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1.
J Chem Phys ; 151(8): 085101, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31470695

RESUMO

Periodic molecular dynamics simulations of proteins may suffer from image interactions. Similarly, the hydrophobic effect required to keep a protein folded may not be enforced by small simulation cells. Accordingly, errors may arise both from the water concentration per se and the image interactions. Intrinsically disordered proteins are particularly sensitive, providing a worst-case estimate of the errors. Following this reasoning, we studied Aß40 (Aß), a disordered peptide central to Alzheimer's disease, by 100 different simulations with variable cell size from very large (20 Å) to very small (3 Å). Even for this very disordered peptide, most properties are not cell-size dependent, justifying the common use of modest-sized (10 Å) cells for simulating proteins. The radius of gyration, secondary structure, intrapeptide, and peptide-water hydrogen bonds are similar relative to standard deviations at any cell size. However, hydrophobic surface area increases significantly in small cells (confidence 95%, two-tailed t-test), as does the standard deviation in exposure and backbone conformations (>40% and >27%). Similar results were obtained for the force fields OPLS3e, Ambersb99-ILDN, and Charmm22*. The similar prevalence of structures and α-ß transitions in long and short simulations indicate small diffusion barriers, which we suggest is a defining hallmark of intrinsically disordered proteins. Whereas hydrophilic exposure dominates in large cells, hydrophobic exposure dominates in small cells, suggesting a weakening of the hydrophobic effect by image interactions and the few water layers available to keep the protein compact, with a critical limit of 2-3 water layers required to enforce the hydrophobic effect.


Assuntos
Peptídeos beta-Amiloides/química , Tamanho Celular , Proteínas Intrinsicamente Desordenadas/química , Simulação de Dinâmica Molecular , Humanos , Interações Hidrofóbicas e Hidrofílicas , Conformação Proteica
2.
Phys Chem Chem Phys ; 21(37): 20999-21006, 2019 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-31528872

RESUMO

A substantial number of diseases leading to loss of neurologic functions such as Morbus Alzheimer, Morbus Parkinson, or Chorea Huntington are related to the fibrillation of particular amyloidogenic peptides. In vitro amyloid fibrillation strongly depends on admixture with other proteins and peptides, lipids, nanoparticles, surfactants and polymers. We investigated amyloid-beta 1-40 peptide (Aß1-40) fibrillation in mixture with thermoresponsive poly(oligo(ethylene glycol)macrylates), in which the polymer's hydrophobicity is tuned by variation of the number of ethylene glycol-units in the side chain (m = 1-9), the end groups (B = butoxy; C = carboxy; D = dodecyl; P = pyridyldisulfide) and the degree of polymerization (n) of the polymers. The polymers were prepared via RAFT-polymerization, obtaining a broad range of molecular masses (Mn = 700 to 14 600 g mol-1 kDa-1, polydispersity indices PDI = 1.10 to 1.25) and tunable cloud point temperatures (Tcp), ranging from 42.4 °C to 80 °C, respectively. Proper combination of hydrophobic end groups with hydrophilic side chains of the polymer allowed to alter the hydrophilicity/hydrophobicity of these polymers, which is shown to enhance Aß1-40 aggregation significantly in case of the endgroup D (with n = 16, 23, 56). We observed that the less hydrophilic polymers (m = 1-2) were able to both decrease and elongate the lag (tlag) and characteristic times (tchar) of Aß1-40 fibril formation in dependence of their end groups, molecular mass and hydrophilicity. On the other hand, highly hydrophilic polymers (m = 3, 5, 9) either decreased, or only marginally influenced the lag and characteristic times of Aß1-40 fibrillation, in all cases forming ß-sheet rich fibrils as observed by TEM and CD-spectroscopy. Our results support that balanced hydrophobic and hydrophilic interactions of a polymer with Aß1-40 is important for inhibiting amyloid-formation pathways.


Assuntos
Peptídeos beta-Amiloides/química , Fragmentos de Peptídeos/química , Polímeros/química , Amiloide/química , Amiloide/ultraestrutura , Peptídeos beta-Amiloides/ultraestrutura , Interações Hidrofóbicas e Hidrofílicas , Fragmentos de Peptídeos/ultraestrutura
3.
Chem Commun (Camb) ; 55(59): 8595-8598, 2019 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-31276123

RESUMO

The amino acid sequence plays an essential role in amyloid formation. Here, using the central core recognition module of the Aß peptide and its reverse sequence, we show that although both peptides assemble into ß-sheets, their morphologies, kinetics and cell toxicities display marked differences. In addition, the native peptide, but not the reverse one, shows notable affinity towards bilayer lipid model membranes that modulates the aggregation pathways to stabilize the oligomeric intermediate states and function as the toxic agent responsible for neuronal dysfunction.


