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Dihydrochalcone molecules destabilize Alzheimer’s amyloid-β protofibrils through binding to the protofibril cavity
Abstract
Alzheimer’s disease (AD) is associated with the aggregation of amyloid-β (Aβ) peptides into toxic fibrillar aggregates. Finding effective inhibitors of Aβ aggregation is a crucial step for the development of drugs against AD. Recent experiments reported that dihydrochalcone (Dih), a compound extracted from daemonorops draco tree, could effectively inhibit Aβ fibrillization and reduce Aβ cytotoxicity. However, the influence of Dih molecules on preformed Aβ fibrils and the atomic-level details of interactions between Dih and Aβ fibrils are largely unknown. In this work, we performed multiple molecular dynamics (MD) simulations of 1.2 μs in total on the Aβ17-42 protofibrils with and without Dih molecules. We found that Dih molecules mostly bind to three different sites of the protofibril: the exterior central hydrophobic core (CHC) spanning residues 17LVFFA21 in β1 region, the protofibril cavity and the C-terminal hydrophobic-groove spanning residues 31IIGLM35 in β2 region. Binding to the C-terminal hydrophobic-groove slightly affects the structures of Aβ17-42 protofibrils, while binding to the exterior CHC and the cavity strongly destabilizes the protofibrils by mostly disrupting the D23-K28 salt bridges and the inter-peptide β-sheet in β1 region. The dynamics process of Dih molecules entering the cavity of Aβ17-42 protofibrils is also investigated. We also examined the effect of Dih molecules on both U-shaped Aβ40/Aβ42 protofibrils and S-shaped Aβ42 protofibrils by carrying out multiple MD simulations. Our simulations show that Dih molecules can destabilize both U-shaped and S-shaped Aβ protofibrils by binding to the protofibril cavity. This study reveals the mechanism by which Dih molecules disrupt Aβ protofibrils, which may offer new clues for the development of drug candidates for the treatment of AD.
... The increased RMSF value (Figure 5b), which accounts for increased flexibility of the fibrillar chains, is consistent with observations, where morin, [76] wgx-50, [77] ellagic acid [61] have been used as ligands. The increased average SASA value (Figure 5c) reflects the increased hydrophilicity of the fibril in the presence of lycopene, resulting in higher mobility and thus larger disorientation similar to reported for dihydrochalcone [78] and ellagic acid [61] molecule. ...
... However, not much change has been observed for chain B-E for Aβ-LYC1 over Aβ-Water system. Our observations well-match with the reported results based on dihydrochalcone, [78] and flavonoid derivative, [82] ellagic acid, [61] brazilin, [49] and a flavonoid compound [53] as the ligand. The higher value of average distance of inter-chain residues, observed in Aβ-LYC1 system elucidates the mechanistic details behind destabilization of the fibril. ...
The therapeutic strategy employing destabilization of the preformed Aβ fibril by various natural compounds, as studied by experimental and computational methods, has been reported significant in curing Alzheimer's disease (AD). However, lycopene (a carotenoid), from terpenes family, needs investigation for its destabilization potential of Aβ fibril. The highest antioxidant potential and ability to cross blood brain barrier makes lycopene a preferred choice as drug lead for treating AD. The current study focuses on investigating the destabilization potential and underpinning mechanism of lycopene on different polymorphic forms of Aβ fibril via Molecular Dynamics (MD) simulation. The key findings highlight binding of lycopene to the outer surface of the chain F of the fibril (2NAO). Herein G9, K16 and V18 residues were found to be involved in van der Waals with the methyl groups of the lycopene. Additionally, Y10 and F20 residues were observed to interact via π-π interactions with CC bonds of the lycopene. The surface mediated binding of lycopene to the fibril is attributed to the large size and structural rigidity of lycopene along with the bulky size of 2NAO and narrow space of fibrillar cavity. The destabilization of the fibril is evident by breakage of inherent H-bonds and hydrophobic interactions in the presence of one lycopene molecule. The lesser β-sheet content explains disorganization of the fibril and bars the higher order aggregation curbing neurotoxicity of the fibril. The higher concentration of the lycopene is not found to be linearly correlated with the extent of destabilization of the fibril. Lycopene is also observed to destabilize the other polymorphic form of Aβ fibril (2BEG), by accessing the fibrillar cavity and lowering the β-sheet content. The destabilization observed by lycopene on two major polymorphs of Aβ fibril explains its potency towards developing an effective therapeutic approach in treating AD.
... The increased RMSF value (Figure 5b), which accounts for increased flexibility of the fibrillar chains, is consistent with observations, where morin, [76] wgx-50, [77] ellagic acid [61] have been used as ligands. The increased average SASA value (Figure 5c) reflects the increased hydrophilicity of the fibril in the presence of lycopene, resulting in higher mobility and thus larger disorientation similar to reported for dihydrochalcone [78] and ellagic acid [61] molecule. ...
... However, not much change has been observed for chain B-E for Aβ-LYC1 over Aβ-Water system. Our observations well-match with the reported results based on dihydrochalcone, [78] and flavonoid derivative, [82] ellagic acid, [61] brazilin, [49] and a flavonoid compound [53] as the ligand. The higher value of average distance of inter-chain residues, observed in Aβ-LYC1 system elucidates the mechanistic details behind destabilization of the fibril. ...
... 42−56 Several groups studied the molecular mechanism by which small molecules disrupt Aβ protofibril (including morin, wgx-50, 12-crown-4 ether, dihydrochalcone, resveratrol and clioquinol hybrid compound, caffeine and ditriazole derivative). They found these hydrophobic molecules disrupt Aβ 17−42 protofibril (a pentamer, PDB ID: 2BEG) 42, 45,49,52,53,55 or Aβ 40 protofibril (a trimer, PDB ID: 2M4J) 46 by mostly binding to the protofibril cavity (or the interior of the hydrophobic core). Recently, Tran et al. investigated the binding modes of a glycopeptidomimetic (3β) molecule on Aβ protofibrils (PDB IDs: 2BEG, 2MXU, and 5KK3) by performing molecular docking calculations and MD simulations. ...
... In MD simulation studies, the increase of RMSD value was often used to examine the destabilization effect of inhibitors on amyloid protofibrils. 40,42,45,48,49,52 As shown in Figure 3a and b, during two independent 1 μs MD simulations, in the absence of EGCG/ EGC, the RMSD value of Aβ 42 tetramer is around 0.27 nm, while, in the presence of EGCG/EGC, it increases to ∼0.39/ 0.36 nm. The increase of the RMSD indicates that both EGCG and EGC destabilize the global or local Aβ 42 protofibril structure. ...
... Hydrophobic interactions play important roles between the inhibitors and the residues of SIRT6. In previous computer simulation studies, hydrophobic interactions between small molecules and proteins have also been reported [44][45][46][47][48]. the snapshot, we can see that the small molecule Compound 9 and Scutellarin insert into the long channel pocket. It has been reported that the binding pocket is between the small domain and the larger Rossmann-fold domain [24]. ...
... Hydrophobic interactions play important roles between the inhibitors and the residues of SIRT6. In previous computer simulation studies, hydrophobic interactions between small molecules and proteins have also been reported [44][45][46][47][48]. ...
Sirtuin 6 (SIRT6) is an NAD+-dependent deacetylase with a significant role in 20% of all cancers, such as colon cancers and rectal adenocarcinoma. However, there is currently no effective drug for cancers related to SIRT6. To explore potential inhibitors of SIRT6, it is essential to reveal details of the interaction mechanisms between inhibitors and SIRT6 at the atomic level. The nature of small molecules from herbs have many advantages as inhibitors. Based on the conformational characteristics of the inhibitor Compound 9 (Asinex ID: BAS13555470), we explored the natural molecule Scutellarin, one compound of Huang Qin, which is an effective herb for curing cancer that has been described in the Traditional Chinese Medicine (TCMS) library. We investigated the interactions between SIRT6 and the inhibitors using molecular dynamics (MD) simulations. We illustrated that the structurally similar inhibitors have a similar binding mode to SIRT6 with residues—Leu9, Phe64, Val115, His133 and Trp188. Hydrophobic and π-stacking interactions play important roles in the interactions between SIRT6 and inhibitors. In summary, our results reveal the interactive mechanism of SIRT6 and the inhibitors and we also provide Scutellarin as a new potential inhibitor of SIRT6. Our study provides a new potential way to explore potential inhibitors from TCMS.
... Hydrophobic interactions and π-π interactions play major roles between the inhibitors and the residues of S_RBD. In previous computer simulation studies, hydrophobic interactions and π-π interactions between small molecules and protein have also been reported [57][58][59][60][61]. ...
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a type of Ribonucleic Acid (RNA) coronavirus and it has infected and killed many people around the world. It is reported that the receptor binding domain of the spike protein (S_RBD) of the SARS-CoV-2 virus is responsible for attachment to human angiotensin converting enzyme II (ACE2). Many researchers are attempting to search potential inhibitors for fighting SARS-CoV-2 infection using theoretical or experimental methods. In terms of experimental and theoretical research, Cefuroxime, Erythromycin, Lincomycin and Ofloxacin are the potential inhibitors of SARS-CoV-2. However, the interactive mechanism of the protein SARS-CoV-2 and the inhibitors are still elusive. Here, we investigated the interactions between S_RBD and the inhibitors using molecular dynamics (MD) simulations. Interestingly, we found that there are two binding sites of S_RBD for the four small molecules. In addition, our analysis also illustrated that hydrophobic and π-π stacking interactions play crucial roles in the interactions between S_RBD and the small molecules. In our work, we also found that small molecules with glycosyl group have more effect on the conformation of S_RBD than other inhibitors, and they are also potential inhibitors for the genetic variants of SARS-CoV-2. This study provides in silico-derived mechanistic insights into the interactions of S_RBD and inhibitors, which may provide new clues for fighting SARS-CoV-2 infection.
... 42 Dihydrochalcone, a natural compound from pepper, has shown its ability to destabilize Ab fibrils, studied by MD simulation. 43 Recently, EGCG (epigallocatechin-3-gallate), a tea catechin, was also reported as a b-sheet breaker converting neurotoxic aggregates into unstructured non-toxic clumps. 44 Finding new drug candidates from plants to treat AD by means of computational studies is an exemplary and continuously evolving process. ...
The clinical signature of Alzheimer’s disease (AD) is the deposition of aggregated Aβ fibrils that are neurotoxic to the brain. It is the major form of dementia affecting older people worldwide, impeding their normal sustenance. Finding and testing various natural compounds to target and disrupt stable Aβ fibrils seems to be a promising and attractive therapeutic approach. The four phenolic compounds from plant sources were taken into consideration for the present work, which initially screened by Docking. The ellagic acid (REF) came out to be the best binder of Aβ oligomer from docking studies. Further, to test the destabilization effect of REF on Aβ oligomer, MD simulation was conducted. The simulation outcome obtained henceforth clearly indicates a drift of terminal chains from the Aβ oligomer, leading to the disorganization of the characteristically organized cross β structure of Aβ fibrils. Increased value of RMSD, Rg, RMSF, and SASA are indicative of destabilization of the Aβ fibril in the presence of REF. The disruption of salt bridges and a notable decline in the number of hydrogen bonds and β-sheet content explains the conformational changes in the Aβ fibril structure, ceasing their neurotoxicity. The MM-PBSA results revealed the binding of REF to chain A of the Aβ oligomer. The destabilization potential of ellagic acid, as explained by the MD simulation study, establishes it as a promising drug to cure AD. The molecular level details about the destabilization mechanism of ellagic acid encourage intensive mining of other natural compounds as well for therapeutic intervention for AD.
... 61 At 150 ns, a significant loss in the β-sheet and enhancement in the coil content were observed in chains D and E of the Aβ 42 protofibril in Aβ 42 protofibril−4v complex as compared to the Aβ 42 protofibril alone. The snapshots indicated that 4v disrupts the Aβ 42 protofibril structure by inserting into the hydrophobic core, which is in accordance with the earlier reports on the binding and destabilization of Aβ protofibril structure by small molecules such as wgx-50, 39c dihydrochalcone, 62 and 12-crown-4 ether. 63 3.2.5. ...
