The stability and dynamics of the Aß40/Aß42 interlaced mixed fibrils.

The accumulation of fibrillar amyloid-ß (Aß) aggregates in the brain, predominantly comprising 40- and 42-residue amyloid-ß (Aß40 and Aß42), is a major pathological hallmark of Alzheimer's disease (AD). Aß40 and Aß42 naturally coexist in the brain under normal physiological conditions, and their interplay is generally considered to be a critical factor in the progression of AD. In addition to forming homogeneous oligomers and fibrils, Aß40 and Aß42 are also reported to co-assemble into hetero-oligomers and interlaced mixed fibrils, as evidenced by solid-state nuclear magnetic resonance spectroscopy (NMR), high molecular weight mass spectrometry and cross-seeding experiments. However, the exact molecular mechanisms underlying these processes remain unclear. In this study, we have used a recently resolved structurally uniform 1:1 mixture of Aß40/Aß42 interlaced mixed fibril as a prototype to gain insights into the molecular-level interactions between Aß40 and Aß42. We employed fully atomistic molecular dynamics simulation and compared the results with a homogeneous U-shaped Aß40 fibrillar model. Our simulations using two different force fields provide conclusive evidence that the Aß40/Aß42 interlaced mixed fibril is energetically more favorable than the homogeneous Aß40 fibrillar model. Furthermore, we also show that the increase in stability observed in the mixed model stems primarily from the packing interfaces and the stacking interfaces between C-termini. Our simulation results provide valuable mechanistic insights that are not readily accessible in experiment and could have significant implications for both the pathogenesis of AD and the development of current therapeutic strategies.Communicated by Ramaswamy H. Sarma.

Non-equilibrium virus particle dynamics: Microsecond MD simulations of the complete Flock House virus capsid under different conditions.

Flock House virus (FHV) is an animal virus and considered a model system for non-enveloped viruses. It has a small, icosahedral capsid (T=3) and a bipartite positive-sense RNA genome. We present an extensive study of the FHV capsid dynamics from all-atom molecular dynamics simulations of the complete capsid. The simulations explore different biologically relevant conditions (neutral/low pH, with/without RNA in the capsid) using the CHARMM force field. The results show that low pH destabilizes the capsid, causing radial expansion, and RNA stabilizes the capsid. The finding of low pH destabilization is biologically relevant because the capsid is exposed to low pH in the endosome, where conformational changes occur leading to genome release. We also observe structural changes at the fivefold and twofold symmetry axes that likely relate to the externalization of membrane active γ peptides through the fivefold vertex and extrusion of RNA at the twofold axis. Simulations using the Amber force field at neutral pH are also performed and display similar characteristics to the CHARMM simulations.

Effect of an Amyloidogenic SARS-COV-2 Protein Fragment on α-Synuclein Monomers and Fibrils.

Aggregates of α-synuclein are thought to be the disease-causing agent in Parkinson's disease. Various case studies have hinted at a correlation between COVID-19 and the onset of Parkinson's disease. For this reason, we use molecular dynamics simulations to study whether amyloidogenic regions in SARS-COV-2 proteins can initiate and modulate aggregation of α-synuclein. As an example, we choose the nine-residue fragment SFYVYSRVK (SK9), located on the C-terminal of the envelope protein of SARS-COV-2. We probe how the presence of SK9 affects the conformational ensemble of α-synuclein monomers and the stability of two resolved fibril polymorphs. We find that the viral protein fragment SK9 may alter α-synuclein amyloid formation by shifting the ensemble toward aggregation-prone and preferentially rod-like fibril seeding conformations. However, SK9 has only a small effect on the stability of pre-existing or newly formed fibrils. A potential mechanism and key residues for potential virus-induced amyloid formation are described.

Effect of an amyloidogenic SARS-COV-2 protein fragment on α-synuclein monomers and fibrils.