Assuntos
Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/metabolismo , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Sequência de Aminoácidos , Peptídeos beta-Amiloides/toxicidade , Animais , Linhagem Celular Tumoral , Colesterol/química , Humanos , Cinética , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Fragmentos de Peptídeos/toxicidade , Fosfatidilcolinas/química , Conformação Proteica em Folha beta , Multimerização Proteica , Ratos , Esfingomielinas/química
4.
Phys Chem Chem Phys ; 21(28): 15686-15694, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31271401

RESUMO

Alzheimer's disease is associated with the abnormal self-assembly of amyloid-ß (Aß) peptide into toxic oligomers and fibrils. Recent experiments reported that Aß16-22, containing the central hydrophobic core (CHC) of Aß, formed antiparallel ß-sheet fibrils, while its E22Q mutant self-assembled into parallel ß-sheet fibrils. However, the molecular mechanisms underlying E22Q-mutation-induced parallel ß-sheet fibril formation are not well understood. Herein, we performed molecular dynamics (MD) simulations to study the dimerization processes of Aß16-22 and Aß16-22E22Q peptides. ß-Sheet dimers with diverse hydrogen bond arrangements were observed and they exhibited highly dynamic and interconverting properties. An antiparallel-to-parallel ß-sheet transition occurred in the assembly process of the E22Q mutant, but not in that of Aß16-22. During this conformational transformation process, the inter-molecular Q22-Q22 hydrogen bonds were first formed and acted as a binder to facilitate the two chains forming a parallel orientation, then the hydrophobic interactions between residues in the CHC region consolidated this arrangement and drove the main-chain H-bond formation, hence resulting in parallel ß-sheet formation. However, parallel ß-sheets were less populated than antiparallel ß-sheets of Aß16-22E22Q dimers. In order to explore whether parallel ß-sheets became dominant in larger size oligomers, we investigated the conformational ensembles of Aß16-22 and Aß16-22E22Q octamers by conducting replica exchange molecular dynamics (REMD) simulations. The REMD simulations revealed that the population of parallel ß-strand alignment increased with an increase of the size of ordered Aß16-22E22Q ß-sheet oligomers, implying that the formation of full parallel ß-sheets requires larger sized oligomers. Our findings provide a mechanistic explanation for the E22Q-mutation-induced formation of parallel ß-sheet fibrils observed experimentally.


Assuntos
Simulação de Dinâmica Molecular , Mutação , Conformação Proteica em Folha beta/genética , Amiloide/química , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/genética , Humanos , Conformação Proteica
5.
Chemistry ; 25(51): 11852-11858, 2019 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-31361361

RESUMO

Proteolysis of amyloid-ß (Aß) is a promising approach against Alzheimer's disease. However, it is not feasible to employ natural hydrolases directly because of their cumbersome preparation and purification, poor stability, and hazardous immunogenicity. Therefore, artificial enzymes have been developed as potential alternatives to natural hydrolases. Since specific cleavage sites of Aß are usually embedded inside the ß-sheet structures that restrict access by artificial enzymes, this strongly hinders their efficiency for practical applications. Herein, we construct a NIR (near-IR) controllable artificial metalloprotease (MoS2 -Co) using a molybdenum disulfide nanosheet (MoS2 ) and a cobalt complex of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (Codota). Evidenced by detailed experimental and theoretical studies, the NIR-enhanced MoS2 -Co can circumvent the restriction by simultaneously inhibition of ß-sheet formation and destroying ß-sheet structures of the preformed Aß aggregates in living cell. Furthermore, our designed MoS2 -Co is an easy to graft Aß-target agent that prevents misdirected or undesirable hydrolysis reactions, and has been demonstrated to cross the blood brain barrier. This method can be adapted for hydrolysis of other kinds of amyloids.


