Dissecting the effect of ALS mutation S375G on the conformational properties and aggregation dynamics of TDP-43370-375 fragment.

The aggregation of transactive response deoxyribonucleic acid (DNA) binding protein of 43 kDa (TDP-43) into ubiquitin-positive inclusions is closely associated with amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration, and chronic traumatic encephalopathy. The 370-375 fragment of TDP-43 (370GNNSYS375, TDP-43370-375), the amyloidogenic hexapeptides, can be prone to forming pathogenic amyloid fibrils with the characteristic of steric zippers. Previous experiments reported the ALS-associated mutation, serine 375 substituted by glycine (S375G) is linked to early onset disease and protein aggregation of TDP-43. Based on this, it is necessary to explore the underlying molecular mechanisms. By utilizing all-atom molecular dynamics (MD) simulations of 102 µs in total, we investigated the impact of S375G mutation on the conformational ensembles and oligomerization dynamics of TDP-43370-375 peptides. Our replica exchange MD simulations show that S375G mutation could promote the unstructured conformation formation and induce peptides to form a loose packed oligomer, thus inhibiting the aggregation of TDP-43370-375. Further analyses suggest that S375G mutation displays a reduction effect on the number of total hydrogen bonds and contacts among TDP-43370-375 peptides. Hydrogen bonding and polar interactions among TDP-43370-375 peptides, as well as Y374-Y374 π-π stacking interaction, are attenuated by S375G mutation. Additional microsecond MD simulations demonstrate that S375G mutation could prohibit the conformational conversion to ß-structure-rich aggregates and possess an inhibitory effect on the oligomerization dynamics of TDP-43370-375. This study offers for the first time of molecular insights into the S375G mutation affecting the aggregation of TDP-43370-375 at the atomic level, and may open new avenues in the development of future site-specific mutation therapeutics.

Destabilization mechanism of R3-R4 tau protofilament by purpurin: a molecular dynamics study.

The accumulation of tau protein aggregates is a common feature observed in many neurodegenerative diseases. However, the structural characteristics of tau aggregates can vary among different tauopathies. It has been established that the structure of the tau protofilament in Chronic traumatic encephalopathy (CTE) is similar to that of Alzheimer's disease (AD). In addition, a previous study found that purpurin, an anthraquinone, could inhibit and disassemble the pre-formed 306VQIVYK311 isoform of AD-tau protofilament. Herein, we used all-atom molecular dynamic (MD) simulation to investigate the distinctive features between CTE-tau and AD-tau protofilament and the influence of purpurin on CTE-tau protofilament. Our findings revealed notable differences at the atomic level between CTE-tau and AD-tau protofilaments, particularly in the ß6-ß7 angle and the solvent-accessible surface area (SASA) of the ß4-ß6 region. These structural disparities contributed to the distinct characteristics observed in the two types of tau protofilaments. Our simulations substantiated that purpurin could destabilize the CTE-tau protofilament and decrease ß-sheet content. Purpurin molecules could insert the ß4-ß6 region and weaken the hydrophobic packing between ß1 and ß8 through π-π stacking. Interestingly, each of the three rings in purpurin exhibited unique binding preferences with the CTE-tau protofilament. Overall, our study sheds light on the structural distinctions between CTE-tau and AD-tau protofilaments, as well as the destabilizing mechanism of purpurin on CTE-tau protofilament, which may be helpful to the development of drugs to prevent CTE.

Inhibitive and Destructive Mechanisms of Chronic Traumatic Encephalopathy-Related R3-R4 Tau Peptide Chains and Protofibril by Epigallocatechin Gallate: Evidence from Molecular Dynamics Simulation.

Chronic traumatic encephalopathy (CTE), a unique tauopathy, is pathologically associated with the aggregation of hyperphosphorylated tau protein 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 ß-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 ß-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.

Molecular Mechanism in the Disruption of Chronic Traumatic Encephalopathy-Related R3-R4 Tau Protofibril by Quercetin and Gallic Acid: Similarities and Differences.