Using molecular dynamic simulations we study whether amyloidogenic regions in viral proteins can initiate and modulate formation of α-synuclein aggregates, thought to be the disease-causing agent in Parkinson's Disease. As an example we choose the nine-residue fragment SFYVYSRVK (SK9), located on the C-terminal of the Envelope protein of SARS-COV-2. We probe how the presence of SK9 affects the conformational ensemble of α-synuclein monomers and the stability of two resolved fibril polymorphs. We find that the viral protein fragment SK9 may alter α-synuclein amyloid formation by shifting the ensemble toward aggregation-prone and preferentially rod-like fibril seeding conformations. However, SK9 has only little effect of the stability of pre-existing or newly-formed fibrils.

Small Peptides for Inhibiting Serum Amyloid A Aggregation.

Deposition of human serum amyloid A (SAA) amyloids in blood vessels, causing inflammation, thrombosis, and eventually organ damage, is commonly seen as a consequence of certain cancers and inflammatory diseases and may also be a risk after SARS-COV-2 infections. Several attempts have been made to develop peptide-based drugs that inhibit or at least slow down SAA amyloidosis. We use extensive all-atom molecular dynamic simulations to compare three of these drug candidates for their ability to destabilize SAA fibrils and to propose for the best candidate, the N-terminal sequence SAA1-5, a mechanism for inhibition. As the lifetime of peptide drugs can be increased by replacing l-amino acids with their mirror d-amino acids, we have also studied corresponding d-peptides. We find that DRI-SAA1-5, formed of d-amino acids with the sequence of the peptide reversed, has similar inhibitory properties compared to the original l-peptide and therefore may be a promising candidate for drugs targeting SAA amyloidosis.

Presence of a SARS-CoV-2 Protein Enhances Amyloid Formation of Serum Amyloid A.

A marker for the severeness and disease progress of COVID-19 is overexpression of serum amyloid A (SAA) to levels that in other diseases are associated with a risk for SAA amyloidosis. To understand whether SAA amyloidosis could also be a long-term risk of SARS-CoV-2 infections, we have used long all-atom molecular dynamic simulations to study the effect of a SARS-CoV-2 protein segment on SAA amyloid formation. Sampling over 40 µs, we find that the presence of the nine-residue segment SK9, located at the C-terminus of the envelope protein, increases the propensity for SAA fibril formation by three mechanisms: it reduces the stability of the lipid-transporting hexamer shifting the equilibrium toward monomers, it increases the frequency of aggregation-prone configurations in the resulting chains, and it raises the stability of SAA fibrils. Our results therefore suggest that SAA amyloidosis and related pathologies may be a long-term risk of SARS-CoV-2 infections.

Presence of a SARS-COV-2 protein enhances Amyloid Formation of Serum Amyloid A.

A marker for the severeness and disease progress of COVID-19 is overexpression of serum amyloid A (SAA) to levels that in other diseases are associated with a risk for SAA amyloidosis. In order to understand whether SAA amyloidosis could also be a long-term risk of SARS-COV-2 infections we have used long all-atom molecular dynamic simulations to study the effect of a SARS-COV-2 protein segment on SAA amyloid formation. Sampling over 40 µs we find that presence of the nine-residue segment SK9, located at the C-terminus of the Envelope protein, increases the propensity for SAA fibril formation by three mechanisms: it reduces the stability of the lipid-transporting hexamer shifting the equilibrium toward monomers, it increases the frequency of aggregation-prone configurations in the resulting chains, and it raises the stability of SAA fibrils. Our results therefore suggest that SAA amyloidosis and related pathologies may be a long-term risk of SARS-COV-2 infections.

Atomistic dynamics of a viral infection process: Release of membrane lytic peptides from a non-enveloped virus.

Molecular simulations have played an instrumental role in uncovering the structural dynamics and physical properties of virus capsids. In this work, we move beyond equilibrium physicochemical characterization of a virus system to study a stage of the infection process that is required for viral proliferation. Despite many biochemical and functional studies, the molecular mechanism of host cell entry by non-enveloped viruses remains largely unresolved. Flock House virus (FHV) is a model system for non-enveloped viruses and is the subject of the current study. FHV infects through the acid-dependent endocytic pathway, where low pH triggers externalization of membrane-disrupting (γ) peptides from the capsid interior. Using all-atom equilibrium and enhanced sampling simulations, the mechanism and energetics of γ peptide liberation and the effect of pH on this process are investigated. Our computations agree with experimental findings and reveal nanoscopic details regarding the pH control mechanism, which are not readily accessible in experiments.