Assuntos
Peptídeos beta-Amiloides/química , Barreira Hematoencefálica/metabolismo , Dissulfetos/química , Metaloproteases/química , Molibdênio/química , Doença de Alzheimer , Barreira Hematoencefálica/química , Humanos , Metaloproteases/metabolismo , Espectrofotometria Infravermelho
6.
Chem Biol Interact ; 309: 108707, 2019 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-31194956

RESUMO

Alzheimer's disease (AD) is a slow but progressive neurodegenerative disease. One of the pathological hallmarks of AD is the progressive accumulation of ß-amyloid (Aß) in the form of senile plaques, and Aß insult to neuronal cells has been identified as one of the major causes of AD onset. In the present study, we investigated the anti-AD potential of four flavonoids, naringenin, didymin, prunin, and poncirin, by evaluating their ability to inhibit acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and ß-site amyloid precursor protein cleaving enzyme 1 (BACE1). All four flavonoids displayed promising inhibitory activity against AChE, BChE, and BACE1. Structure-activity relationships suggested that glycosylation of naringenin at sugar moieties, and at different positions of the glycosidic linkage, might be closely associated with anti-AD potential. Kinetic and docking studies showed the lowest binding energy and highest affinity for the mixed, competitive, and non-competitive type inhibitors didymin, prunin, and poncirin. Hydrophobic interactions and the number of hydrogen bonds determined the strength of the protein-inhibitor interaction. We also examined the neuroprotective mechanisms by which flavonoids act against Aß25-35-induced toxicity in PC12 cells. Exposure of PC12 cells to 10 µM Aß25-35 for 24 h resulted in a significant decrease in cell viability. In addition, pretreatment of PC12 cells with different concentrations of flavonoids for 1 h significantly reversed the effects of Aß. Furthermore, treatment with the most active flavonoid, didymin, significantly reduced BACE1, APPsß, and C99 expression levels in a dose-dependent manner, without affecting amyloid precursor protein (APP) levels in the amyloidogenic pathway. Together, our results indicate that flavonoids, and in particular didymin, exhibit inhibitory activity in vitro, and may be useful in the development of therapeutic modalities for the treatment of AD.


Assuntos
Acetilcolinesterase/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Ácido Aspártico Endopeptidases/metabolismo , Butirilcolinesterase/metabolismo , Flavanonas/química , Glicosídeos/farmacologia , Agregados Proteicos/efeitos dos fármacos , Acetilcolinesterase/química , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/farmacologia , Animais , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Sítios de Ligação , Butirilcolinesterase/química , Domínio Catalítico , Sobrevivência Celular/efeitos dos fármacos , Glicosídeos/química , Cinética , Simulação de Acoplamento Molecular , Células PC12 , Fragmentos de Peptídeos/farmacologia , Substâncias Protetoras/química , Substâncias Protetoras/farmacologia , Ratos , Relação Estrutura-Atividade
7.
Eur J Med Chem ; 177: 247-258, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31158742

RESUMO

Alzheimer's disease (AD) is a chronic, fatal and complex neurodegenerative disorder, which is characterized by cholinergic system dysregulation, metal dyshomeostasis, amyloid-ß (Aß) aggregation, etc. Therefore in most cases, single-target or single-functional agents are insufficient to achieve the desirable effect against AD. Multi-Target-Directed Ligand (MTDL), which is rationally designed to simultaneously hit multiple targets to improve the pharmacological profiles, has been developed as a promising approach for drug discovery against AD. To identify the multifunctional agents for AD, we developed an innovative method to successfully conceal the metal chelator into acetylcholinesterase (AChE) inhibitor. Briefly, the "hidden" agents first cross the Blood Brain Barrier (BBB) to inhibit the function of AChE, and the metal chelator will then be released via the enzymatic hydrolysis by AChE. Therefore, the AChE inhibitor, in this case, is not only a single-target agent against AD, but also a carrier of the metal chelator. In this study a total of 14 quinoline derivatives were synthesized and biologically evaluated. Both in vitro and in vivo results demonstrated that compound 9b could cross the BBB efficiently, then release 8a, the metabolite of 9b, into brain. In vitro, 9b had a potent AChE inhibitory activity, while 8a displayed a significant metal ion chelating function, therefore in combination, both 9b and 8a exhibited a considerable inhibition of Aß aggregation, one of the observations that plays important roles in the pathogenesis of AD. The efficacy of 9b against AD was further investigated in both a zebrafish model and two different mice models.


Assuntos
Quelantes/farmacologia , Inibidores da Colinesterase/farmacologia , Nootrópicos/farmacologia , Quinolinas/farmacologia , Acetilcolinesterase/química , Acetilcolinesterase/metabolismo , Peptídeos beta-Amiloides/química , Animais , Barreira Hematoencefálica/metabolismo , Quelantes/síntese química , Quelantes/farmacocinética , Quelantes/toxicidade , Inibidores da Colinesterase/síntese química , Inibidores da Colinesterase/farmacocinética , Inibidores da Colinesterase/toxicidade , Desenho de Drogas , Canal de Potássio ERG1/antagonistas & inibidores , Masculino , Camundongos , Microssomos Hepáticos/metabolismo , Simulação de Acoplamento Molecular , Estrutura Molecular , Nootrópicos/síntese química , Nootrópicos/farmacocinética , Nootrópicos/toxicidade , Fragmentos de Peptídeos/química , Multimerização Proteica/efeitos dos fármacos , Quinolinas/síntese química , Quinolinas/farmacocinética , Quinolinas/toxicidade , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Peixe-Zebra
8.
J Chem Phys ; 150(22): 225101, 2019 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-31202253