Clinical studies have identified that abnormal self-assembly of amyloid-β (Aβ) peptide into toxic fibrillar aggregates is associated with the pathology of Alzheimer’s disease (AD). The most acceptable therapeutic approach to stop the progression of AD is to inhibit the formation of β-sheet-rich structures. Recently, we designed and evaluated a series of novel mono-triazole derivatives 4(a–x), where compound 4v was identified as the most potent inhibitor of Aβ42 aggregation and disaggregates preformed Aβ42 fibrils significantly. Moreover, 4v strongly averts the Cu²⁺-induced Aβ42 aggregation and disaggregates the preformed Cu²⁺-induced Aβ42 fibrils, halts the generation of reactive oxygen species, and shows neuroprotective effects in SH-SY5Y cells. However, the underlying molecular mechanism of inhibition of Aβ42 aggregation by 4v and disaggregation of preformed Aβ42 fibrils remains obscure. In this work, molecular dynamics (MD) simulations have been performed to explore the conformational ensemble of the Aβ42 monomer and a pentameric protofibril structure of Aβ42 in the presence of 4v. The MD simulations highlighted that 4v binds preferentially at the central hydrophobic core region of the Aβ42 monomer and chains D and E of the Aβ42 protofibril. The dictionary of secondary structure of proteins analysis indicated that 4v retards the conformational conversion of the helix-rich structure of the Aβ42 monomer into the aggregation-prone β-sheet conformation. The binding free energy calculated by the molecular mechanics Poisson–Boltzmann surface area method revealed an energetically favorable process with ΔGbinding = −44.9 ± 3.3 kcal/mol for the Aβ42 monomer–4v complex. The free energy landscape analysis highlighted that the Aβ42 monomer–4v complex sampled conformations with significantly higher helical contents (35 and 49%) as compared to the Aβ42 monomer alone (17%). Compound 4v displayed hydrogen bonding with Gly37 (chain E) and π–π interactions with Phe19 (chain D) of the Aβ42 protofibril. Further, the per-residue binding free energy analysis also highlighted that Phe19 (chain D) and Gly37 (chain E) of the Aβ42 protofibril showed the maximum contribution in the binding free energy. The decreased binding affinity and residue–residue contacts between chains D and E of the Aβ42 protofibril in the presence of 4v indicate destabilization of the Aβ42 protofibril structure. Overall, the structural information obtained through MD simulations indicated that 4v stabilizes the native helical conformation of the Aβ42 monomer and persuades a destabilization in the protofibril structure of Aβ42. The results of the study will be useful in the rational design of potent inhibitors against amyloid aggregation.
... These results are in agreement with simulation studies conducted on Ab protofibrils with dihydrochalcone as the ligand. 53 Salt bridges, electrostatic interactions (intra-peptide and inter-peptide), involving D23 and K28 residues are reported to be a major contributor towards the stability of fibril structures and govern the fibrillation rate of full length Ab 17-42 . 78,125-128 A substantial increase in the interchain distance between D23 of chain A and K28 of chain B is observed (see Fig. 7B), resulting in the interruption of salt bridge formation. ...
Aggregation and deposition of neurotoxic Aβ fibrils result in etiology of Alzheimer’s disease (AD). It has been clinically recognized as a major form of dementia across the globe. Finding and testing various natural compounds to target Aβ fibrils to disrupt their stable structure seems to be a promising and attractive therapeutic strategy. The destabilization effect of caffeine on Aβ fibrils is comprehended by in-silico studies wherein a series of molecular dynamics (MD) simulations each of 100 ns were conducted. The simulation outcomes obtained henceforth clearly indicated a drift of terminal chains from the protofibril leading to disorganization of the characteristically organized cross β structure of Aβ fibrils. The structural instability of Aβ17-42 protofibril is explained by enhanced fluctuations in RMSD, the radius of gyration and RMSF values in the presence of caffeine. The key interactions providing stability comprising D23-K28 salt bridges, intra- and inter-chain hydrogen bonding and hydrophobic interactions consisting interchain A21-V36 and F19-G38 and intrachain L34-V36, found disrupted due to an increase in the distance between them. The loss of β-sheet structure with the introduction of turns and α-helix in terminal chains may further inhibit the formation of higher order aggregates, necessary to stop the progression of the disease. The atomistic details by MD studies on the underlying destabilization mechanism of Aβ17-42 protofibril by caffeine encourage further investigation in exploring the potency of natural compounds to treat AD by disrupting preformed neurotoxic Aβ protofibrils.
... Using multiple molecular dynamics (MD) simulations, Jin et al. reported that dihydrochalcone, a compound extracted from the daemonorops draco tree, could effectively inhibit Aβ 1-42 fibrillization and reduce Aβ-induced cytotoxicity by destabilizing the Aβ PFs. In this process, dihydrochalcone was shown to bind to the cavity of the Aβ 1-40 /Aβ 1-42 PFs themselves and disrupt the D23-K28 salt bridge and inter-peptide β-sheet in the β1 region [51]. In addition, Zhou et al. reported that 1,2-(dimethoxymethano)fullerene (DMF), a water-soluble fullerene derivative, strongly inhibited Aβ 1-42 aggregation by binding with Aβ PFs on three dominant binding sites, namely, the central hydrophobic core (17LVFFA21), the turn site (27NKGAI31), and the C-terminal β-sheet site comprised of glycine and hydrophobic residues (31IIGLMVGGVVI41), by MD stimulations [52]. ...
Worldwide, Alzheimer’s disease (AD) is the most common age-related neurodegenerative disease and is characterized by unique pathological hallmarks in the brain, including plaques composed of amyloid β-protein (Aβ) and neurofibrillary tangles of tau protein. Genetic studies, biochemical data, and animal models have suggested that Aβ is responsible for the pathogenesis of AD (i.e., the amyloid hypothesis). Indeed, Aβ molecules tend to aggregate, forming oligomers, protofibrils, and mature fibrils. However, while these Aβ species form amyloid plaques of the type implicated in AD neurodegeneration, recent clinical trials designed to reduce the production of Aβ and/or the plaque burden have not demonstrated clinical efficacy. In addition, recent studies using synthetic Aβ peptides, cell culture models, Arctic transgenic mice, and human samples of AD brain tissues have suggested that the pre-fibrillar forms of Aβ, particularly Aβ protofibrils, may be the most critical species, compared with extracellular fibrillar forms. We recently reported that protofibrils of Aβ1-42 disturbed membrane integrity by inducing reactive oxygen species generation and lipid peroxidation, resulting in decreased membrane fluidity, intracellular calcium dysregulation, depolarization, and synaptic toxicity. Therefore, the therapeutic reduction of protofibrils may prevent the progression of AD by ameliorating neuronal damage and cognitive dysfunction through multiple mechanisms.
... Chakraborty et al. [146] described the multifunctional neuroprotective activity of neohesperidin dihydrochalcone as a potential novel scaffold for therapeutics against neurodegenerative disorders, such like Alzheimer's disease. Jin et al. [147] noted that some natural dihydrochalcones might be good drug candidates to treat Alzheimer's disease. It has been shown that a compound extracted from Daemonorops draco tree inhibited amyloid-β peptide aggregation and stopped production of the toxic fibrillary aggregates, which are responsible for the formation of neurofibrillary tangles, one of the main symptoms of dementia [146]. ...
Dihydrochalcones are a class of secondary metabolites, for which demand in biological and pharmacological applications is still growing. They posses several health-endorsing properties and, therefore, are promising candidates for further research and development. However, low content of dihydrochalcones in plants along with their low solubility and bioavailability restrict the development of these compounds as clinical therapeutics. Therefore, chemomicrobial and enzymatic modifications are required to expand their application. This review aims at analyzing and summarizing the methods of obtaining dihydrochalcones and of presenting their pharmacological actions that have been described in the literature to support potential future development of this group of compounds as novel therapeutic drugs. We have also performed an evaluation of the available literature on beneficial effects of dihydrochalcones with potent antioxidant activity and multifactorial pharmacological effects, including antidiabetic, antitumor, lipometabolism regulating, antioxidant, anti-inflammatory, antibacterial, antiviral, and immunomodulatory ones. In addition, we provide useful information on their properties, sources, and usefulness in medicinal chemistry.
... 44−47 Destabilization of the Aβ protofibril is mainly due to partial insertion of RR-AFC into the hydrophobic core; binding of small molecules (i.e., dihydrochalcone, wgx-50, and 12-crown-4 ether) at the hydrophobic core also destabilized the Aβ protofibril reported by previous studies. 48,49 Binding at this site causes rearrangements of β2 sheets at the C-terminal region of the Aβ protofibril ( Figure 4B,D). Secondary structural analysis from the MD simulation of the blind docked complex (set II-1) and flexible docked complex (set IV-3) using DSSP shows that there is a slight decrease in βsheet percentage corresponding to the increase in coil contents (Table 2 and Figure S13). ...
Aggregation of amyloid beta (Aβ) peptides leads to formation of fibrilar, soluble oligomers, and their deposition is a key event in progression of Alzheimer’s disease (AD). Recent experimental studies of Arg-Arg-7-amino-4-trifluoromethylcoumarin (RR-AFC) showed significant Aβ aggregation inhibition, but its molecular mechanism is not yet clear. Hence, the present study aims at exploring the underlying mechanism of destabilization and inhibition of aggregation of the Aβ protofibril by RRAFC at the molecular level. Molecular docking analysis shows that RR-AFC binds to chain A of the Aβ protofibril through hydrogen bonding interactions. Comparative molecular dynamics simulations depict the binding of RR-AFC at the edge of chain A, and its partially inserted conformation at the hydrophobic core destabilizes the Aβ protofibril. It’s binding causes loss of hydrophobic contacts, leading to a partial opening of tightly packed β-sheet protofibrils. The hydration effect of salt bridge between the amino group of Lys28 and the oxygen atom of RR-AFC contributes in destabilization of Aβ protofibrils. Binding free energy calculations of RR-AFC and the Aβ protofibril showed that van der Waals interactions are dominant over the others. Thus, our results revealed that RR-AFC interacts mainly with the hydrophobic core along with positively charged residues of the Aβ protofibril for effective destabilization. Thus, this structural information could be useful to design new inhibitors to control the aggregation of Aβ protofibrils in AD patients.
... Docking studies using the known ss NMR structures may not provide accurate picture of ligand-protein interactions [56]. However, they are a useful tool to predict potential ligand binding sites of Aβ aggregates [55,57,58]. ...
... In addition, dihydrochalcone also binds to N-terminal residues 4Phe-Arg-His6 and 10Tyr12Val-His-His-Gln15. These results indicate that dihydrochalcone has similar binding sites and a similar destabilization effect on Aβ1-40 protofibrils as on Aβ1-42 [103]. ...
The amyloid-β 1-42 (Aβ1-42) peptide is produced by proteolytic cleavage of the amyloid precursor protein (APP) by sequential reactions that are catalyzed by γ and β secretases. Aβ1-42, together with the Tau protein are two principal hallmarks of Alzheimer’s disease (AD) that are related to disease genesis and progression. Aβ1-42 possesses a higher aggregation propensity, and it is able to form fibrils via nucleated fibril formation. To date, there are compounds available that prevent Aβ1-42 aggregation, but none have been successful in clinical trials, possibly because the Aβ1-42 structure and aggregation mechanisms are not thoroughly understood. New molecules have been designed, employing knowledge of the Aβ1-42 structure and are based on preventing or breaking the ionic interactions that have been proposed for formation of the Aβ1-42 fibril U-shaped structure. Recently, a new Aβ1-42 fibril S-shaped structure was reported that, together with its aggregation and catalytic properties, could be helpful in the design of new inhibitor molecules. Therefore, in silico and in vitro methods have been employed to analyze the Aβ1-42 fibril S-shaped structure and its aggregation to obtain more accurate Aβ1-42 oligomerization data for the design and evaluation of new molecules that can prevent the fibrillation process.
i>Sanguis Draconis , the prepared resin of the fruit of Daemonorops draco Bl., is a precious traditional medicine from Southeast Asia. It has the effects of activating blood circulation and calming pain, removing blood stasis and hemostasis, generating muscle and astringeting sores. It is used for traumatic injury, abdominal stasis and pain, traumatic bleeding, etc. The chemical composition of Sanguis Draconis is complex, mainly including flavonoids, diterpenoid acids and other effective components. Modern pharmacological studies show that Sanguis Draconis has excellent pharmacological activities such as anti-inflammatory, antibacterial, anti-oxidant, anti-tumor, promoting wound healing and so on. This paper comprehensively arranges and summarizes the research status of the variety sources, chemical components and pharmacological effects of Sanguis Draconis , so as to provide scientific basis and new ideas for future research of Sanguis Draconis .