Effect of Lauric Acid on the Stability of Aß42 Oligomers.

While Alzheimer's disease is correlated with the presence of Aß fibrils in patient brains, the more likely agents are their precursors, soluble oligomers that may form pores or otherwise distort cell membranes. Using all-atom molecular dynamics simulation, we study how the presence of fatty acids such as lauric acid changes the stability of pore-forming oligomers built from three-stranded Aß42 chains. Such a change would alter the distribution of amyloids in the fatty acid-rich brain environment and therefore could explain the lower polymorphism observed in Aß fibrils derived from brains of patients with Alzheimer's disease. We find that lauric acid stabilizes both ring-like and barrel-shaped models, with the effect being stronger for barrel-like models than for ring-like oligomers.

Structural and dynamic asymmetry in icosahedrally symmetric virus capsids.

A common characteristic of virus capsids is icosahedral symmetry, yet these highly symmetric structures can display asymmetric features within their virions and undergo asymmetric dynamics. The fields of structural and computational biology have entered a new realm in the investigation of virus infection mechanisms, with the ability to observe symmetry-breaking features. This review will cover important studies on icosahedral virus structure and dynamics, covering both symmetric and asymmetric conformational changes. However, the main emphasis of the review will be towards recent studies employing cryo-electron microscopy or molecular dynamics simulations, which can uncover asymmetric aspects of these systems relevant to understanding viral physical-chemical properties and their biological impact.

Structural and dynamics studies of the TetR family protein, CprB from Streptomyces coelicolor in complex with its biological operator sequence.

In Streptomycetes, tetracycline repressor family of transcription regulators (TetR-FTRs) controls various biological processes including antibiotic biosynthesis, cellular morphology and innate resistance. Here, we focus on understanding the structural basis of transcription regulation by CprB, a member of TetR-FTRs from S. coelicolor. CprB is implicated as a receptor of γ-butyrolactones, a class of quorum sensing molecules, responsible for initiating secondary metabolic pathways. In order to understand the molecular mechanism of DNA recognition, the X-ray structure of CprB in complex with its biological relevant operator sequence was solved to a resolution of 3.95Å. Furthermore, to refine and compliment the results, atomistic molecular dynamics simulations were carried out using the X-ray structure as the template. The studies reveal that CprB binds to DNA as dimer of dimers with this mode of interaction results in minimal distortion in the DNA, enabling these proteins to recognize multiple sequences with varying affinity. Another crucial finding from our simulation results was that the positively charged N-terminal arm of CprB brings extra stability to the protein-DNA complex by interacting with the minor-groove of the DNA and anchoring itself to the phosphate backbone. Corroborating electrophoretic mobility shift assay and fluorescence anisotropy experiments showed that the mutant ΔN6-CprB exhibited about 7-8 fold reduced DNA binding. Comparison with other TetR-FTRs reveals that this strategy is also employed by over 25% of TetR-FTRs, where N-terminal anchoring mechanism is used to enhance selectivity for a particular DNA sequence.

Glycation induces conformational changes in the amyloid-ß peptide and enhances its aggregation propensity: molecular insights.