Chronic traumatic encephalopathy (CTE) is a unique progressive neurodegenerative tauopathy pathologically related to the aggregation of the tau protein to neurofibrillary tangles. Disrupting tau oligomers (protofibril) is a promising strategy to prevent CTE. Quercetin (QE) and gallic acid (GA), two polyphenol small molecules abundant in natural crops, were proved to inhibit recombinant tau and the R3 fragment of human full-length tau in vitro. However, their disruptive effect on CTE-related protofibril and the underlying molecular mechanism remain elusive. Cryo-electron microscopy resolution reveals that the R3-R4 fragment of tau forms the core of the CTE-related tau protofibril. In this study, we conducted extensive all-atom molecular dynamics simulations on CTE-related R3-R4 tau protofibril with and without QE/GA molecules. The results disclose that both QE and GA can disrupt the global structure of the protofibril, while GA shows a relatively strong effect. The binding sites, exact binding patterns, and disruptive modes for the two molecules show similarities and differences. Strikingly, both QE and GA can insert into the hydrophobic cavity of the protofibril, indicating they have the potential to compete for the space in the cavity with aggregation cofactors unique to CTE-related protofibril and thus impede the further aggregation of the tau protein. Due to relatively short time scale, our study captures the early disruptive mechanism of CTE-related R3-R4 tau protofibril by QE/GA. However, our research does provide valuable knowledge for the design of supplements or drugs to prevent or delay the development of CTE.

Dissecting the Inhibitory Mechanism of the αB-Crystallin Domain against Aß42 Aggregation and Its Effect on Aß42 Protofibrils: A Molecular Dynamics Simulation Study.

Alzheimer's disease (AD) is related to the misfolding and aggregation of amyloid-ß (Aß) protein, and its major pathological hallmark is fibrillary ß-amyloid plaques. Impeding the formation of Aß ß-structure-rich aggregates and dissociating Aß 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 Aß 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 Aß42 aggregation. Our simulations show that αBC greatly impedes the formation of ß-structure contents. We find that the binding of αBC to the Aß42 monomer is driven by polar, hydrophobic, and H-bonding interactions. To explore whether αBC could destabilize Aß42 protofibrils, we also carried out MD simulations of Aß42 protofibrils with and without αBC. The results show that αBC interacts with three binding sites of the Aß42 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 ß-structure content, kink angle, and K28-A42 salt bridges. Overall, this study not only discloses the molecular mechanism of αBC against Aß42 aggregation but also demonstrates the disruption effects of αBC on Aß42 protofibrils, which yields an avenue for designing anti-AD drug candidates.

The destructive mechanism of Aß1-42 protofibrils by norepinephrine revealed via molecular dynamics simulations.

Amyloid-ß (Aß) fibrillary plaques represent the main hallmarks of Alzheimer's disease (AD), in addition to tau neurofibrillary tangles. Disrupting early-formed Aß protofibrils is considered to be one of the primary therapeutic strategies to interfere with AD. Our previous work showed that norepinephrine (NE), an important neurotransmitter in the brain, can effectively inhibit the aggregation of the Aß1-42 peptide. However, whether and how NE molecules disassemble Aß1-42 protofibrils remains to be elucidated. Herein we investigate the influence of NE (in protonated and deprotonated states) on the recently cryo-EM solved LS-shaped Aß1-42 protofibrils and the underlying molecular mechanism by performing all-atom molecular dynamics simulations. Our simulations showed that protonated and deprotonated NE exhibited distinct disruptive mechanisms on Aß1-42 protofibrils. Protonated NE could significantly disrupt the N-terminal (residues D1-H14) structure of Aß1-42 protofibrils and destabilize the global structure of the protofibril. It preferentially bound with N-terminal residues of Aß1-42 protofibrils and formed hydrogen bonds with E3, D7, E11, Q15, E22, and D23 residues and π-π stackings with H6, H13, and F20 residues, and thus destroyed the hydrogen bonds between H6 and E11 and increased the kink angle around Y10. Compared to protonated NE, deprotonated NE displayed a higher disruptive capability on Aß1-42 protofibrils, and stronger hydrophobic and π-π stacking interactions with the protofibril structure. This study revealed the molecular mechanism of NE in the destruction of Aß1-42 protofibrils, which may be helpful in the design of potent drug candidates against AD.

Mechanistic insight into the disruption of Tau R3-R4 protofibrils by curcumin and epinephrine: an all-atom molecular dynamics study.