Chronic traumatic encephalopathy (CTE), a unique tauopathy,
is pathologically associated with the aggregation of hyperphosphorylated tauprotein into fibrillar aggregates. Inhibiting tau aggregation and disaggregating
tau protofibril might be promising strategies to prevent or delay the
development of CTE. Newly resolved tau fibril structures from deceased
CTE patients’ brains show that the R3-R4 fragment of tau forms the core of the
fibrils and the structures are distinct from other tauopathies. An in vitro
experiment finds that epigallocatechin gallate (EGCG) can effectively inhibit
human full-length tau aggregation and disaggregate preformed fibrils. However,
its inhibitive and destructive effects on the CTE-related R3-R4 tau and the
underlying molecular mechanisms remain elusive. In this study, we performed
extensive all-atom molecular dynamics simulations on the CTE-related R3-R4
tau dimer/protofibril with and without EGCG. The results reveal that EGCG
could reduce the β-sheet structure content of the dimer, induce the dimer to form loosely packed conformations, and impede the
interchain interactions, thus inhibiting the further aggregation of the two peptide chains. Besides, EGCG could reduce the structural
stability, decrease the β-sheet structure content, reduce the structural compactness, and weaken local residue−residue contacts of the
protofibril, hence making the protofibril disaggregated. We also identified the dominant binding sites and pivotal interactions. EGCG
preferentially binds with hydrophobic, aromatic, and positively/negatively charged residues of the dimer, while it tends to bind with
polar, hydrophobic, aromatic, and positively charged residues of the protofibril. Hydrophobic, hydrogen-bonding, π−π stacking, and
cation−π interactions synergistically drive the binding of EGCG on both the dimer and the protofibril, but anion−π interaction only
exists in the interaction of EGCG with the dimer. Our work unravels EGCG’s inhibitive and destructive effects on the CTE-related
R3-R4 tau dimer/protofibril and the underlying molecular mechanisms, which provides useful implications for the design of drugs to
prevent or delay the progression of CTE.
Valerolactone as a renewable, polar aprotic solvent was evaluated in the selective hydrogenation of carbon‐carbon double bond of chalcones from both catalytic and mechanistic aspects. By using triphenylphosphine‐modified cationic rhodium catalyst systems reasonably high activities (TOFinitial∼1200 h⁻¹) were achieved in the selective hydrogenation of (E)‐chalcone (CHL), as a common model substrate for α,β‐unsaturated ketones, to dihydrochalcone (DHC). Various derivatives of CHL, which were substituted on the styryl phenyl groups also showed exclusive selectivity for the hydrogenation of the C=C double bond. The optimal PPh3/Rh ratio was between 2.5 to 3, but higher ratios were advantageously also tolerated, which is beneficial for the process chemistry. The solvent GVL proved to stable enough under the reaction conditions for waste‐free recycling. Kinetic studies and NMR spectra from the analogous deuteration of chalcone by using the cationic Rh‐PPh3 system clearly show the olefin insertion as the rate‐determining step with definite irreversibility. By investigating the substituent effect on the phenyl ring of the styryl moiety of chalcone, a quasi‐linear correlation was found between the electronic parameters of the para‐substituents and the activity with electron donor groups being favorable for the reaction rate. In summary, an environmentally friendly and feasible method is presented for the production of valuable DHC intermediates in view of their pharmaceutical importance.
Scientific review of articles based on Indonesian local wisdom has been carried out, especially on the potential of 11 types of rattan in Suku Anak Dalam (SAD) jambi province, Indonesia. Scientific studies are carried out with reference to the same type of species, related to the study of their potential activity pharmacologically and toxicity studies on values LD 50. The article review study was conducted using the PRISMA method. Retrieval of article sources from science direct, springer and pubmed databases is performed. Limitations are carried out related to data collection, that only take original research articles and review articles with article sources in English. Interesting data have been obtained here, no study of the pharmacological effect on 11 types of rattan has been carried out in this article review study. Rattan by species Daemonorops draco, Calamus scipionum, Calamus manan, Calamus ornatus, Calamus axillaris and Calamus caesius studies of pharmacological effects have been carried out. Only on Daemonorops draco obtained data that is quite capable and gets attention by world researchers because it is known to have the potential for anti-cancer activities. However, in other types of rattan, the data obtained is not so much and it is still very feasible to conduct a study search on drug discovery based on natural product sources as drug candidates. Rattan by species Calamus fabellatus, Calamus javensis, Korthalsia echinometra and Daemonorops geniculate no scientific studies have been found related to its potential activities pharmacologically. In the study of acute toxicity LD 50 , has not been done at all on the 11 types of rattan. In conclusion, the "Gap of Science" of the topic of rattan available on the environment Suku Anak Dalam (SAD) Bukit Dua Belas Jambi Province still has great potential for scientific and in-depth studies related to the potential and study of drug discovery by utilizing natural product sources that can be done by all researchers with an interest in natural products as a source of treatment.
Alzheimer's disease (AD) is related to the misfolding and aggregation of amyloid-β (Aβ) protein, and its major pathological hallmark is fibrillary β-amyloid plaques. Impeding the formation of Aβ β-structure-rich aggregates and dissociating Aβ fibrils are considered potent strategies to suppress the onset and progression of AD. As a molecular chaperone, human αB-crystallin has received extensive attention in the inhibition of protein aggregation. Previous experiments reported that the structured core region of αB-crystallin (αBC) exhibits a better preventive effect on Aβ aggregation and toxicity than the full-length protein. However, the molecular mechanism behind the effect of inhibition remains mostly unknown. Herein, we carried out six 500 ns molecular dynamics (MD) simulations to investigate the inhibitory mechanism of αBC on Aβ42 aggregation. Our simulations show that αBC greatly impedes the formation of β-structure contents. We find that the binding of αBC to the Aβ42 monomer is driven by polar, hydrophobic, and H-bonding interactions. To explore whether αBC could destabilize Aβ42 protofibrils, we also carried out MD simulations of Aβ42 protofibrils with and without αBC. The results show that αBC interacts with three binding sites of the Aβ42 protofibril, and the binding is mainly driven by polar and H-bonding interactions. The binding of αBC at these three sites has a preferred dissociation effect on the β-structure content, kink angle, and K28-A42 salt bridges. Overall, this study not only discloses the molecular mechanism of αBC against Aβ42 aggregation but also demonstrates the disruption effects of αBC on Aβ42 protofibrils, which yields an avenue for designing anti-AD drug candidates.
Biflavones are a kind of natural compound with a variety of biological activities, which have the capability of reversing diabetes and neurodegenerative diseases. The human islet amyloid polypeptide (hIAPP) is closely related to the pathological process of type II diabetes mellitus (T2DM). The development of new inhibitors is crucial to prevent hIAPP aggregation against T2DM. However, the influences of biflavones on hIAPP aggregation are unknown. In this work, we utilized a series of biophysical and biochemical techniques to seek the inhibitory effects of two biflavones on hIAPP fibril formation and their interaction mechanism. The biflavones namely amentoflavone (1), and bilobetin (2), distinctly prevented the self-assembly behavior of hIAPP, and depolymerized the aged aggregates to small oligomers and monomers. In addition, the two compounds displayed strong binding affinity to hIAPP mainly through hydrophobic and hydrogen bonding interactions, and the hydroxyl substitution in 1 was superior to the methoxy substitution in 2 at the same C8 position in impeding hIAPP aggregation. 1 and 2 also decreased hIAPP-induced cytotoxicity by reducing peptide oligomerization. This work offers useful data for understanding the roles of biflavones in hIAPP fibrillation and for the treatment of T2DM and other amyloidosis related diseases.
Functional amyloids are abundant in living organisms from prokaryotes to eukaryotes playing diverse biological functions. In contrast to the irreversible aggregation of most known pathological amyloids, we postulate that naturally-occurring functional amyloids are reversible under the evolutionary pressure to be able to module the fibrillization process, reuse the composite peptides, or perform their biological functions. β-endorphin, an endogenous opioid peptide hormone, forms such kind of reversible amyloid fibrils in secretory granules for efficient storage and returns to the functional state of monomers upon release into the blood. The environment change between the low pH in secretory granules and the neutral pH in extracellular spaces is believed to drive the reversible fibrillization of β-endorphin. Here, we investigate the critical role of a buried glutamate, Glu8, in the pH-responsive disassembly of β-endorphin fibrils using all-atom molecular dynamics simulations along with structure-based pKa prediction. The fibril was stable at pH 5.5 or lower with all the buried Glu8 residues protonated and neutrally charged. After switching to the neutral pH, Glu8 residues of peptides at the outer layers of the ordered fibrils became deprotonated due to partial solvent exposure, causing sheet-to-coil conformational changes and subsequent exposure of adjacent Glu8 residues in the inner chains. Via iterative deprotonation of Glu8 and induced structural disruption, all Glu8 residues would be progressively deprotonated. Electrostatic repulsion between deprotonated Glu8 residues along with their high solvation tendency disrupted the hydrogen bonding between the β1 strands and increased the solvent exposure of those otherwise buried residues in the cross-β core. Overall, our computational study reveals that strategic positioning of ionizable residues into the cross-β core is a potential approach to design reversible amyloid fibrils as pH-responsive smart bio-nanomaterials.
The aggregation of amyloid-β protein (Aβ) into fibrillary deposits is implicated in Alzheimer's disease (AD), and inhibiting Aβ aggregation and clearing Aβ fibrils are considered as promising strategies to treat...
Stimulator of interferon genes (STING), which is an integral ER-membrane protein, could induce an antiviral state and boost antitumor immunity. Recent experiments reported that different small molecules could modulate the conformation of the STING. However, the mechanism of small molecules modulating the conformation of STING is still unknown. To illustrate the conformational modulated mechanism of STING by small molecules at atomic level, we investigated the interactions between STING and the small molecules: cGAMP and diABZI with molecular dynamics (MD) simulations method. Interestingly, we found that the residues of STING in the binding pocket are more flexible in the monomers of STING than that in the dimer of STING. We also demonstrated that cGAMP and diABZI have a similar binding mode to STING monomers/dimer, and π–π stacking interactions play important roles for the agonists and STING. Our study proposed mechanistic insights into the STING conformation modulated by small molecules and we suggested that the special molecule (e. g. diABZI) could induce the conformational transition of STING from the “open” monomers to the “closed” dimer state. Our research may provide a clue for the development of cancer immunotherapy.Graphic abstract
Here, we have studied the ameliorative effects of Withania somnifera derivatives (Withanolide A, Withanolide B, Withanoside IV, and Withanoside V) on the fibril formation of amyloid-β 42 for Alzheimer's disease. We analyzed reduction in the aggregation of β amyloid protein with these Ashwagandha derivatives by Thioflavin T assay in the oligomeric and fibrillar state. We have tested the cytotoxic activity of these compounds against human SK-N-SH cell line for 48 h, and the IC 50 value found to be 28.61 ± 2.91, 14.84 ± 1.45, 18.76 ± 0.76 and 30.14 ± 2.59 μM respectively. After the treatment of the cells with half the concentration of IC 50 value, there was a remarkable decrease in the number of apoptotic cells stained by TUNEL assay indicating the DNA damage and also observed significant decrease of reactive oxygen species. Also, the binding and molecular stability of these derivatives with amyloid β was also studied using bioinformatics tools where these molecules were interacted at LVFFA region which is inhibition site of amyloid-β 42. These studies revealed that the Withanolides and Withanosides interact with the hydrophobic core of amyloid-β 42 in the oligomeric stage, preventing further interaction with the monomers and diminishing aggregation.