RESUMO

Understanding the key factors that govern the rate of protein aggregation is of immense interest since protein aggregation is associated with a number of neurodegenerative diseases. Previous experimental and theoretical studies have revealed that the hydrophobicity, charge, and population of the fibril-prone monomeric state control the fibril formation rate. Because the fibril structures consist of cross beta sheets, it is widely believed that those sequences that have a high beta content (ß) in the monomeric state should have high aggregation rates as the monomer can serve as a template for fibril growth. However, this important fact has never been explicitly proven, motivating us to carry out this study. Using replica exchange molecular dynamics simulation with implicit water, we have computed ß of 19 mutations of amyloid beta peptide of 42 residues (Aß42) for which the aggregation rate κ has been measured experimentally. We have found that κ depends on ß in such a way that the higher the propensity to aggregation, the higher the beta content in the monomeric state. Thus, we have solved a long-standing problem of the dependence of fibril formation time of the ß-structure on a quantitative level.


Assuntos
Peptídeos beta-Amiloides/química , Fragmentos de Peptídeos/química , Multimerização Proteica , Peptídeos beta-Amiloides/genética , Interações Hidrofóbicas e Hidrofílicas , Simulação de Dinâmica Molecular , Mutação , Fragmentos de Peptídeos/genética , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Termodinâmica
9.
Nanoscale ; 11(18): 9185-9193, 2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31038146

RESUMO

Recent advances in nanotechnology have developed a lot of opportunities for biological applications. In this work, multifunctional MoS2/AuNR nanocomposites with unique high NIR absorption were designed via combining MoS2 nanosheets and gold nanorods (AuNRs). The nanocomposites were synthesized through electrostatic self-assembly and showed high stability and good biocompatibility. Then they were used to modulate the aggregation of amyloid-ß peptides, destabilize mature fibrils under NIR irradiation, and eliminate Aß-induced ROS against neurotoxicity. The inhibition and destabilization effects were confirmed by Thioflavin T (ThT) fluorescence assay and transmission electron microscopy (TEM). Cell viability assay and ROS assay revealed that MoS2/AuNR nanocomposites could alleviate Aß-induced oxidative stress and cell toxicity. More importantly, both MoS2 nanosheets and AuNRs can be used as NIR photothermal agents, MoS2/AuNR nanocomposites have enhanced ability of disrupting Aß fibrils and improved cell viability by generating local heat under low power NIR irradiation. Our results provide new insights into the design of new multifunctional systems for the treatment of amyloid-related diseases.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Raios Infravermelhos , Nanocompostos/química , Fragmentos de Peptídeos/metabolismo , Amiloide/química , Amiloide/toxicidade , Peptídeos beta-Amiloides/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Dissulfetos/química , Ouro/química , Humanos , Molibdênio/química , Nanocompostos/toxicidade , Nanotubos/química , Fragmentos de Peptídeos/química , Espécies Reativas de Oxigênio/metabolismo , Espectroscopia de Infravermelho com Transformada de Fourier
10.
Dalton Trans ; 48(21): 7451-7461, 2019 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-31086893

RESUMO

A significant abundance of copper (Cu) and iron in amyloid ß (Aß) plaques, and several heme related metabolic disorders are directly correlated with Alzheimer's disease (AD), and these together with co-localization of Aß plaques with heme rich deposits in the brains of AD sufferers indicates a possible association of the said metals with the disease. Recently, the Aß peptides have been found to bind heme and Cu individually as well as simultaneously. Another significant finding relevant to this is the lower levels of nitrite and nitrate found in the brains of patients suffering from AD. In this study, a combination of absorption and electron paramagnetic resonance spectroscopy and kinetic assays have been used to study the interaction of nitrite with the metal bound Aß complexes. The data indicate that heme(III)-Cu(i)-Aß, heme(II)-Cu(i)-Aß, heme(II)-Aß and Cu(i)-Aß can reduce nitrite to nitric oxide (NO), an important biological messenger also related to AD, and thus behave as nitrite reductases. However these complexes reduce nitrite at different rates with heme(III)-Cu(i)-Aß being the fastest following an inner sphere electron transfer mechanism. The rest of the metal-Aß adducts follow an outer sphere electron transfer mechanism during nitrite reduction. Protonation from the Arg5 residue triggering the N-O bond heterolysis in heme(III) bound nitrite with a simultaneous electron transfer from the Cu(i) center to produce NO is the rate determining step, indicating a proton transfer followed by electron transfer (PTET) mechanism.