The cytotoxicity of the amyloid beta (Aß) peptide, implicated in the pathogenesis of Alzheimer's disease (AD), can be enhanced by its post-translational glycation, a series of non-enzymatic reactions with reducing sugars and reactive dicarbonyls. However, little is known about the underlying mechanisms that potentially enhance the cytotoxicity of the advanced glycation modified Aß. In this work, fully atomistic molecular dynamics (MD) simulations are exploited to obtain direct molecular insights into the process of early Aß self-assembly in the presence and absence of glycated lysine residues. Analyses of data exceeding cumulative timescales of 1 microsecond for each system reveal that glycation results in a stronger enthalpy of association between Aß monomers and lower conformational entropy, in addition to a sharp overall increase in the beta-sheet content. Further analyses reveal that the enhanced interactions originate, in large part, due to markedly stronger, as well as new, inter-monomer salt bridging propensities in the glycated variety. Interestingly, these conformational and energetic effects are broadly reflected in preformed protofibrillar forms of Aß small oligomers modified with glycation. Our combined results imply that glycation consolidates Aß self-assembly regardless of its point of occurrence in the pathway. They provide a basis for further mechanistic studies and therapeutic endeavors that could potentially result in novel ways of combating AGE related AD progression.

Unraveling origins of the heterogeneous curvature dependence of polypeptide interactions with carbon nanostructures.

Emerging nanotechnology has rapidly broadened interfacial prospects of biological molecules with carbon nanomaterials (CNs). A prerequisite for effectively harnessing such hybrid materials is a multi-faceted understanding of their complex interfacial interactions as functions of the physico-chemical characteristics and the surface topography of the individual components. In this article, we address the origins of the curvature dependence of polypeptide adsorption on CN surfaces (CNSs), a phenomenon bearing an acute influence upon the behavior and activity of CN-protein conjugates. Our benchmark molecular dynamics (MD) simulations with the amphiphilic full-length amyloid beta (Aß) peptide demonstrate that protein adsorption is strongest on the concave (inner) CN surface, weakest on the convex (outer) surface, and intermediary on the planar surface, in agreement with recent experimental reports. The curvature effects, however, are found to manifest non-uniformly between the amino acid subtypes. To understand the underlying interplay of the chemical nature of the amino acids and surface topography of the CNs, we performed high-level quantum chemical (QM) calculations with amino acid analogs (AAA) representing their five prominent classes, and convex, concave and planar CN fragments. Molecular electrostatic potential maps reveal pronounced curvature dependence in the mixing of electron densities, and a resulting variance in the stabilization of the non-covalently bound molecular complexes. Interestingly, our study revealed that the interaction trends of the high-level QM calculations were captured well by the empirical force field. The findings in this study have important bearing upon the design of carbon based bio-nanomaterials, and additionally, provide valuable insights into the accuracy of various computational techniques for probing non-bonded interfacial interactions.

Aß self-association and adsorption on a hydrophobic nanosurface: competitive effects and the detection of small oligomers via electrical response.

Treatment of Alzheimer's disease (AD) is impeded by the lack of effective early diagnostic methods. Small, soluble Aß globulomers play a major role in AD neurotoxicity, and detecting their presence in aqueous fluids could lead to suitable sensors. We evaluate the adsorption behavior of small Aß oligomers on the surface of a single walled carbon nanotube of high curvature. While the intrinsic self-assembly propensity of Aß is markedly hindered by adsorption, the oligomeric units show high degrees of surface immobilization. Immobilized complexes are capable of oligomeric growth, but with a shifted monomer-oligomer equilibrium compared to the free states. In the presence of an ionic solution and suitable external electric fields, magnitudes of the current blockades are found to be sensitive to the oligomeric number of the adsorbed complex. However, this sensitivity gradually diminishes with increasing oligomeric size. The results provide a proof-of-concept basis for further investigations in the design of sensors for detecting the toxic small oligomers of Aß.

Molecular investigations of protriptyline as a multi-target directed ligand in Alzheimer's disease.

Alzheimer's disease (AD) is a complex neurodegenerative disorder involving multiple cellular and molecular processes. The discovery of drug molecules capable of targeting multiple factors involved in AD pathogenesis would greatly facilitate in improving therapeutic strategies. The repositioning of existing non-toxic drugs could dramatically reduce the time and costs involved in developmental and clinical trial stages. In this study, preliminary screening of 140 FDA approved nervous system drugs by docking suggested the viability of the tricyclic group of antidepressants against three major AD targets, viz. Acetylcholinesterase (AChE), ß-secretase (BACE-1), and amyloid ß (Aß) aggregation, with one member, protriptyline, showing highest inhibitory activity. Detailed biophysical assays, together with isothermal calorimetry, fluorescence quenching experiments, kinetic studies and atomic force microscopy established the strong inhibitory activity of protriptyline against all three major targets. The molecular basis of inhibition was supported with comprehensive molecular dynamics simulations. Further, the drug inhibited glycation induced amyloid aggregation, another important causal factor in AD progression. This study has led to the discovery of protriptyline as a potent multi target directed ligand and established its viability as a promising candidate for AD treatment.