The accumulation and deposition of fibrillar aggregates of α-synuclein (α-syn) into Lewy bodies are the major hallmarks of Parkinson's disease (PD) for which there is no cure yet. Disrupting preformed α-syn fibrils is considered one of the rational therapeutic strategies to combat PD. Experimental studies reported that epigallocatechin gallate (EGCG), a polyphenol extracted from green tea, can disrupt α-syn fibrils into benign amorphous aggregates. However, the molecular mechanism of action is poorly understood. Herein, we performed molecular dynamics simulations on a newly released Greek-key-like α-syn fibril with or without EGCG to investigate the influence of EGCG on α-syn fibril. Our simulations show that EGCG disrupts the local β-sheet structure, E46-K80 salt-bridge crucial for the stabilization of the Greek-key-like structure, and hydrophobic interactions stabilizing the inter-protofibril interface and destabilizes the global structure of the α-syn fibril. Interaction analyses reveal that hydrophobic and hydrogen-bonding interactions between EGCG and α-syn fibrils play important roles in the destabilization of the fibril. We find that the disruption of the E46-K80 salt-bridge closely correlates with the formation of hydrogen-bonds (H-bonds) between EGCG and E46/K80. Our results provide mechanistic insights into the disruption modes of α-syn fibril by EGCG, which may pave the way for designing drug candidates targeting α-syn fibrillization to treat PD.
Alzheimer’s disease (AD) is a neurological disorder, growing epidemic across worldwide due to no effective medical aid available in the market. AD is known to be directly associated with toxicity...
Alzheimer's Disease (AD) is a devastating neurodegenerative disease affects millions of people in the world. The abnormal aggregation of amyloid b protein (Aβ) is regarded as the key event in AD onset. Meanwhile, the Aβ oligomers are believed to be the most toxic species of Aβ. Recent studies show that the Aβ dimers, which are the smallest form of Aβ oligomers, also have the neurotoxicity in the absence of other oligomers in physiological conditions. In this review we focus on the pathogenesis, structure and potential therapeutic molecules against small Aβ oligomers, as well as the nano-particles (NPs) in the treatment of AD. In this review, we firstly focus on the pathogenic mechanism of Aβ oligomers, especially the Aβ dimers. The toxicity of Aβ dimer or oligomers which attributes to the interactions with various receptors and the disruption of membrane or intracellular environments were introduced. Then the structure properties of Aβ dimers and oligomers are summarized. Although some structural information such as the secondary structure content are characterized by experimental technologies, the full detailed structures are still absent. Following that, the small molecules targeting Aβ dimers or oligomers are collected, nevertheless, all of these ligands are failed to come into the market, rising controversy of the Aβ-related "amyloid cascade hypothesis". At last, the recent progresses about the nano-particles as the potential drugs or the drug delivery to the Aβ oligomers are present.
Variational autoencoders are artificial neural networks with the capability to reduce highly dimensional sets of data to smaller dimensional, latent representations. In this work, these models are applied to molecular dynamics simulations of the self-assembly of coarse-grained peptides to obtain a singled-valued order parameter for amyloid aggregation. This automatically learned order parameter is constructed by time-averaging the latent parameterizations of internal coordinate representations and compared to the nematic order parameter which is commonly used to study ordering of similar systems in literature. It is found that the latent space value provides more tailored insight into the aggregation mechanism’s details, correctly identifying fibril formation in instances where the nematic order parameter fails to do so. A means is provided by which the latent space value can be analyzed so that the major contributing internal coordinates are identified, allowing for a direct interpretation of the latent space order parameter in terms of the behavior of the system. The latent model is found to be an effective and convenient way of representing the data from the dynamic ensemble and provides a means of reducing the dimensionality of a system whose scale exceeds molecular systems so-far considered with similar tools. This bypasses a need for researcher speculation on what elements of a system best contribute to summarizing major transitions, and suggests latent models are effective and insightful when applied to large systems with a diversity of complex behavior.
Previous studies have indicated that 5-hydroxycyclopenicillone (HCP), an active compound derived from marine sponge, could inhibit oligomerization of amyloid β-protein (Aβ). However, the molecular basis for the interaction between HCP and Aβ remains unclear. Herein, all-atom molecular dynamics (MD) simulations were used to explore the conformational conversion of an Aβ40 monomer at different concentrations (0-40 mM) of HCP at the atomic level. It is confirmed that the conformational transition of the Aβ40 monomer is prevented by HCP in a concentration-dependent manner in silico. In 40 mM HCP solution, the initial α-helix-rich conformation of Aβ40 monomer is kept under the action of HCP. The intra-peptide hydrophobic collapse and D23-K28 salt bridge are prevented by HCP. Moreover, it is indicated that the non-polar binding energy dominates the binding between HCP and Aβ40 monomer as evaluated by molecular mechanics Poisson-Boltzmann surface area method. And, the residues of F4, Y10, V12, L17 and L34 in Aβ40 might contribute to the binding energy in HCP-Aβ40 complex. All these results elucidate the molecular mechanism underlying the inhibitory effects of HCP against the conformational transformation of Aβ40, providing a support that HCP may be developed as a potential anti-Aβ compound for the treatment of Aβ-related diseases.
Communicated by Ramaswamy H. Sarma
Conformational transition of proteins and peptides into highly stable, β-sheet rich structures is observed in many amyloid-associated neurodegenerative disorders, yet the precise mechanism of amyloid formation at the molecular level remains poorly understood due to the complex molecular structures. Short peptides provide simplified models for studying the molecular basis of the assembly mechanism in that governs β-sheet fibrillation processes underlying the formation and inhibition of amyloid-like structures. Herein, we report a supramolecular co-assembly strategy for the inhibition and transformation of stable β-sheet-rich amyloid-derived dipeptide self-assemblies into adaptable secondary structural fibrillar assemblies by mixing with bipyridine derivatives. The interplay between the type and mixing ratio of bipyridine derivatives allowed the variable co-assembly process with stimuli-responsive functional properties, studied by various experimental characterizations and computational methods. Furthermore, the resulting co-assemblies showed functional redox- and photo-responsive properties, making them promising candidates for controllable drug release and fluorescent imprint. This work presents a co-assembly strategy not only to explore the mechanism of amyloid-like structure formation and inhibition at the molecular level, but also to manipulate amyloid-like structures into responsive supramolecular co-assemblies for material science and biotechnology applications.
Extracellular plaques, the hallmark of Alzheimer’s disease brains, consist of insoluble amyloid fibrils that result from the aggregation of amyloid beta peptides. None of the few therapeutic options currently adopted, address the cause of the disease. Instead, they reduce symptom of the disease. Inhibition of aggregation or destabilization of aggregates therefore, emerges as a preferable therapeutic approach. Designing inhibitors or destabilizers demands comprehensive knowledge of the residues of amyloid beta responsible for the phenomenal structural stability of the aggregate. For the purpose, we have compared the effect on structural destabilization of 13 in silico mutations (single and double) with the wild type counterpart of beta-strand-turn-beta-strand motif of the amyloid beta protofibrils by molecular dynamics simulation. Besides the already known salt bridge interaction between K28 and D23, our analyses expose more significant role of K28 as the only positive charge present in the vicinity. Amongst the two consecutive aromatic residues, F19 is involved in stacking interaction; although effect of F20 mutation is more pronounced. Face to face arrangement of A21 and V36 acts as a pillar maintaining the necessary optimum distance between consecutive chains to promote stabilizing interactions. In addition to providing stability to the first beta-strand, large sized negatively charged E22 facilitates salt bridge formation by ensuring fixed relative position of D23 and in turn K28. Likewise, the hydrophobic residues I32 and L34 pack the protofibril core, once again fostering salt bridge interaction. Prospectively, these findings may be compiled for efficient identification or design of scaffolds accountable for protofibril destabilization.
Communicated by Ramaswamy H. Sarma
Amyloid aggregation modulators offer a promising treatment strategy for Alzheimer's disease (AD). We have recently reported a novel di-triazole based compound 6n as a multi-target-directed ligand (MTDL) against AD. 6n effectively inhibits Aβ42 aggregation, metal-induced Aβ42 aggregation, reactive oxygen species (ROS) generation, and rescues SH-SY5Y cells from Aβ42 induced neurotoxicity. However, the underlying inhibitory mechanism remains uncovered. In this regard, molecular dynamics (MD) simulations were performed to understand the effect of 6n on the structure and stability of monomeric Aβ42 and a pentameric protofibril structure of Aβ42. Compound 6n binds preferably to the central hydrophobic core (CHC) and C-terminal regions of the Aβ42 monomer as well as the protofibril structure. The secondary structure analysis suggests that 6n prevents the aggregation of the Aβ42 monomer and disaggregates Aβ42 protofibrils by sustaining the helical content in the Aβ42 monomer and converting the β-sheet into random coil conformation in the Aβ42 protofibril structure. A significant decrease in the average number of hydrogen bonds, binding affinity, and residue-residue contacts between chains D-E of the Aβ42 protofibril in the presence of 6n indicates destabilization of the Aβ42 protofibril structure. The MM-PBSA (molecular mechanics Poisson-Boltzmann surface area) analysis highlighted favourable binding free energy (ΔGbinding) for the Aβ42 monomer-6n and Aβ42 protofibril-6n complex. Overall, MD results highlighted that 6n stabilizes the native α-helix conformation of the Aβ42 monomer and induces a sizable destabilization in the Aβ42 protofibril structure. This work provides theoretical insights into the inhibitory mechanism of 6n against amyloid aggregation and will be beneficial as a molecular guide for structure-based drug design against AD.
Amyloid beta (Aβ) peptide aggregation is considered as one of the key hallmarks of Alzheimer’s disease (AD). Moreover, Aβ peptide aggregation increases considerably in the presence of metal ions and triggers the generation of reactive oxygen species (ROS), which ultimately leads to oxidative stress and neuronal damage. Based on the ‘multi–target–directed ligands’ (MTDLs) strategy, we designed, synthesized and evaluated a novel series of triazole based compounds for AD treatment via experimental and computational methods. Among the designed MTDLs [4(a-x)], the triazole derivative 4v exhibited most potent inhibition of self induced Aβ42 aggregation (78.02%) with an IC50 value of 4.578 ± 0.109 μM and also disassembled the preformed Aβ42 aggregates significantly. In addition, compound 4v showed excellent metal chelating ability and maintained copper in the redox–dormant state to prevent the generation of ROS in copper−ascorbate redox cycling. Further, 4v significantly inhibited Cu2+–induced Aβ42 aggregation and disassembled the Cu2+–induced Aβ42 protofibrils as compared to the reference compound clioquinol (CQ). Importantly, 4v did not show cytotoxicity and was able to inhibit the toxicity induced by Aβ42 aggregates in SH–SY5Y cells. Molecular docking results confirmed the strong binding of 4v with Aβ42 monomer and Aβ42 protofibril structure. The experimental and molecular docking results highlighted that 4v is a promising multifunctional lead compound for AD.
Peptides and proteins have been found to possess an inherent tendency to convert from their native functional states into intractable amyloid aggregates. This phenomenon is associated with a range of increasingly common human disorders, including Alzheimer and Parkinson diseases, type II diabetes, and a number of systemic amyloidoses. In this review, we describe this field of science with particular reference to the advances that have been made over the last decade in our understanding of its fundamental nature and consequences. We list the proteins that are known to be deposited as amyloid or other type of aggregates in human tissues and the disorders with which they are associated, as well as the proteins that exploit the amyloid motif to play specific functional roles in humans. In addition, we summarize the genetic factors that have provided insight into the mechanisms of disease onset. We describe recent advances in our knowledge of the structures of amyloid fibrils and their oligomeric precursors and of the mechanisms by which they are formed and proliferate to generate cellular dysfunction. We show evidence that a complex proteostasis network actively combats protein aggregation and that such an efficient system can fail in some circumstances and give rise to disease. Finally, we anticipate the development of novel therapeutic strategies with which to prevent or treat these highly debilitating and currently incurable conditions. Expected final online publication date for the Annual Review of Biochemistry Volume 86 is June 20, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
Alzheimer’s disease (AD) is the most common dementia affecting tens of million people worldwide. The primary neuropathological hallmark in AD is amyloid plaques composed of amyloid-β peptide (Aβ). Several familial mutations found in Aβ sequence result in early onset of AD. Previous studies showed that the mutations located at N-terminus of Aβ, such as the English (H6R) and Tottori (D7N) mutations, promote fibril formation and increase cytotoxicity. However, A2T mutant located at the very N-terminus of Aβ shows low-prevalence incidence of AD, whereas, another mutant A2V causes early onset of AD. To understand the molecular mechanism of the distinct effect and develop new potential therapeutic strategy, here, we examined the effect of full-length and N-terminal A2V/T variants to wild type (WT) Aβ40 by fibrillization assays and NMR studies. We found that full-length and N-terminal A2V accelerated WT fibrillization and induced large chemical shifts on the N-terminus of WT Aβ, whereas, full-length and N-terminal A2T retarded the fibrillization. We further examined the inhibition effect of various N-terminal fragments (NTFs) of A2T to WT Aβ. The A2T NTFs ranging from residue 1 to residue 7 to 10, but not 1 to 6 or shorter, are capable to retard WT Aβ fibrillization and rescue cytotoxicity. The results suggest that in the presence of full-length or specific N-terminal A2T can retard Aβ aggregation and the A2T NTFs can mitigate its toxicity. Our results provide a novel targeting site for future therapeutic development of AD.