Assuntos
Peptídeos beta-Amiloides/química , Cobre/química , Heme/química , Nitrito Redutases/química , Espectroscopia de Ressonância de Spin Eletrônica , Transporte de Elétrons , Humanos , Ferro/química , Cinética , Óxido Nítrico/química , Nitritos/química , Oxirredução , Placa Amiloide/química
11.
Int J Mol Sci ; 20(9)2019 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-31071995

RESUMO

Human apolipoprotein E (apoE) is a major component of lipoprotein particles, and under physiological conditions, is involved in plasma cholesterol transport. Human apolipoprotein E found in three isoforms (E2; E3; E4) is a member of a family of apolipoproteins that under pathological conditions are detected in extracellular amyloid depositions in several amyloidoses. Interestingly, the lipid-free apoE form has been shown to be co-localized with the amyloidogenic Aß peptide in amyloid plaques in Alzheimer's disease, whereas in particular, the apoE4 isoform is a crucial risk factor for late-onset Alzheimer's disease. Evidence at the experimental level proves that apoE self-assembles into amyloid fibrilsin vitro, although the misfolding mechanism has not been clarified yet. Here, we explored the mechanistic insights of apoE misfolding by testing short apoE stretches predicted as amyloidogenic determinants by AMYLPRED, and we computationally investigated the dynamics of apoE and an apoE-Αß complex. Our in vitro biophysical results prove that apoE peptide-analogues may act as the driving force needed to trigger apoE aggregation and are supported by the computational apoE outcome. Additional computational work concerning the apoE-Αß complex also designates apoE amyloidogenic regions as important binding sites for oligomeric Αß; taking an important step forward in the field of Alzheimer's anti-aggregation drug development.


Assuntos
Doença de Alzheimer/genética , Peptídeos beta-Amiloides/química , Amiloidose/genética , Apolipoproteínas E/química , Doença de Alzheimer/patologia , Amiloide/química , Amiloide/genética , Amiloide/ultraestrutura , Peptídeos beta-Amiloides/genética , Peptídeos beta-Amiloides/ultraestrutura , Amiloidose/patologia , Apolipoproteínas E/ultraestrutura , Sítios de Ligação , Colesterol/química , Colesterol/genética , Humanos , Placa Amiloide/genética , Placa Amiloide/patologia , Placa Amiloide/ultraestrutura , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/patologia , Dobramento de Proteína , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/ultraestrutura
12.
J Chromatogr A ; 1601: 350-356, 2019 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-31101465

RESUMO

This study reports a capillary isotachophoresis (ITP) - electrospray ionization mass spectrometry (ESI-MS) method for the determination of several amyloid ß (Aß) peptides, which are biomarkers of Alzheimer's disease (AD) in cerebrospinal fluids (CSF). For the first time, these peptides have been detected directly from CSF by MS without recourse to an immunocapture-based sample pre-treatment. The antibody-free approach is based on the marriage between capillary ITP, a powerful on-line electrokinetic preconcentration technique, and MS for simultaneous detection of different Aß peptides. To ensure a good performance, the ITP process of fluorescently labelled Aß peptides was for the first time implemented and verified with laser induced fluorescent detection, prior to methodology transfer to MS detection. Better detection sensitivity was achieved with labelled Aß peptides for both detection modes. Using hydroxyl ions as the terminating and acetate as the leading ions, our method allows efficient ITP preconcentration under alkaline conditions of the slowly migrating Aß peptides to reach quantifiable concentration down to 50 pM. The developed ITP-MS approach allows reliable quantification of different fluorescently derivatized Aß peptides, i.e. Aß 1-42, Aß 1-40 and Aß 1-38 down to sub nM ranges in CSF samples from AD and non-demented (healthy control) patients. Good agreement (<20% deviation) for the determination of Aß 1-42/Aß 1-40 ratio in CSF was achieved between results obtained with this new ITP-MS and our recently developed method based on large volume sample stacking coupled to CE. Discrimination of one AD patient from two healthy controls was successfully made with the Aß 1-42/Aß 1-40 ratio obtained by the developed ITP-MS method for the tested cerebrospinal fluid samples.