The accumulation of Tau protein aggregates is a pathological hallmark of tauopathy, including chronic traumatic encephalopathy (CTE). Inhibiting Tau aggregation or disrupting preformed Tau fibrils is considered one of the rational therapeutic strategies to combat tauopathy. Previous studies reported that curcumin (Cur, a molecule of a labile natural product) and epinephrine (EP, an important neurotransmitter) could effectively inhibit the formation of Tau fibrillar aggregates and disassociate preformed fibrils. However, the underlying molecular mechanisms remain elusive. In this study, we performed multiple molecular dynamics simulations for 17.5 µs in total to investigate the influence of Cur and EP on the C-shaped Tau protofibril associated with CTE. Our simulations show that the protofibrillar pentamer is the smallest stable Tau R3-R4 protofibril. Taking the pentamer as a protofibril model, we found that both Cur and EP molecules could affect the shape of the Tau pentamer by changing the ß2-ß3 and ß7-ß8 angles, leading to a more extended structure. Cur and EP display a disruptive effect on the local ß-sheets and the formation of hydrogen bonds, and thus destabilize the global protofibril structure. The contact number analysis shows that Cur has a higher binding affinity with the Tau pentamer than EP, especially in the nucleating segment PHF6. Hydrophobic, π-π and cation-π interactions together facilitate the binding of Cur and EP with the Tau pentamer. Cur exhibits stronger hydrophobic and π-π interactions with Tau than EP, and EP displays a stronger cation-π interaction. Our findings provide molecular insights into the disruptive mechanisms of the Tau R3-R4 protofibrils by curcumin and epinephrine, which may be useful for the design of effective drug candidates for the treatment of CTE.

Receptor Activator of Nuclear Factor (Nf)-kb Ligand Promotes T Helper 17 Cell Differentiation through Fas.

T helper 17 (Th17) cells play important role in the defense against pathogens and autoimmune diseases. Many cytokines can induce Th17 cell differentiation. However, the mechanism of Th17 cell differentiation is not well clarified. RankL, a member of the TNF superfamily, binds with Rank and then participates in the proliferation and differentiation of many kinds of cells. Recent studies showed that RankL-Rank signaling is closely related to Th17 differentiation and function. The detail of the Rank-RankL pathway in Th17 cell differentiation is still unclear. To illustrate the role of Rank-RankL in Th17 differentiation, naive CD4 + T cells were differentiated into Th17 cells with or without RankL stimulation. During Th17 differentiation, the expression of Rank obviously increased. The RankL stimulation significantly increased Th17 cell differentiation indicated by increased IL-17-positive cell number, highly expressed IL-17 and IL-22 and elevated IL-17 secretion. These effects were canceled by Rank-Fc addition. In further study, RankL treatment during Th17 differentiation up-regulated Fas expression. Fas knockdown inhibited the Th17 differentiation promoted by RankL. In this study, it was confirmed that Rank-RankL signaling could promote Th17 cell differentiation through Fas induction.

Mechanisms of melatonin binding and destabilizing the protofilament and filament of tau R3-R4 domains revealed by molecular dynamics simulation.

The accumulation of ß-amyloid (Aß) and tau protein is considered to be an important pathological characteristic of Alzheimer's disease (AD). Failure of medicine targeting Aß has drawn more attention to the influence of tau protein and its fibrillization on neurodegeneration. Increasing evidence shows that melatonin (Mel) can effectively inhibit the formation of tau fibrils and disassemble preformed tau fibrils. However, the underlying mechanism is poorly understood. In this work, we investigated the kinetics of melatonin binding and destabilizing the tetrameric protofilament and octameric filament of tau R3-R4 domains by performing microsecond all-atom molecular dynamics simulations. Our results show that Mel is able to disrupt the C-shaped structure of the tau protofilament and filament, and destabilizes the association between N- and C-termini. Mel predominantly binds to ß1 and ß6-ß8 regions and favors contact with the elongation surface, which is dominantly driven by hydrogen bonding interactions and facilitated by other interactions. The strong π-π stacking interaction of Mel with Y310 impedes the intramolecular CH-π interaction between I308 and Y310, and the cation-π interaction of Mel with R379 interferes with the formation of the D348-R379 salt bridge. Moreover, Mel occupies the protofilament surface in the tetrameric protofilament and prevents the formation of intermolecular hydrogen bonds between residues K331 and Q336 in the octameric filament. Our work provides molecular insights into Mel hindering tau fibrillization or destabilizing the protofilament and filament, and the revealed inhibitory mechanisms provide useful clues for the design of efficient anti-amyloid agents.

Trends in HSPB5 research: a 36-year bibliometric analysis.