Significance
The absence of fully reproducible protein aggregation assays has contributed to the systematic failures in clinical trials for Alzheimer’s disease (AD) of compounds targeting the aggregation process of the amyloid-β peptide (Aβ). To address this problem, we report the identification of a library of compounds against Aβ aggregation using a drug discovery strategy based on highly quantitative aggregation rate measurements. We then demonstrate, both in Caenorhabditis elegans and human cerebrospinal fluid, that this approach can systematically provide a rich variety of related small molecules to take forward into a drug discovery process. We therefore report an approach that should substantially help overcome the very high level of attrition associated with drug discovery programs for AD.
Significance
Alzheimer’s disease is the most prevalent neurodegenerative disease still with no known cure. The disease is characterized by the development of extracellular plaques and intracellular neurofibrillary tangles. The senile plaques consist mainly of the peptide amyloid-β (Aβ) in aggregated form, called amyloid fibrils. It is believed that the Aβ amyloid fibrils play an important role in disease progression and cell-to-cell transmissibility, and small Aβ oligomers are often assumed to be the most neurotoxic species. Here, we determined the 3D structure of a disease-relevant Aβ(1–42) fibril polymorph combining data from solid-state NMR spectroscopy and mass-per-length measurements from EM. The 3D structure is composed of two molecules per fibril layer, forming a double-horseshoe–like cross–β-sheet entity with maximally buried hydrophobic side chains.
Amyloid-β (Aβ) is a 39-42 residue protein produced by the cleavage of the amyloid precursor protein (APP), which subsequently aggregates to form cross-β amyloid fibrils that are a hallmark of Alzheimer’s disease (AD). The most prominent forms of Aβ are Aβ1-40 and Aβ1-42, that differ by two amino acids (I and A) at the C terminus. However, Aβ42 is more neurotoxic and essential to the etiology of AD. Here we present an atomic resolution structure of a monomorphic form of AβM01-42 amyloid fibrils derived from over 500 13C-13C, 13C-15N distance and backbone angle structural constraints obtained from high field magic angle spinning NMR spectra. The structure (PDB ID: 5KK3) shows that the fibril core consists of a dimer of Aβ42 molecules, each containing four β-strands in a S-shaped amyloid fold, and arranged in a manner that generates two hydrophobic cores that are capped at the end of the chain by a salt bridge. The outer surface of the monomers presents hydrophilic sidechains to the solvent. The interface between the monomers of the dimer shows clear contacts between M35 of one molecule and L17 and Q15 of the second. Intermolecular 13C-15N constraints demonstrate that the amyloid fibrils are parallel in register. The RMSD of the backbone structure (Q15-A42) is 0.71±0.12 Å and of all heavy atoms is 1.07±0.08 Å. The structure provides a point of departure for the design of drugs that bind to the fibril surface and therefore interfere with secondary nucleation and for other therapeutic approaches to mitigate Aβ42 fibril formation.
Seven dihydrochalcone compounds were isolated from the leaves of Malus crabapples, cv. "Radiant", and their chemical structures were elucidated by UV, IR, ESI-MS, ¹H-NMR and (13)C-NMR analyses. These compounds, which include trilobatin (A1), phloretin (A2), 3-hydroxyphloretin (A3), phloretin rutinoside (A4), phlorizin (A5), 6''-O-coumaroyl-4'-O-glucopyranosylphloretin (A6), and 3'''-methoxy-6''-O-feruloy-4'-O-glucopyranosyl-phloretin (A7), all belong to the phloretin class and its derivatives. Compounds A6 and A7 are two new rare dihydrochalcone compounds. The results of a MTT cancer cell growth inhibition assay demonstrated that phloretin and these derivatives showed significant positive anticancer activities against several human cancer cell lines, including the A549 human lung cancer cell line, Bel 7402 liver cancer cell line, HepG2 human ileocecal cancer cell line, and HT-29 human colon cancer cell line. A7 had significant effects on all cancer cell lines, suggesting potential applications for phloretin and its derivatives. Adding a methoxyl group to phloretin dramatically increases phloretin's anticancer activity.
Missense mutations in alanine-673 of the amyloid precursor protein (APP) which corresponds to the second alanine of the Aβ sequence have dramatic impact on risk for Alzheimer disease: Ala2Val (A2V) is causative, while Ala2Thr (A2T) is protective. Assuming a crucial role of amyloid-Aβ in neurodegeneration, we hypothesized that both A2V and A2T mutations cause distinct changes in Aβ properties that may at least partially explain these completely different phenotypes. Using human APP-overexpressing primary neurons, we observed significantly decreased Aβ production in the A2T mutant along with an enhanced Aβ generation in the A2V mutant confirming earlier data from non-neuronal cell lines. More importantly, Thioflavin T fluorescence assays revealed that the mutations, while having little effect on Aβ42 peptide aggregation, dramatically change the properties of the Aβ40 pool with A2V accelerating and A2T delaying aggregation of the Aβ peptides. In line with the kinetic data, Aβ A2T demonstrated an increase in the solubility at equilibrium, an effect that was also observed in all mixtures of A2T mutant with the wild type Aβ40. We propose that in addition to the reduced β-secretase cleavage of APP, the impaired propensity to aggregate may be part of the protective effect conferred by A2T substitution. The interpretation of the protective effect of this mutation is thus much more complicated than proposed previously.
By the virtual screening method we have screened out Dihydrochalcone as a top-lead for the Alzheimer's disease using the database of about 32364 natural compounds. The binding affinity of this ligand to amyloid beta (A[Formula: see text]) fibril has been thoroughly studied by computer simulation and experiment. Using the Thioflavin T (ThT) assay we have obtained the inhibition constant IC50 [Formula: see text]M. This result is in good agreement with the estimation of the binding free energy obtained by the molecular mechanic-Poisson Boltzmann surface area method and all-atom simulation with the force field CHARMM 27 and water model TIP3P. Cell viability assays indicated that Dihydrochalcone could effectively reduce the cytotoxicity induced by A[Formula: see text]. Thus, both in silico and in vitro studies show that Dihydrochalcone is a potential drug for the Alzheimers disease.
The generation of toxic oligomers during the aggregation of the amyloid-β (Aβ) peptide Aβ42 into amyloid fibrils and plaques has emerged as a central feature of the onset and progression of Alzheimer's disease, but the molecular pathways that control pathological aggregation have proved challenging to identify. Here, we use a combination of kinetic studies, selective radiolabeling experiments, and cell viability assays to detect directly the rates of formation of both fibrils and oligomers and the resulting cytotoxic effects. Our results show that once a small but critical concentration of amyloid fibrils has accumulated, the toxic oligomeric species are predominantly formed from monomeric peptide molecules through a fibril-catalyzed secondary nucleation reaction, rather than through a classical mechanism of homogeneous primary nucleation. This catalytic mechanism couples together the growth of insoluble amyloid fibrils and the generation of diffusible oligomeric aggregates that are implicated as neurotoxic agents in Alzheimer's disease. These results reveal that the aggregation of Aβ42 is promoted by a positive feedback loop that originates from the interactions between the monomeric and fibrillar forms of this peptide. Our findings bring together the main molecular species implicated in the Aβ aggregation cascade and suggest that perturbation of the secondary nucleation pathway identified in this study could be an effective strategy to control the proliferation of neurotoxic Aβ42 oligomers.
Congo red (CR) is a commonly used histological amyloid dye and a weak amyloid inhibitor. There is currently no experimentally available structure of CR bound to an amyloid fibril and the binding modes, and the mechanisms governing its inhibitory and optical properties are poorly understood. In this work, we present the first, to our knowledge, atomistically detailed picture of CR binding to protofibrils of the Alzheimer Aβ(9-40) peptide. We identify three major binding modes, with the primary mode residing in the grooves formed by the β-sheets, and observe a restriction of the torsional rotation of the CR molecule upon binding. Our simulations reveal a novel, to our knowledge, electrostatic steering mechanism that plays an important role in the initial recognition and binding of CR to the positively charged surface residues of the fibril. Our simulations provide new, to our knowledge, insights into the striking spectrophotometric and inhibitory properties of CR. In particular, we show that birefringence upon CR binding is due to the anisotropic orientation of the CR dipoles resulting from the spatial ordering of these molecules in the grooves along the fibril axis. The fluorescent enhancement of the bound CR, in turn, is associated with the torsional restriction of this molecule upon binding.
A description of the ab initio quantum chemistry package GAMESS is presented. Chemical systems containing atoms through radon can be treated with wave functions ranging from the simplest closed-shell case up to a general MCSCF case, permitting calculations at the necessary level of sophistication. Emphasis is given to novel features of the program. The parallelization strategy used in the RHF, ROHF, UHF, and GVB sections of the program is described, and detailed speecup results are given. Parallel calculations can be run on ordinary workstations as well as dedicated parallel machines. © John Wiley & Sons, Inc.
Classical Monte Carlo simulations have been carried out for liquid water in the NPT ensemble at 25 °C and 1 atm using six of the simpler intermolecular potential functions for the water dimer: Bernal–Fowler (BF), SPC, ST2, TIPS2, TIP3P, and TIP4P. Comparisons are made with experimental thermodynamic and structural data including the recent neutron diffraction results of Thiessen and Narten. The computed densities and potential energies are in reasonable accord with experiment except for the original BF model, which yields an 18% overestimate of the density and poor structural results. The TIPS2 and TIP4P potentials yield oxygen–oxygen partial structure functions in good agreement with the neutron diffraction results. The accord with the experimental OH and HH partial structure functions is poorer; however, the computed results for these functions are similar for all the potential functions. Consequently, the discrepancy may be due to the correction terms needed in processing the neutron data or to an effect uniformly neglected in the computations. Comparisons are also made for self‐diffusion coefficients obtained from molecular dynamics simulations. Overall, the SPC, ST2, TIPS2, and TIP4P models give reasonable structural and thermodynamic descriptions of liquid water and they should be useful in simulations of aqueous solutions. The simplicity of the SPC, TIPS2, and TIP4P functions is also attractive from a computational standpoint.
The amyloid-β lowering capacity of anti-Aβ antibodies has been demonstrated in transgenic models of Alzheimer's disease (AD) and in AD patients. While the mechanism of immunotherapeutic amyloid-β removal is controversial, antibody-mediated sequestration of peripheral Aβ versus microglial phagocytic activity and disassembly of cerebral amyloid (or a combination thereof) has been proposed. For successful Aβ immunotherapy, we hypothesized that high affinity antibody binding to amyloid-β plaques and recruitment of brain effector cells is required for most efficient amyloid clearance. Here we report the generation of a novel fully human anti-Aβ antibody, gantenerumab, optimized in vitro for binding with sub-nanomolar affinity to a conformational epitope expressed on amyloid-β fibrils using HuCAL(®) phage display technologies. In peptide maps, both N-terminal and central portions of Aβ were recognized by gantenerumab. Remarkably, a novel orientation of N-terminal Aβ bound to the complementarity determining regions was identified by x-ray analysis of a gantenerumab Fab-Aβ(1-11) complex. In functional assays gantenerumab induced cellular phagocytosis of human amyloid-β deposits in AD brain slices when co-cultured with primary human macrophages and neutralized oligomeric Aβ42-mediated inhibitory effects on long-term potentiation in rat brain. In APP751(swedish)xPS2(N141I) transgenic mice, gantenerumab showed sustained binding to cerebral amyloid-β and, upon chronic treatment, significantly reduced small amyloid-β plaques by recruiting microglia and prevented new plaque formation. Unlike other Aβ antibodies, gantenerumab did not alter plasma Aβ suggesting undisturbed systemic clearance of soluble Aβ. These studies demonstrated that gantenerumab preferentially interacts with aggregated Aβ in the brain and lowers amyloid-β by eliciting effector cell-mediated clearance.