Assuntos
Doença de Alzheimer/líquido cefalorraquidiano , Peptídeos beta-Amiloides/líquido cefalorraquidiano , Biomarcadores/líquido cefalorraquidiano , Isotacoforese , Espectrometria de Massas por Ionização por Electrospray , Idoso , Idoso de 80 Anos ou mais , Peptídeos beta-Amiloides/química , Feminino , Humanos , Masculino
13.
Chemphyschem ; 20(13): 1680-1689, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-31087613

RESUMO

We employed deuterium solid-state NMR techniques under static conditions to discern the details of the µs-ms timescale motions in the flexible N-terminal subdomain of Aß1-40 amyloid fibrils, which spans residues 1-16. In particular, we utilized a rotating frame (R1ρ ) and the newly developed time domain quadrupolar Carr-Purcell-Meiboom-Gill (QCPMG) relaxation measurements at the selectively deuterated side chains of A2, H6, and G9. The two experiments are complementary in terms of probing somewhat different timescales of motions, governed by the tensor parameters and the sampling window of the magnetization decay curves. The results indicated two mobile "free" states of the N-terminal domain undergoing global diffusive motions, with isotropic diffusion coefficients of 0.7-1 ⋅ 108 and 0.3-3 ⋅ 106 ad2 s-1 . The free states are also involved in the conformational exchange with a single bound state, in which the diffusive motions are quenched, likely due to transient interactions with the structured hydrophobic core. The conformational exchange rate constants are 2-3 ⋅ 105  s-1 and 2-3 ⋅ 104  s-1 for the fast and slow diffusion free states, respectively.


Assuntos
Peptídeos beta-Amiloides/química , Amiloide/química , Fragmentos de Peptídeos/química , Deutério , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Domínios Proteicos
14.
Comput Biol Chem ; 80: 259-269, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31048244

RESUMO

In order to study the effects of peptide exposure to oxidative attack, we chose a model reaction in which the hydroxyl radical discretely abstracts a hydrogen atom from the α-carbon of each residue of a highly amyloidogenic region in the human calcitonin hormone, hCT15-19. Based on a combined Molecular Mechanics / Quantum Mechanics approach, the extended and folded L- and D-configuration and radical intermediate hCT15-19 peptides were optimized to obtain their compactness, secondary structure and relative thermodynamic data. The results suggest that the epimerization of residues is generally an exergonic process that can explain the cumulative nature of molecular aging. Moreover, the configurational inversion induced conformational changes can cause protein dysfunction. The epimerization of the central residue to the D-configuration induced a hairpin structure in hCT15-19, concomitant with a possible oligomerization of human calcitonin into Aß(1-42)-like amyloid fibrils present in patients suffering from Alzheimer's disease.


Assuntos
Proteínas Amiloidogênicas/química , Calcitonina/química , Fragmentos de Peptídeos/química , Peptídeos beta-Amiloides/química , Teoria da Densidade Funcional , Humanos , Ligações de Hidrogênio , Modelos Químicos , Simulação de Dinâmica Molecular , Oxirredução , Estrutura Secundária de Proteína , Estereoisomerismo , Termodinâmica
15.
J Chem Phys ; 150(18): 185102, 2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31091893

RESUMO

Studies have found strong correlations between polymorphism and structural variations in amyloid-ß (Aß) fibrils and the diverse clinical subtypes of Alzheimer's disease (AD). Thus, a detailed understanding of the conformational behavior of Aß fibrils may be an aid to elucidate the pathological mechanisms involved in AD. However, a key point that has been inadvertently underestimated or dismissed is the role of the protonated state at the C-terminal residue of amyloid-ß peptides, which can give rise to intrinsic differences in the morphology and stability of the fibrils. For instance, the effects of the salt bridge formed between the C-terminal residue A42 and the residue K28 on the S-shaped Aß protofibril structure remain unknown and may be different from those in the U-shaped Aß protofibril structures. To address this effect, we explore the stability of the S-shaped protofibrils capped with different C-terminal modifications, including carboxyl group in its deprotonated (COO-) and protonated (COOH) states, by using molecular dynamics simulations. Our findings indicated that the C-terminal deprotonated protofibril is significantly more stable than its C-terminal protonated counterpart due to a well-defined and highly stable zipper-like salt-bridge-chain formed by the ε-NH3 + groups on the sidechain of residue K28 and the C-terminal COO- group at the A42 residue. The revealed underlying molecular mechanism for the different stability of the protofibrils provides insights into the diversity of polymorphism in Aß fibrils.