HSPB5 (heat shock protein B5), also known as αB-crystallin, is one of the most widespread and populous of the ten human small heat shock proteins (sHsps). Over the past decades, extensive research has been conducted on HSPB5. However, few studies have statistically analyzed these publications. Herein, we conducted a bibliometric analysis to track the global research trend and current development status of HSPB5 research from the Web of Science Core Collection (WoSCC) database between 1985 and 2020. Our results demonstrate that 1220 original articles cited 54,778 times in 391 scholarly journals were published. Visualization analyses reveal that the Journal of Biological Chemistry was the most influential journal with 85 articles. The USA dominated this field with 520 publications (42.62%), followed by Japan with 149 publications (12.21%), and Kato contributed the largest number of publications. Most related publications were published in journals focusing on biochemistry molecular biology, cell biology, neurosciences neurology, and ophthalmology. In addition, keyword co-occurrence analyses identify three predominant research topics: expression of HSPB5, chaperone studies for HSPB5, and pathological studies of HSPB5. This study provides valuable guidance for researchers and leads to collaborative opportunities between diverse research interests to be integrated for HSPB5 research.

Serotonin and Melatonin Show Different Modes of Action on Aß42 Protofibril Destabilization.

Alzheimer's disease (AD) is associated with the aberrant self-assembly of amyloid-ß (Aß) protein into fibrillar deposits. The disaggregation of Aß fibril is believed as one of the major therapeutic strategies for treating AD. Previous experimental studies reported that serotonin (Ser), one of the indoleamine neurotransmitters, and its derivative melatonin (Mel) are able to disassemble preformed Aß fibrils. However, the fibril-disruption mechanisms are unclear. As the first step to understand the underlying mechanism, we investigated the interactions of Ser and Mel molecules with the LS-shaped Aß42 protofibril by performing a total of nine individual 500 ns all-atom molecular dynamics (MD) simulations. The simulations demonstrate that both Ser and Mel molecules disrupt the local ß-sheet structure, destroy the salt bridges between K28 side chain and A42 COO-, and consequently destabilize the global structure of Aß42 protofibril. The Mel molecule exhibits a greater binding capacity than the Ser molecule. Intriguingly, we find that Ser and Mel molecules destabilize Aß42 protofibril through different modes of action. Ser preferentially binds with the aromatic residues in the N-terminal region through π-π stacking interactions, while Mel binds not only with the N-terminal aromatic residues but also with the C-terminal hydrophobic residues via π-π and hydrophobic interactions. This work reveals the disruptive mechanisms of Aß42 protofibril by Ser and Mel molecules and provides useful information for designing drug candidates against AD.

Molecular dynamics simulations reveal the destabilization mechanism of Alzheimer's disease-related tau R3-R4 Protofilament by norepinephrine.

Aggregation of Tau protein into neurofibrillary tangles is associated with the pathogenesis of Alzheimer's disease (AD) which has no cure yet. Clearing neurofibrillary tangles is one of major therapeutic strategies. Experimental studies reported that norepinephrine (NE) has the ability to disrupt Tau filament and cause Tau degradation. However, the underlying mechanism remains elusive. Herein, we performed molecular dynamic simulations to investigate the influence of NE on the C-shaped Tau R3-R4 protofilament. Our simulations show that NE compound destabilizes Tau protofilament by mostly disrupting ß6/ß8 and altering the ß2-ß3 and ß6-ß7 angles. NE binds mainly with aromatic residues Y310/P312/H374/F378 through ππ stacking and charged residues E338/E342/D348/D358/E372 via hydrogen-bonding interactions. Our results, together with the findings that exercise can markedly increase NE level, suggest that exercise might be a potent therapy against AD. This study reveals the disruptive mechanism of Tau protofilament by NE molecules, which may provide new clues for AD drug candidate design.

Critical nucleus of Greek-key-like core of α-synuclein protofibril and its disruption by dopamine and norepinephrine.

The formation of amyloid fibrils by α-synuclein (αS) protein inside the Lewy bodies and Lewy neurites is the prominent pathological hallmark of Parkinson's disease (PD). The fibrillation of αS in vitro is described by a nucleation-elongation process involving the formation of a critical nucleus. Finding the critical/smallest nuclei and effective inhibitors of αS aggregation is a crucial step for the development of drugs against PD. Recent experiments reported that dopamine (DA) and norepinephrine (NE), two prominent naturally occurring neurotransmitters, can effectively disrupt the preformed αS fibrils. The level of DA/NE in blood can be markedly increased by exercise. However, the size and structure of the critical nucleus and the disruptive mechanism by DA/NE are largely unknown. In this work, we performed multiple molecular dynamics (MD) simulations to find the critical nucleus size and examine the influences of DA/NE molecules on preformed αS44-96 (Greek-key-like core of full length αS) protofibrils. Our results show that the trimer is the critical nucleus for the αS44-96 fibril formation, and the tetramer is the minimal stable nucleus. When DA/NE molecules bind to the fibril-like trimer and tetramer, they strongly destabilize the αS protofibrils by disrupting the ß-sheet structure and inter-chain E46-K80 salt bridges. Two common binding sites are identified for both DA and NE molecules on αS oligomers: residues 57-70 and 81-83. A different binding site is also observed, which is located at the N-terminal region (residues 45-52). The binding of DA/NE molecules to αS oligomers is mostly driven by hydrophobic and electrostatic interactions. We found two disruptive modes, and binding to the turn region of αS oligomers but disrupting the adjacent ß-sheet structure is the dominant one. Our work identified the critical nucleus of Greek-key-like core of αS protofibrils and revealed the disruptive mechanism of αS protofibrils by DA/NE molecules, which may be helpful to the design of effective drugs against αS aggregation.