Deriving atomic charges and building a force field library for a new molecule are key steps when developing a force field required for conducting structural and energy-based analysis using molecular mechanics. Derivation of popular RESP charges for a set of residues is a complex and error prone procedure because it depends on numerous input parameters. To overcome these problems, the R.E.D. Tools (RESP and ESP charge Derive, ) have been developed to perform charge derivation in an automatic and straightforward way. The R.E.D. program handles chemical elements up to bromine in the periodic table. It interfaces different quantum mechanical programs employed for geometry optimization and computing molecular electrostatic potential(s), and performs charge fitting using the RESP program. By defining tight optimization criteria and by controlling the molecular orientation of each optimized geometry, charge values are reproduced at any computer platform with an accuracy of 0.0001 e. The charges can be fitted using multiple conformations, making them suitable for molecular dynamics simulations. R.E.D. allows also for defining charge constraints during multiple molecule charge fitting, which are used to derive charges for molecular fragments. Finally, R.E.D. incorporates charges into a force field library, readily usable in molecular dynamics computer packages. For complex cases, such as a set of homologous molecules belonging to a common family, an entire force field topology database is generated. Currently, the atomic charges and force field libraries have been developed for more than fifty model systems and stored in the RESP ESP charge DDataBase. Selected results related to non-polarizable charge models are presented and discussed.
Recent advances in hardware and software have enabled increasingly long molecular dynamics (MD) simulations of biomolecules, exposing certain limitations in the accuracy of the force fields used for such simulations and spurring efforts to refine these force fields. Recent modifications to the Amber and CHARMM protein force fields, for example, have improved the backbone torsion potentials, remedying deficiencies in earlier versions. Here, we further advance simulation accuracy by improving the amino acid side-chain torsion potentials of the Amber ff99SB force field. First, we used simulations of model alpha-helical systems to identify the four residue types whose rotamer distribution differed the most from expectations based on Protein Data Bank statistics. Second, we optimized the side-chain torsion potentials of these residues to match new, high-level quantum-mechanical calculations. Finally, we used microsecond-timescale MD simulations in explicit solvent to validate the resulting force field against a large set of experimental NMR measurements that directly probe side-chain conformations. The new force field, which we have termed Amber ff99SB-ILDN, exhibits considerably better agreement with the NMR data.
The amyloid-beta(1-42) (Abeta42) peptide rapidly aggregates to form oligomers, protofibils and fibrils en route to the deposition of amyloid plaques associated with Alzheimer's disease. We show that low-temperature and low-salt conditions can stabilize disc-shaped oligomers (pentamers) that are substantially more toxic to mouse cortical neurons than protofibrils and fibrils. We find that these neurotoxic oligomers do not have the beta-sheet structure characteristic of fibrils. Rather, the oligomers are composed of loosely aggregated strands whose C termini are protected from solvent exchange and which have a turn conformation, placing Phe19 in contact with Leu34. On the basis of NMR spectroscopy, we show that the structural conversion of Abeta42 oligomers to fibrils involves the association of these loosely aggregated strands into beta-sheets whose individual beta-strands polymerize in a parallel, in-register orientation and are staggered at an intermonomer contact between Gln15 and Gly37.
Synapse loss correlates with a cognitive decline in Alzheimer's disease (AD), but whether this is caused by fibrillar deposits known as senile plaques or soluble oligomeric forms of amyloid beta (Abeta) is controversial. By using array tomography, a technique that combines ultrathin sectioning of tissue with immunofluorescence, allowing precise quantification of small structures, such as synapses, we have tested the hypothesis that oligomeric Abeta surrounding plaques contributes to synapse loss in a mouse model of AD. We find that senile plaques are surrounded by a halo of oligomeric Abeta. Analysis of >14,000 synapses (represented by PSD95-stained excitatory synapses) shows that there is a 60% loss of excitatory synapses in the halo of oligomeric Abeta surrounding plaques and that the density increases to reach almost control levels in volumes further than 50 microm from a plaque in an approximately linear fashion (linear regression, r(2) = 0.9; P < 0.0001). Further, in transgenic cortex, microdeposits of oligomeric Abeta associate with a subset of excitatory synapses, which are significantly smaller than those not in contact with oligomeric Abeta. The proportion of excitatory synapses associated with Abeta correlates with decreasing density (correlation, -0.588; P < 0.0001). These data show that senile plaques are a potential reservoir of oligomeric Abeta, which colocalizes with the postsynaptic density and is associated with spine collapse, reconciling the apparently competing schools of thought of "plaque" vs. "oligomeric Abeta" as the synaptotoxic species in the brain of AD patients.
Rapid progress in deciphering the biological mechanism of Alzheimer's disease (AD) has arisen from the application of molecular and cell biology to this complex disorder of the limbic and association cortices. In turn, new insights into fundamental aspects of protein biology have resulted from research on the disease. This beneficial interplay between basic and applied cell biology is well illustrated by advances in understanding the genotype-to-phenotype relationships of familial Alzheimer's disease. All four genes definitively linked to inherited forms of the disease to date have been shown to increase the production and/or deposition of amyloid β-protein in the brain. In particular, evidence that the presenilin proteins, mutations in which cause the most aggressive form of inherited AD, lead to altered intramembranous cleavage of the β-amyloid precursor protein by the protease called γ-secretase has spurred progress toward novel therapeutics. The finding that presenilin itself may be the long-sought γ-secretase, coupled with the recent identification of β-secretase, has provided discrete biochemical targets for drug screening and development. Alternate and novel strategies for inhibiting the early mechanism of the disease are also emerging. The progress reviewed here, coupled with better ability to diagnose the disease early, bode well for the successful development of therapeutic and preventative drugs for this major public health problem.
The last decades have witnessed a growing global burden of Alzheimer’s disease (AD). Evidence indicates the onset and progression of AD is associated with β-amyloid (Aβ) peptide fibrillation. As such, there is a strong passion with discovering potent Aβ fibrillation inhibitors that can be developed into antiamyloiddogenic agents for AD treatment. Current challenges arisen with this development involve with Aβ oligomer toxicity suppression and Blood Brain Barrier penetration capability. Considering most natural or biological events, one would observe that there is usually a “seed” to direct natural materials to assembly in response to a certain stimulation. Inspired by this, several materials or compounds, including nanoparticle, peptide or peptide mimics, and organic molecules, have been designed for the purpose of redirecting or impeding Aβ aggregation. Achieving these tasks requires comprehensive understanding on (1) how initial Aβ assembly into insoluble deposits, (2) main concerns arisen with fibrillation inhibition, and (3) current major methodologies to disrupt the aggregation. Herein, the objective of this review is to address these three areas, and enable the prompt for a promising therapeutic agent design for AD treatment.
Elucidating pathological fibril structure
Amyloid-β (Aβ) is a key pathological contributor to Alzheimer's disease. Gremer et al. used cryoelectron microscopy data to build a high-quality de novo atomic model of Aβ fibrils (see the Perspective by Pospich and Raunser). The complete structure reveals all 42 amino acids (including the entire N terminus) and provides a structural basis for understanding the effect of several disease-causing and disease-preventing mutations. The fibril consists of two intertwined protofilaments with an unexpected dimer interface that is different from those proposed previously. The structure has implications for the mechanism of fibril growth and will be an important stepping stone to rational drug design.
Science , this issue p. 116 ; see also p. 45
The two hallmarks of Alzheimer’s disease (AD) are the presence of neurofibrillary tangles (NFT) made of aggregates of the hyperphosphorylated tau protein and of amyloid plaques composed of amyloid-β (Aβ) peptides, primarily Aβ1–40 and Aβ1–42. Targeting the production, aggregation, and toxicity of Aβ with small molecule drugs or antibodies is an active area of AD research due to the general acceptance of the amyloid cascade hypothesis, but thus far all drugs targeting Aβ have failed. From a review of the recent literature and our own experience based on in vitro, in silico, and in vivo studies, we present some reasons to explain this repetitive failure.
The dimer of the amyloid-beta peptide Aβ of 42 residues is the smallest toxic species in Alzheimer’s disease, but its equilibrium structures are unknown. Here we determined the equilibrium ensembles generated by the four atomistic OPLS-AA, CHARMM22*, AMBER99sb-ildn and AMBERsb14 force fields with the TIP3P water model. Based on 144 microsecond replica exchange molecular dynamics simulations (with 750 ns per replica), we find that the four force fields lead to random coil ensembles with calculated cross-collision sections, hydrodynamics properties and small-angle X-ray scattering profiles independent on the force field. There are, however, marked differences in secondary structure, with the AMBERsb14 and CHARMM22* ensembles overestimating the CD-derived helix content, and the OPLS-AA and AMBER99sb-ildn secondary structure contents in agreement with CD data. Also the intramolecular beta-hairpin content spanning residues 17-21 and 30-36 varies between 1.5% and 13%. Overall, there are significant differences in tertiary and quaternary conformations among all force fields, and the key finding, irrespective of the force field, is that the dimer is stabilized by nonspecific interactions, explaining therefore its possible transient binding to multiple cellular partners, and in part its toxicity.
Although the amyloid (abeta peptide, Aβ) hypothesis is 25 years old, is the dominant model of Alzheimer's disease (AD) pathogenesis, and guides the development of potential treatments, it is still controversial. One possible reason is a lack of a mechanistic path from the cleavage products of the amyloid precursor protein (APP) such as soluble Aβ monomer and soluble molecular fragments to the deleterious effects on synaptic form and function. From a review of the recent literature and our own published work including aggregation kinetics and structural morphology, Aβ clearance, molecular simulations, long-term potentiation measurements with inhibition binding, and the binding of a commercial monoclonal antibody, aducanumab, we hypothesize that the N-terminal domains of neurotoxic Aβ oligomers are implicated in causing the disease.
Amyloid beta (Aβ) oligomerization is associated with the onset and progression of Alzheimer’s disease (AD). While the A2V mutation enhances aggregation kinetics and toxicity, mixtures of wild-type (WT) and A2V, and also WT and A2T, peptides retard fibril formation and protect against AD. In this study we simulate the equilibrium ensemble of WT:A2T Aβ40 dimer by means of extensive atomistic replica exchange molecular dynamics and compare our results with previous equivalent simulations of A2V:A2V, WT:WT and WT:A2V Aβ40 dimers for a total time scale of nearly 0.1 milliseconds. Qualitative comparison of the resulting thermodynamic properties, such as the relative binding free energies, with the reported experimental kinetic and thermodynamic data affords us important insight into the conversion from slow-pathway to fast-pathway dimer conformations. The crucial reaction coordinate or driving force of such transformation turns out to be related to hydrophobic interpeptide interactions. Analysis of the equilibrium ensembles shows that the fast-pathway conformations contain interpeptide out-of-register antiparallel β-sheet structures at short interpeptide distances. In contrast, the slow-pathway conformations are formed by the association of peptides at large interpeptide distances and high intrapeptide compactness, such as conformations containing intramolecular three-stranded β-sheets which sharply distinguish fast (WT:WT and A2V:A2V) and slow (WT:A2T and WT:A2V) amyloid-forming sequences. Also, this analysis leads us to predict that a molecule stabilizing the intramolecular three-stranded β-sheet or inhibiting the formation of an interpeptide β-sheet spanning residues 17-20 and 31-37 would further reduce fibril formation and probably the cytotoxicity of Aβ species.
Alzheimer's disease (AD) is characterized by deposition of amyloid-β (Aβ) plaques and neurofibrillary tangles in the brain, accompanied by synaptic dysfunction and neurodegeneration. Antibody-based immunotherapy against Aβ to trigger its clearance or mitigate its neurotoxicity has so far been unsuccessful. Here we report the generation of aducanumab, a human monoclonal antibody that selectively targets aggregated Aβ. In a transgenic mouse model of AD, aducanumab is shown to enter the brain, bind parenchymal Aβ, and reduce soluble and insoluble Aβ in a dose-dependent manner. In patients with prodromal or mild AD, one year of monthly intravenous infusions of aducanumab reduces brain Aβ in a dose- and time-dependent manner. This is accompanied by a slowing of clinical decline measured by Clinical Dementia Rating - Sum of Boxes and Mini Mental State Examination scores. The main safety and tolerability findings are amyloid-related imaging abnormalities. These results justify further development of aducanumab for the treatment of AD. Should the slowing of clinical decline be confirmed in ongoing phase 3 clinical trials, it would provide compelling support for the amyloid hypothesis.