Assuntos
Peptídeos beta-Amiloides/química , Fragmentos de Peptídeos/química , Modelos Químicos , Simulação de Dinâmica Molecular , Conformação Proteica , Estabilidade Proteica , Prótons
16.
Protein Pept Lett ; 26(7): 502-511, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30950343

RESUMO

BACKGROUND: Amyloid fibrils in Alzheimer's disease are composed of amyloid-ß (Aß) peptides of variant lengths. Humanin (HN), a 24 amino acid residue neuroprotective peptide, is known to interact with the predominant Aß isoform in the brain, Aß (1-40). METHODS: Here, we constructed smaller segments of Aß and HN and identified residues in HN important for both HN-HN and HN-Aß interactions. Peptides corresponding to amino acid residues 5- 15 of HN, HN (5-15), HN (5-15, L11S), where Leu11 was replaced with Ser, and residues 17-28 of Aß, Aß (17-28), were synthesized and tested for their ability to block formation of the complex between HN and Aß (1-40). RESULTS: Co-immunoprecipitation and binding kinetics showed that HN (5-15) was more efficient at blocking the complex between HN and Aß (1-40) than either HN (5-15, L11S) or Aß (17-28). Binding kinetics of these smaller peptides with either full-length HN or Aß (1-40) showed that HN (5- 15) was able to bind either Aß (1-40) or HN more efficiently than HN (5-15, L11S) or Aß (17-28). Compared to full-length HN, however, HN (5-15) bound Aß (1-40) with a weaker affinity suggesting that while HN (5-15) binds Aß, other residues in the full length HN peptide are necessary for maximum interactions. CONCLUSION: L11 was more important for interactions with Aß (1-40) than with HN. Aß (17-28) was relatively ineffective at binding to either Aß (1-40) or HN. Moreover, HN, and the smaller HN (5-15), HN (5-15 L11S), and Aß (17-28) peptides, had different effects on regulating Aß (1-40) aggregation kinetics.


Assuntos
Peptídeos beta-Amiloides/química , Peptídeos e Proteínas de Sinalização Intracelular/química , Fragmentos de Peptídeos/química , Sequência de Aminoácidos , Humanos , Cinética , Conformação Proteica
17.
Nutrients ; 11(4)2019 Mar 31.
Artigo em Inglês | MEDLINE | ID: mdl-30935135

RESUMO

Some polyphenols, which are common natural compounds in fruits, vegetables, seeds, and oils, have been considered as potent inhibitors of amyloid beta (Aß) aggregation, one critical pathogenic event in Alzheimer's disease (AD). However, the mechanisms by which polyphenols affect aggregation are not fully understood. In this study, we aimed to investigate the effect of two classes of polyphenols (flavonoids and stilbenes) on the self-assembly of Aß_42, in particular, how this relates to structure. We found that the flavonoids gallocatechin gallate (GCG) and theaflavin (TF) could completely inhibit Aß aggregation, while two stilbenes, resveratrol and its glucoside derivative piceid, could also suppress Aß aggregation, but to a much lesser extent. Intriguingly, resveratrol accelerated the formation of Aß fibrils before its decreasing effect on fibrillation was detected. Atomic force microscopy (AFM) images showed a huge mass of long and thin Aß fibrils formed in the presence of resveratrol. Although the morphology was the same in the presence of piceid, the fibrils were sparse in the presence of picead. In the presence of flavonoids, Aß morphology was unchanged from prior to incubation (0 h), in agreement with amyloid beta kinetics analysis using thioflavin-T fluorescence assay. The electrochemical data showed a higher ability of GCG and TF to interact with Aß than resveratrol and piceid, which could be attributed to the presence of more aromatic rings and hydroxyl groups. In addition, the two flavonoids exhibited a similar propensity for Aß aggregation, despite having some differences in their structure. However, in the case of stilbenes, the addition of a glucoside at C-7 slightly decreased anti-Aß aggregation property compared to resveratrol. These findings contribute to a better understanding of the essential structural features of polyphenols required for inhibiting Aß aggregation, and the possible mechanisms for modulating aggregation.