Surface induced collapse of Aß1-42 with the F19A replacement following adsorption on a single walled carbon nanotube.

Spontaneous adsorption of the Aß peptide on the surface of a single-walled carbon nanotube, resulting in the prevention of its intrinsic propensity to form collapsed states, could be a plausible means to hinder the peptide's initial nucleation and self-assembly. We report here the effects of sharply reducing both aromatic and hydrophobic character within the peptide's central hydrophobic core on its free and surface behavior. In such an altered peptide, complete surface adsorption is found to induce, rather than prevent, the adsorbed peptide's collapse. The weakened surface interactions of the central hydrophobic core allow its greater translational mobility on the surface, thereby facilitating interactions that lead to compaction. Both the adsorption and the subsequent collapse are accompanied by a loss of surface hydration in the modified peptide. We further find that such a two-step dewetting leads to hydration levels comparable to that obtained after compaction of the free peptide. These insights may be leveraged for designing molecular surfaces for disrupting intrinsic Aß behavior.

Critical roles of key domains in complete adsorption of Aß peptide on single-walled carbon nanotubes: insights with point mutations and MD simulations.

Owing to the influence of nanomaterials on biomacromolecular behavior, their potential applications are rapidly gaining attention. Based on atomistic molecular dynamics simulation studies we have recently reported that the full-length Aß peptide, whose self-assembly is associated with Alzheimer's disease, adsorbs rapidly on single-walled carbon nanotubes, thereby losing its natural propensity to collapse. Here, we investigate the mechanistic overlap between the peptide's compactification and its adsorption, while decoupling the roles of hydrophobicity and aromaticity via point mutations. The collapse mechanism is correlated with interactions between the central hydrophobic core (HP1) and the peptide's C-terminal domain, which are almost exactly compensated by interactions arising from the nanotube after complete adsorption. Adsorption is initiated by HP1 and consolidated by strong interactions arising from the N-terminal domain. Altering the hydrophobicity, but not the aromatic character, of the central residue in HP1 decreases the collapse probability. On the other hand, the adsorption propensity is dramatically reduced when either the hydrophobicity or the aromatic character in HP1 is compromised. The hydrophobicity of HP1 is responsible for dewetting transitions that facilitate its initial interactions with the nanotube, which then lead to very favorable interactions with the nanotube.

Adsorption mechanism and collapse propensities of the full-length, monomeric Aß(1-42) on the surface of a single-walled carbon nanotube: a molecular dynamics simulation study.

Though nanomaterials such as carbon nanotubes have gained recent attention in biology and medicine, there are few studies at the single-molecule level that explore their interactions with disease-causing proteins. Using atomistic molecular-dynamics simulations, we have investigated the interactions of the monomeric Aß(1-42) peptide with a single-walled carbon nanotube of small diameter. Starting with peptide-nanotube complexes that delineate the interactions of different segments of the peptide, we find rapid convergence in the peptide's adsorption behavior on the nanotube surface, manifested in its arrested movement, the convergence of peptide-nanotube contact areas and approach distances, and in increased peptide wrapping around the nanotube. In systems where the N-terminal domain is initially distal from nanotube, the adsorption phenomena are initiated by interactions arising from the central hydrophobic core, and precipitated by those arising from the N-terminal residues. Our simulations and free energy calculations together demonstrate that the presence of the nanotube increases the energetic favorability of the open state. We note that the observation of peptide localization could be leveraged for site-specific drug delivery, while the decreased propensity of collapse appears promising for altering kinetics of the peptide's self-assembly.