Recent experimental data elucidated that 12-crown-4 ether molecule can disrupt Aβ40 fibrils but the mechanism of disruption remains elusive. We have performed a series of all-atom molecular dynamics simulations to study the molecular mechanism of Aβ40 fibril disruption by 12-crown-4. In the present study we have used the Aβ40 fibril trimer as it is the smallest unit that maintains a stable U-shaped structure, and serves as the nucleus to form larger fibrils. Our study reveals that 12-crown-4 ether can enter into the hydrophobic core region and form competitive, hydrophobic interactions with key hydrophobic residues; these interactions break the intersheet hydrophobic interactions and lead to the opening of the U-shaped topology and a loss of β-sheet structure. Furthermore, we observed periods of time when 12-crown-4 was in the hydrophobic core and periods of time when it interacted with Lys28 (chain C), a "tug of war"; the 12-crown-4 binding with Lys28 destabilizes the salt-bridge between Asp23 and Lys28. In addition to the two aforementioned binding modes, the 12-crown-4 binds with Lys16, which is known to form a salt-bridge with Glu22 in antiparallel arranged Aβ fibrils. Our results are in good agreement with experimental results and suggest that molecules that have the ability to interact with both the hydrophobic core region and positively charged residues could serve as potential inhibitors of Aβ fibrils.
Technical aspects of the constant pressure molecular dynamics (MD) method proposed by Andersen and extended by Parrinello and Rahman to allow changes in the shape of the MD cell are discussed. The new MD method is extended to treat molecular systems and to include long range charge-charge interactions.Results on the conservation laws, the frequency of oscillation of the MD cell, and the equations which constrain the shape of the MD cell are also given. An additional constraint is introduced to stop the superfluous MD cell rotation which would otherwise complicate the analysis of crystal structures. The method is illustrated by examining the behaviour of solid nitrogen at high pressure.
Protein aggregation underlies an array of human diseases, yet only one small-molecule therapeutic targeting this process has been successfully developed to date. Here, we introduce an in vivo system, based on a β-lactamase tripartite fusion construct, that is capable of identifying aggregation-prone sequences in the periplasm of Escherichia coli and inhibitors that prevent their aberrant self-assembly. We demonstrate the power of the system using a range of proteins, from small unstructured peptides (islet amyloid polypeptide and amyloid β) to larger, folded immunoglobulin domains. Configured in a 48-well format, the split β-lactamase sensor readily differentiates between aggregation-prone and soluble sequences. Performing the assay in the presence of 109 compounds enabled a rank ordering of inhibition and revealed a new inhibitor of islet amyloid polypeptide aggregation. This platform can be applied to both amyloidogenic and other aggregation-prone systems, independent of sequence or size, and can identify small molecules or other factors able to ameliorate or inhibit protein aggregation.
Oligomeric states of the amyloid β-protein (Aβ) appear to be causally related to Alzheimer's disease (AD). Recently, two familial mutations in the amyloid precursor protein gene have been described, both resulting in amino acid substitutions at Ala2 (A2) within Aβ. An A2V mutation causes autosomal recessive early onset AD. Interestingly, heterozygotes enjoy some protection against development of the disease. An A2T substitution protects against AD and age-related cognitive decline in non-AD patients. Here, we use ion mobility-mass spectrometry (IM-MS) to examine the effects of these mutations on Aβ assembly. These studies reveal different assembly pathways for early oligomer formation for each peptide. A2T Aβ42 formed dimers, tetramers, and hexamers, but dodecamer formation was inhibited. In contrast, no significant effects on Aβ40 assembly were observed. A2V Aβ42 also formed dimers, tetramers, and hexamers, but no dodecamers. However, A2V Aβ42 formed trimers, unlike A2T or wild type (wt) Aβ42. In addition, the A2V substitution caused Aβ40 to oligomerize similar to wt Aβ42, as evidenced by the formation of dimers, tetramers, hexamers, and dodecamers. In contrast, wt Aβ40 formed only dimers and tetramers. These results provide a basis for understanding how these two mutations lead to, or protect against, AD. They also suggest that the Aβ N-terminus, in addition to the oft discussed central hydrophobic cluster and C-terminus, can play a key role in controlling disease susceptibility.
Alzheimer's disease (AD) is one of the most common dementia. The aggregation and deposition of the amyloid-β peptide (Aβ) in neural tissue is its characteristic symptom. In order to destabilize and dissolve Aβ fibrils, a number of candidate molecules have been proposed. Wgx-50 is a compound extracted from Sichuan pepper (Zanthoxylum bungeanum), and a potential candidate drug for treating Alzheimer's disease (AD). Our early experiments show it is effective in disassembling Aβ42 aggregations. To explain the molecular mechanism of the destabilization of Aβ42 protofibril by wgx-50, a series of molecular dynamics simulations were performed in this work. It is found that there were three possible stable binding sites including two sites in hydrophobic grooves on surface of Aβ protofibril that made no significant changes in Aβ structures, and one site in the interior that caused destabilization of the protofibril. In this site, wgx-50 was packed against the side chains of I32 and L34, disrupted the D23-K28 salt bridges, and partially opened the tightly compacted two β-sheets. The results were confirmed by simulations at 320K, where deeper insertion of wgx-50 into the whole protofibril was observed. The molecular mechanism of this novel drug candidate wgx-50 to disaggregate Aβ protofibril may provide some insight on the strategy of structure-based drug design for Alzheimer's disease.
Increasing evidence has suggested that formation and propagation of misfolded aggregates of 42-residue human amyloid β (Aβ(1-42)), rather than of the more abundant Aβ(1-40), provokes the Alzheimer's disease cascade. However, structural details of misfolded Aβ(1-42) have remained elusive. Here we present the atomic model of an Aβ(1-42) amyloid fibril, from solid-state NMR (ssNMR) data. It displays triple parallel-β-sheet segments that differ from reported structures of Aβ(1-40) fibrils. Remarkably, Aβ(1-40) is incompatible with the triple-β-motif, because seeding with Aβ(1-42) fibrils does not promote conversion of monomeric Aβ(1-40) into fibrils via cross-replication. ssNMR experiments suggest that C-terminal Ala42, absent in Aβ(1-40), forms a salt bridge with Lys28 to create a self-recognition molecular switch that excludes Aβ(1-40). The results provide insight into the Aβ(1-42)-selective self-replicating amyloid-propagation machinery in early-stage Alzheimer's disease.
The misfolding and aggregation of amyloid beta (Aβ) peptides into amyloid fibrils are key events in the amyloid cascade hypothesis for the etiology of Alzheimer's disease (AD). Using thioflavin-T (ThT) fluorescence assay, atomic force microscopy, circular dichroism, size exclusion chromatography, surface plasmon resonance (SPR), and cytotoxicity tests, we demonstrate that tabersonine, an ingredient extracted from the bean of Voacanga africana, disrupts the Aβ(1-42) aggregation and ameliorates Aβ aggregate-induced cytotoxicity. A small amount of tabersonine (e.g., 10 μM) can effectively inhibit the formation of Aβ(1-42) (e.g., 80 μM) fibrils or convert mature fibrils into largely innocuous amorphous aggregates. SPR results indicate that tabersonine binds to Aβ(1-42) oligomers in a dose-dependent way. Molecular dynamics (MD) simulations further confirm that tabersonine can bind to oligomers such as the pentamer of Aβ(1-42). Tabersonine preferentially interact with the β-sheet grooves of Aβ(1-42) containing aromatic and hydrophobic residues. The various binding sites and modes explain the diverse inhibitory effects of tabersonine on Aβ aggregation. Given that tabersonine is a natural product and a precursor for vincristine used in cancer chemotherapy, the biocompatibility and small size essential for permeating the blood brain barrier make it a potential therapeutic drug candidate for treating AD.
Researchers present an in-depth review on the contribution of biophysical and biochemical studies and computer simulations to characterize the molecular structures of Aβ1-40/1-42 monomers, oligomers, protofibrils, and amyloid fibrils in aqueous solution. They focus their current knowledge of the Aβ1-40/1-42 nucleus and the structures and dynamics of Aβ1-40/1-42 oligomers in proximity of or at the membrane. They also the available information regarding the interactions of Aβ monomers and oligomers with ion metals, cellular partners, and potential inhibitors.
Curcumin, a chemical constituent present in the spice turmeric is known to prevent the aggregation of amyloid peptide implicated in the pathophysiology of Alzheimer's disease. While curcumin is known to bind directly to various amyloid aggregates, no systematic investigations have been carried out to understand its ability to bind to the amyloid aggregates including oligomers and fibrils. In this study, we constructed computational models of (i) Aβ hexapeptide (16) KLVFFA(21) octamer steric zipper β-sheet assembly and (ii) full length Aβ fibril β-sheet assembly. Curcumin binding in these models was evaluated by molecular docking and molecular dynamics (MD) simulation studies. In both the models, curcumin was oriented in a linear extended conformation parallel to fiber axis and exhibited better stability in the Aβ hexapeptide (16) KLVFFA(21) octamer steric zipper model (Ebinding = -10.05 kcal/mol) compared to full length Aβ fibril model (Ebinding = -3.47 kcal/mol). Analysis of MD trajectories of curcumin bound to full length Aβ fibril shows good stability with minimum Cα-atom RMSD shifts. Interestingly, curcumin binding led to marked fluctuations in the (14) HQKLVFFA(21) region that constitute the fibril spine with RMSF values ranging from 1.4-3.6 Å. These results show that curcumin binding to Aβ shifts the equilibrium in the aggregation pathway by promoting the formation of nontoxic aggregates. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
Aggregation of the amyloid β protein (Aβ) peptide with 40 or 42 residues is one key feature in Alzheimer’s disease (AD). The 1,4-naphthoquinon-2-yl-l-tryptophan (NQTrp) molecule was reported to alter Aβ self-assembly and reduce toxicity. Though nuclear magnetic resonance experiments and various simulations provided atomic information about the interaction of NQTrp with Aβ peptides spanning the regions of residues 12–28 and 17–42, none of these studies were conducted on the full-length Aβ1–42 peptide. To this end, we performed extensive atomistic replica exchange molecular dynamics simulations of Aβ1–42 dimer with two NQTrp molecules in explicit solvent, by using a force field known to fold diverse proteins correctly. The interactions between NQTrp and Aβ1–42, which change the Aβ interface by reducing most of the intermolecular contacts, are found to be very dynamic and multiple, leading to many transient binding sites. The most favorable binding residues are Arg5, Asp7, Tyr10, His13, Lys16, Lys18, Phe19/Phe20, and Leu34/Met35, providing therefore a completely different picture from in vitro and in silico experiments with NQTrp with shorter Aβ fragments. Importantly, the 10 hot residues that we identified explain the beneficial effect of NQTrp in reducing both the level of Aβ1–42 aggregation and toxicity. Our results also indicate that there is room to design more efficient drugs targeting Aβ1–42 dimer against AD.
Alzheimer's disease (AD) is the most common form of dementia and the sixth leading cause of death in the United States. Plaques comprised of aggregated amyloid-beta protein (Aβ) accumulate between the neural cells in the brain and are associated with dementia and cellular death. Many strategies have been investigated to prevent Aβ aggregation and thus the build-up of Aβ plaques; however a promising therapeutic has not yet been identified. In this study, a peptoid-based mimic of the peptide KLVFF (residues 16-20 of Aβ) was tested for its ability to modulate Aβ aggregation. Peptoid JPT1 includes chiral, aromatic side chains to induce formation of a stable helical secondary structure that allows for greater interaction between the aromatic side chains and the cross beta sheet of Aβ. JPT1 was found to modulate Aβ40 aggregation, specifically decreasing lag time and the total number of aggregates formed. These results suggest that peptoids may be able to limit the formation of Aβ aggregates that are associated with AD.