Assuntos
Peptídeos beta-Amiloides/química , Polifenóis/química , Agregados Proteicos , Amiloide/química , Antioxidantes/farmacologia , Microscopia de Força Atômica , Conformação Proteica
18.
Biosens Bioelectron ; 133: 183-191, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30928737

RESUMO

Alzheimer's disease (AD) is a chronic central neurodegenerative disease. The pathological features of AD are the extracellular deposition of senile plaques formed by amyloid-ß oligomers (AßOs) and the intracellular accumulation of neurofibrillary tangles. However, due to the lack of effective method and experimental models to study the cognitive decline, communication at cell resolution and the implementation of interventions, the diagnosis and treatment on AD still progress slowly. In this paper, we established a pathological model of AD in vitro based on AßOs-induced hippocampal neuronal network chip for multi-site dynamic analysis of the neuronal electrical activity and network connection. The multiple characteristic parameters, including positive and negative spike intervals, firing rate and peak-to-peak values, were extracted through the analysis of spike signals, and two firing patterns from the interneurons and pyramidal neurons were recorded. The spatial firing patterns mapping and cross-correlation between channels were performed to validate the degeneration of neuronal network connectivity. Moreover, an electrical stimulation with frequency at 40 Hz was exerted to preliminarily explore the therapeutic effect on the pathological model of AD. This neuronal network chip enables the implementation of AD models in vitro for studying basic mechanisms of neurodegeneration within networks and for the parallel testing of various potential therapies. It can be a novel technique in the research of AD pathological model in vitro.


Assuntos
Doença de Alzheimer/diagnóstico , Peptídeos beta-Amiloides/isolamento & purificação , Técnicas Biossensoriais , Sistemas Microeletromecânicos/métodos , Peptídeos beta-Amiloides/química , Estimulação Elétrica , Eletrólitos/química , Hipocampo/química , Hipocampo/efeitos da radiação , Humanos , Interneurônios/química , Interneurônios/efeitos da radiação , Dispositivos Lab-On-A-Chip , Rede Nervosa/química , Rede Nervosa/efeitos da radiação , Emaranhados Neurofibrilares/química , Emaranhados Neurofibrilares/efeitos da radiação , Células Piramidais/química , Células Piramidais/efeitos da radiação
19.
Biosens Bioelectron ; 133: 192-198, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30928738

RESUMO

A highly efficient quench-type electrochemiluminescence (ECL) immunosensor was proposed for the trace detection of amyloid ß-protein (Aß1-42). In this work, tin disulfide nanoflowers (SnS2 NFs) with large specific surface area, favorable catalytic property and chemical stability were prepared and used as substrate material. Taking advantage of the excellent catalytic ability and biocompatibility, palladium nanoparticles (Pd NPs) were in situ reduced on SnS2 NFs to obtain SnS2-Pd. Moreover it could combine with large amounts of luminol and achieve a strong ECL signal output. In addition, copper doped mesoporous tungsten trioxide (Cu:WO3) nanoparticles were selected to quench ECL emission of luminol@SnS2-Pd via resonance energy transfer, where luminol@SnS2-Pd was the donor and Cu:WO3 was the acceptor. On this basis, a quench-type ECL immunosensor was constructed for detection of Aß1-42. Under optimum conditions, the fabricated ECL immunosensor showed sensitive response to Aß1-42 concentration from 0.1 pg/mL to 50 ng/mL with a low detection limit of 5.4 fg/mL (S/N = 3). It is expected to be a promising analytical tool for the sensitive detection of Aß1-42 and other biomarkers with high specificity, good reproducibility and long-term stability.


Assuntos
Peptídeos beta-Amiloides/isolamento & purificação , Biomarcadores/química , Técnicas Biossensoriais , Nanopartículas Metálicas/química , Peptídeos beta-Amiloides/química , Catálise , Cobre/química , Dissulfetos/química , Técnicas Eletroquímicas , Glucose Oxidase/química , Ouro/química , Humanos , Limite de Detecção , Luminol/química , Paládio/química , Estanho/química
20.
J Mol Model ; 25(5): 124, 2019 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-31020417

RESUMO

Besides their biomolecular relevance, amyloids, generated by the self-assembly of peptides and proteins, are highly organized structures useful for nanotechnology applications. The introduction of halogen atoms in these peptides, and thus the possible formation of halogen bonds, allows further possibilities to finely tune the amyloid nanostructure. In this work, we performed molecular dynamics simulations on different halogenated derivatives of the ß-amyloid peptide core-sequence KLVFF, by using a modified AMBER force field in which the σ-hole located on the halogen atom is modeled with a positively charged extra particle. The analysis of equilibrated structures shows good agreement with crystallographic data and experimental results, in particular concerning the formation of halogen bonds and the stability of the supramolecular structures. The modified force field described here allows describing the atomistic details contributing to peptides aggregation, with particular focus on the role of halogen bonds. This framework can potentially help the design of novel halogenated peptides with desired aggregation propensity. Graphical abstract Molecular dynamics investigation of halogenated amyloidogenic peptides.


Assuntos
Peptídeos beta-Amiloides/química , Amiloide/química , Simulação de Dinâmica Molecular , Halogenação , Halogênios/química , Modelos Moleculares
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