome unfolding and its diffusive dynamics are both key to correctly reproduce the E. coli growth rate.

Insights into the Mixture of Aß24 and Aß42 Peptides from Atomistic Simulations.

Amyloid-ß (Aß) oligomers play a central role in Alzheimer's disease (AD). Plaques of AD patients consist of Aß40 and Aß42 peptides and truncated Aß peptides. The Aß24 peptide, identified in human AD brains, was found to impair Aß42 clearance through the brain-blood barrier. The Aß24 peptide was also shown to reduce Aß42 aggregation kinetics in pure buffer, but the underlying mechanism is unknown at atomistic level. In this study, we explored the conformational ensemble of the equimolar mixture of Aß24 and Aß42 by replica exchange molecular dynamics simulations and compared it to our previous results on the pure Aß42 dimer. Our simulations demonstrate that the truncation at residue 24 changes the secondary, tertiary, and quaternary structures of the dimer, offering an explanation of the slower aggregation kinetics of the mixture.

Self-Assembly of Amyloid-Beta (Aß) Peptides from Solution to Near In Vivo Conditions.

Understanding the atomistic resolution changes during the self-assembly of amyloid peptides or proteins is important to develop compounds or conditions to alter the aggregation pathways and suppress the toxicity and potentially aid in the development of drugs. However, the complexity of protein aggregation and the transient order/disorder of oligomers along the pathways to fibril are very challenging. In this Perspective, we discuss computational studies of amyloid polypeptides carried out under various conditions, including conditions closely mimicking in vivo and point out the challenges in obtaining physiologically relevant results, focusing mainly on the amyloid-beta Aß peptides.

Molecular dynamics simulation of cancer cell membrane perforated by shockwave induced bubble collapse.

It has been widely accepted that cancer cells are softer than their normal counterparts. This motivates us to propose, as a proof-of-concept, a method for the efficient delivery of therapeutic agents into cancer cells, while normal cells are less affected. The basic idea of this method is to use a water jet generated by the collapse of the bubble under shockwaves to perforate pores in the cell membrane. Given a combination of shockwave and bubble parameters, the cancer membrane is more susceptible to bending, stretching, and perforating than the normal membrane because the bending modulus of the cancer cell membrane is smaller than that of the normal cell membrane. Therefore, the therapeutic agent delivery into cancer cells is easier than in normal cells. Adopting two well-studied models of the normal and cancer membranes, we perform shockwave induced bubble collapse molecular dynamics simulations to investigate the difference in the response of two membranes over a range of shockwave impulse 15-30 mPa s and bubble diameter 4-10 nm. The simulation shows that the presence of bubbles is essential for generating a water jet, which is required for perforation; otherwise, pores are not formed. Given a set of shockwave impulse and bubble parameters, the pore area in the cancer membrane is always larger than that in the normal membrane. However, a too strong shockwave and/or too large bubble results in too fast disruption of membranes, and pore areas are similar between two membrane types. The pore closure time in the cancer membrane is slower than that in the normal membrane. The implications of our results for applications in real cells are discussed in some details. Our simulation may be useful for encouraging future experimental work on novel approaches for cancer treatment.

Impact of A2T and D23N mutations on C99 homodimer conformations.

The proteolytic cleavage of C99 by γ-secretase is the last step in the production of amyloid-ß (Aß) peptides. Previous studies have shown that membrane lipid composition, cholesterol concentration, and mutation in the transmembrane helix modified the structures and fluctuations of C99. In this study, we performed atomistic molecular dynamics simulations of the homodimer of the 55-residue congener of the C-terminal domain of the amyloid protein precursor, C99(1-55), in a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine-cholesterol lipid bilayer and compared the conformational ensemble of wild-type (WT) sequence to those of the A2T and D23N variants. These mutations are particularly interesting as the protective Alzheimer's disease (AD) A2T mutation is known to decrease Aß production, whereas the early onset AD D23N mutation does not affect Aß production. We found noticeable differences in the structural ensembles of the three sequences. In particular, A2T varies from both WT and D23N by having long-range effects on the population of the extracellular juxtamembrane helix, the interface between the G29xxx-G33xxx-G37 motifs, and the fluctuations of the transmembrane helical topologies.

Molecular Dynamics Simulations of the Tau Amyloid Fibril Core Dimer at the Surface of a Lipid Bilayer Model: I. In Alzheimer's Disease.

A tau R3-R4 domain spanning residues 306-378 was shown to form an amyloid fibril core of a full-length tau in the brain of patients with Alzheimer's disease. Recently, we studied the dynamics of a tau R3-R4 monomer at the surface of a lipid bilayer model and revealed deep insertion of the amino acids spanning the PHF6 motif (residues 306-311) and its flanking residues. Here, we explore the membrane-associated conformational ensemble of a tau R3-R4 dimer by means of atomistic molecular dynamics. Similar to the monomer simulation, the R3-R4 dimer has the propensity to form ß-hairpin-like conformation. Unlike the monomer, the dimer shows insertion of the C-terminal R4 region and transient adsorption of the PHF6 motif. Taken together, these results reveal the multiplicity of adsorption and insertion modes of tau into membranes depending on its oligomer size.

Molecular Dynamics Simulations of the Tau R3-R4 Domain Monomer in the Bulk Solution and at the Surface of a Lipid Bilayer Model.

The aggregation of the tau protein plays a significant role in Alzheimer's disease, and the tau R3-R4 domain spanning residues 306-378 was shown to form the amyloid fibril core of a full-length tau. The conformations of the tau R3-R4 monomer in the bulk solution and at the surface of membranes are unknown. In this study, we address these questions by means of atomistic molecular dynamics. The simulations in the bulk solution show a very heterogeneous ensemble of conformations with low ß and helical contents. The tau R3-R4 monomer has the propensity to form transient ß-hairpins within the R3 repeat and between the R3 and R4 repeats and parallel ß-sheets spanning the R3 and R4 repeats. The simulations also show that the surface of the membrane does not induce ß-sheet insertion and leads to an ensemble of structures very different from those in the bulk solution. They also reveal the dynamical properties of the membrane-bound state of the tau R3-R4 monomer, enabling insertion of the residues 306-318 and 376-378.

Elastic moduli of normal and cancer cell membranes revealed by molecular dynamics simulations.

Recent studies indicate that there are mechanical differences between normal cells and cancer cells. Because the cell membrane takes part in a variety of vital processes, we test the hypothesis of whether or not two fundamental alterations in the cell membrane, i.e., the overexpression of phosphatidylserine lipids in the outer leaflet and a reduction in cholesterol concentration, could cause the softening in cancer cells. Adopting ten models of normal and cancer cell membranes, we carry out 1 µs all-atom molecular dynamics simulations to compare the structural properties and elasticity properties of two membrane types. We find that the overexpression of the phosphatidylserine lipids in the outer leaflet does not significantly alter the area per lipid, the membrane thickness, the lipid order parameters and the elasticity moduli of the cancer membranes. However, a reduction in the cholesterol concentration leads to clear changes in those quantities, especially decreases in the bending, tilt and twist moduli. This implies that the reduction of cholesterol concentration in the cancer membranes could contribute to the softening of cancer cells.

Small Oligomers of Aß42 Protein in the Bulk Solution with AlphaFold2.

Aggregation of amyloid-ß (Aß42) protein is one hallmark of Alzheimer's disease, and the conformations of the smallest Aß42 oligomers are largely unknown. Here, we explore the application of the deep learning AlphaFold2 method to the structure determination of Aß42 monomers up to hexamers. The results shed light on the early Aß42 aggregation steps in the bulk solution.