Evolution has fine-tuned proteins to accomplish a variety of tasks. Yet, with aging, some proteins assemble into harmful amyloid aggregates associated with neurodegenerative diseases, such as Alzheimer’s disease (AD), which presents a complex and costly challenge to our society. Thus, far, drug after drug has failed to slow the progression of AD, characterized by the self-assembly of the 39–43 amino acid β-amyloid (Aβ) protein into extracellular senile plaques that form a cross-β structure. While there is experimental evidence that the Aβ small oligomers are the primary toxic species, standard tools of biology have failed to provide structures of these transient, inhomogeneous assemblies. Despite extensive experimental studies, researchers have not successfully characterized the nucleus ensemble, the starting point for rapid fibril formation. Similarly scientists do not have atomic data to show how the compounds that reduce both fibril formation and toxicity in cells bind to Aβ42 oligomers. In this context, computer simulations are important tools for gaining insights into the self-assembly of amyloid peptides and the molecular mechanism of inhibitors.
Recent experiments have shown that the mutation Tottori (D7N) alters the toxicity, assembly and rate of fibril formation of the wild type (WT) amyloid beta (Aβ) Aβ40 and Aβ42 peptides. We used all-atom molecular dynamics simulations in explicit solvent of the monomer and dimer of both alloforms with their WT and D7N sequences. The monomer simulations starting from a random coil and totaling 3 μs show that the D7N mutation changes the fold and the network of salt bridges in both alloforms. The dimer simulations starting from the amyloid fibrillar states and totaling 4.4 μs also reveal noticeable changes in terms of secondary structure, salt bridge, and topology. Overall, this study provides physical insights into the enhanced rate of fibril formation upon D7N mutation and an atomic picture of the D7N-mediated conformational change on Aβ40 and Aβ42 peptides.
In vitro, β-amyloid (Aβ) peptides form polymorphic fibrils, with molecular structures that depend on growth conditions, plus various oligomeric and protofibrillar aggregates. Here, we investigate structures of human brain-derived Aβ fibrils, using seeded fibril growth from brain extract and data from solid-state nuclear magnetic resonance and electron microscopy. Experiments on tissue from two Alzheimer's disease (AD) patients with distinct clinical histories showed a single predominant 40 residue Aβ (Aβ40) fibril structure in each patient; however, the structures were different from one another. A molecular structural model developed for Aβ40 fibrils from one patient reveals features that distinguish in-vivo- from in-vitro-produced fibrils. The data suggest that fibrils in the brain may spread from a single nucleation site, that structural variations may correlate with variations in AD, and that structure-specific amyloid imaging agents may be an important future goal. PAPERFLICK:
An N·log(N) method for evaluating electrostatic energies and forces of large periodic systems is presented. The method is based on interpolation of the reciprocal space Ewald sums and evaluation of the resulting convolutions using fast Fourier transforms. Timings and accuracies are presented for three large crystalline ionic systems. The Journal of Chemical Physics is copyrighted by The American Institute of Physics.
This review is intended to reflect the recent progress on therapeutic applications of nanomaterials in amyloid diseases. The progress on anti-amyloid functions of various nanomaterials including inorganic nanoparticles, polymeric nanoparticles, carbon nanomaterials and biomolecular aggregates, is reviewed and discussed. The main functionalization strategies for general nanoparticle modifications are reviewed for potential applications of targeted therapeutics. The interaction mechanisms between amyloid peptides and nanomaterials are discussed from the perspectives of dominant interactions and kinetics. The encapsulation of anti-amyloid drugs, targeted drug delivery, controlled drug release and drug delivery crossing blood brain barrier by application of nanomaterials would also improve the therapeutics of amyloid diseases.
For a successful analysis of the relation between amino acid sequence and protein structure, an unambiguous and physically meaningful definition of secondary structure is essential. We have developed a set of simple and physically motivated criteria for secondary structure, programmed as a pattern-recognition process of hydrogen-bonded and geometrical features extracted from x-ray coordinates. Cooperative secondary structure is recognized as repeats of the elementary hydrogen-bonding patterns “turn” and “bridge.” Repeating turns are “helices,” repeating bridges are “ladders,” connected ladders are “sheets.” Geometric structure is defined in terms of the concepts torsion and curvature of differential geometry. Local chain “chirality” is the torsional handedness of four consecutive Cα positions and is positive for right-handed helices and negative for ideal twisted β-sheets. Curved pieces are defined as “bends.” Solvent “exposure” is given as the number of water molecules in possible contact with a residue. The end result is a compilation of the primary structure, including SS bonds, secondary structure, and solvent exposure of 62 different globular proteins. The presentation is in linear form: strip graphs for an overall view and strip tables for the details of each of 10.925 residues. The dictionary is also available in computer-readable form for protein structure prediction work.
The 'toxic Aβ oligomer' hypothesis has attracted considerable attention among Alzheimer's disease researchers as a way of resolving the lack of correlation between deposited amyloid-β (Aβ) in amyloid plaques-in terms of both amount and location-and cognitive impairment or neurodegeneration. However, the lack of a common, agreed-upon experimental description of the toxic Aβ oligomer makes interpretation and direct comparison of data between different research groups impossible. Here we critically review the evidence supporting toxic Aβ oligomers as drivers of neurodegeneration and make some suggestions that might facilitate progress in this complex field.
Experimental and computational studies of Aβ amyloids have suggested that for any given segment there are one or more preferred parallel and (or) antiparallel structural states. The preferred organizations of the Aβ fragments do not appear to present straight forward rules with respect to length, hydrophobicity, and charge. Because polymorphism is presented by different Aβ segments, clearly a combination of these segments would lead to polymorphic full-length Aβ, although the relative populations in the full sequence are likely to be different. How the Aβ peptides assemble and form toxic entities and what is the mechanism of toxicity are major questions that persist in Alzheimer research. Two types of models of the three-dimensional structures of Aβ oligomers have been reported from computational and experimental studies. Because metal ions can coordinate with different residues in each structural model, the variety of the morphologies can increase quickly.
Alzheimer's disease is a progressive, neurodegenerative disorder that is the leading cause of senile dementia, afflicting millions of individuals worldwide. Since the identification of the amyloid beta-peptide (Abeta) as the principal toxic entity in the progression of Alzheimer's disease, numerous attempts have been made to reduce endogenous Abeta production and deposition, including designing inhibitors of the proteases that generate the peptide, generating antibodies against Abeta aggregates, utilizing metal chelators, and identifying small molecules that target the peptide during the aggregation pathway. The last approach is particularly attractive, as Abeta is normally present in vivo, but aggregation is a purely pathological event. Studies conducted in vitro and in vivo have suggested that administration of flavonoids, compounds naturally present in many foods, including wine and tea, can prevent and reverse Abeta aggregation, but mechanistic details are lacking. In this work, we employ atomistic, explicit-solvent molecular dynamics (MD) simulations to identify the mechanism of Abeta fibril destabilization by morin, one of the most effective anti-aggregation flavonoids, using a model of the mature Abeta fibril. Through the course of 24 simulations totaling 4.3 mus, we find that morin can bind to the ends of the fibrils to block the attachment of an incoming peptide and can penetrate into the hydrophobic core to disrupt the Asp23-Lys28 salt bridges and interfere with backbone hydrogen bonding. The combination of hydrophobicity, aromaticity, and hydrogen bonding capacity of morin imparts the observed behavior.
Recent experimental data demonstrate that small, soluble amyloid-beta42 oligomers play an important role in Alzheimer's disease because they exhibit neurotoxic properties and also act as seed for fibril growth. We performed all-atom molecular dynamics simulations in explicit solvent of 0.7 micros in total on five Abeta9-42 oligomers (monomer through pentamer) starting from the fibril conformation. The initial conformation proves to be stable in the trimer to pentamer, and the two parallel in-register beta-sheets as well as the connecting turn are preserved. The dimer undergoes larger conformational changes in its C-terminus, and the predominant conformation detected exhibits an additional antiparallel beta-sheet in one of the subunits. This conformational rearrangement allows efficient shielding of hydrophobic residues from the solvent, which is not possible for a dimer in the fibril conformation. In addition to the presence of the hydrogen bonds in the beta-sheets, the larger oligomers are stabilized by interchain D23-K28 salt bridges, whereas a D23-N27 interaction is found in the dimer. The degree of structural similarity to the fibril conformation detected for the oligomers in the simulation may also offer a structural explanation for the experimental finding that trimers and tetramers act as more potent seeds in fibril formation than dimers because only small conformational changes will be required for fibril growth. The fact that the dimer predominantly exists in conformations distinct from the larger oligomers and the fibril is also interesting for the design of anti-Alzheimer drugs, because it suggests that multiple drugs might be required to target the structurally different neurotoxic oligomers.
Amyloid fibrils represent a stable form of many misfolded proteins associated with numerous diseases. Among these are Parkinson's disease (alpha-synuclein), Type II diabetes (islet amyloid polypeptide), and Alzheimer's disease (amyloid beta-peptide, Abeta). The appearance of Abeta fibrils in neural tissue is a hallmark of Alzheimer's disease, and many studies have been conducted to determine and analyze the structure of these protein aggregates. The principal toxic species in Alzheimer's disease are believed to be soluble, oligomeric aggregates of Abeta, but numerous studies have found that the insoluble fibrillated form of the peptide also contributes to neurotoxicity. Thus, to design therapeutic agents to combat the progression of Alzheimer's disease, it is worthwhile to understand the thermodynamics of destabilizing these aggregates and the features that contribute to their stability. In this work, we present a systematic study of several factors that influence the stability of Abeta(42) fibrils following in silico mutation. We have employed standard molecular dynamics, as well as center-of-mass pulling and umbrella sampling, to study the thermodynamics of peptide dissociation from the core of a model protofibril at physiological temperature. Results indicate that a finite level of hydration around the Asp23-Lys28 salt bridge is crucial to protofibril stability, while mutation of Phe19 to glycine has no effect on the binding free energy of the terminal peptide. Packing between Ile32 and the aliphatic portion of the Lys28 side chain serves to regulate the level of hydration in the core of the protofibril and thus rigidify the Asp23-Lys28 salt bridge. These observations are important for designing compounds that target Abeta aggregates; interrupting these native interactions may destabilize these assemblies and ameliorate their toxicity.
Recent experiments have shown that the congener Abeta(1-40)[D23-K28], in which the side chains of charged residues Asp23 and Lys28 are linked by a lactam bridge, forms amyloid fibrils that are structurally similar to the wild type (WT) Abeta peptide, but at a rate that is nearly 1000 times faster. We used all atom molecular dynamics simulations in explicit water, and two force fields, of the WT dimer, a monomer with the lactam bridge (Abeta(10-35)-lactam[D23-K28]), and the monomer and dimers with harmonically constrained D23-K28 salt bridge (Abeta(10-35)[D23-K28]) to understand the origin of the enhanced fibril rate formation. The simulations show that the assembly competent fibril-like monomer (N*) structure, which is present among the conformations sampled by the isolated monomer, with strand conformations in the residues spanning the N and C termini and a bend involving residues D(23) VGSNKG(29), are populated to a much greater extent in Abeta(10-35)[D23-K28] and Abeta(10-35)-lactam[D23-K28] than in the WT, which has negligible probability of forming N*. The salt bridge in N* of Abeta(10-35)[D23-K28], whose topology is similar to that found in the fibril, is hydrated. The reduction in the free energy barrier to fibril formation in Abeta(10-35)[D23-K28] and in Abeta(10-35)-lactam[D23-K28], compared to the WT, arises largely due to entropic restriction which enables the bend formation. A decrease in the entropy of the unfolded state and the lesser penalty for conformational rearrangement including the formation of the salt bridge in Abeta peptides with D23-K28 constraint results in a reduction in the kinetic barrier in the Abeta(1-40)-lactam[D23-K28] congener compared to the WT. The decrease in the barrier, which is related to the free energy cost of forming a bend, is estimated to be in the range (4-7)k(B)T. Although a number of factors determine the growth of fibrils, the decrease in the free energy barrier, relative to the WT, to N* formation is a major factor in the rate enhancement in the fibril formation of Abeta(1-40)[D23-K28] congener. Qualitatively similar results were obtained using simulations of Abeta(9-40) peptides and various constructs related to the Abeta(10-35) systems that were probed using OPLS and CHARMM force fields. We hypothesize that mutations or other constraints that preferentially enhance the population of the N* species would speed up aggregation rates. Conversely, ligands that lock it in the fibril-like N* structure would prevent amyloid formation.