Proline hydroxylation at different sites in hypoxia-inducible factor 1α modulates its interactions with the von Hippel-Lindau tumor suppressor protein.

Hypoxia-inducible factor 1 (HIF-1) plays an essential role in the regulation of hypoxia in humans. This regulation is mediated by the interaction of the von Hippel-Lindau tumor suppressor protein (pVHL) with the hydroxylated HIF-1α at proline564 (Pro564). Experimental studies reported that Pro567 could also be hydroxylated. However, the conformational dynamics of the complex of pVHL with hydroxylated HIF-1α at Pro564 is not well understood, and whether hydroxylated Pro567 plays the similar essential role as Pro564 in regulating HIF-1α-pVHL interaction remains elusive. Herein, we performed all-atom molecular dynamics (MD) simulations on the pVHL/HIF-1α complexes with single hydroxylation at Pro564 and Pro567, double hydroxylation at both Pro564 and Pro567, and without hydroxylation. Our multiple MD simulations and binding energy calculations show that hydroxylation at Pro567 is less favorable for the binding of HIF-1α to pVHL, whereas hydroxylation at Pro564 results in an increase of structural rigidity of the pVHL/HIF-1α complex and an enhancement of the interactions between HIF-1α and pVHL. The different roles revealed here for Pro564 and Pro567 in regulating HIF-1α-pVHL interactions, together with the previous finding that HIF-prolyl hydroxylase PHD-3 participates in a negative feedback loop controlling the HIF-1 level, suggest that hydroxylated HIF-1α at Pro567 may perturb or may not participate in this negative feedback loop. Intriguingly, our simulation data and community network analysis demonstrate that the binding of hydroxylated HIF-1α at Pro564 to the ß-domain of pVHL allosterically induces the conformational change of the α-domain via an optimal communication pathway from Pro564 of HIF-1α to S168 of the pVHL α-domain. This study reveals the different roles of Pro564 and Pro567 hydroxylation in HIF-1α in HIF-1α-pVHL interactions, which will be beneficial for developing effective strategies to treat hypoxia-related diseases and understanding the molecular basis of hypoxic training/exercise.

Human Neuronal Calcium Sensor-1 Protein Avoids Histidine Residues To Decrease pH Sensitivity.

pH is highly regulated in mammalian central nervous systems. Neuronal calcium sensor-1 (NCS-1) can interact with numerous target proteins. Compared to that in the NCS-1 protein of Caenorhabditis elegans, evolution has avoided the placement of histidine residues at positions 102 and 83 in the NCS-1 protein of humans and Xenopus laevis, possibly to decrease the conformational sensitivity to pH gradients in synaptic processes. We used all-atom molecular dynamics simulations to investigate the effects of amino acid substitutions between species on human NCS-1 by substituting Arg102 and Ser83 for histidine at neutral (R102H and S83H) and acidic pHs (R102Hp and S83Hp). Our cumulative 5 µs simulations revealed that the R102H mutation slightly increases the structural flexibility of loop L2 and the R102Hp mutation decreases protein stability. Community network analysis illustrates that the R102H and S83H mutations weaken the interdomain and strengthen the intradomain communications. Secondary structure contents in the S83H and S83Hp mutants are similar to those in the wild type, whereas the global structural stabilities and salt-bridge probabilities decrease. This study highlights the conformational dynamics effects of the R102H and S83H mutations on the local structural flexibility and global stability of NCS-1, whereas protonated histidine decreases the stability of NCS-1. Thus, histidines at positions 102 and 83 may not be compatible with the function of NCS-1 whether in the neutral or protonated state.