RESUMEN
Protein aggregation is known as the main mechanism of amyloid fibrillation in amyloidosis diseases. Recent studies confirmed that compounds with one or two indole rings have inhibitory potential against amyloid fibrillation. Herein, the interaction of two similar compounds 'bis(indolyl)-2-methyl-phenyl-methene' and 'bis(indolyl)-2-chloro-phenyl-methene' with an amyloid core model was investigated. To this aim, molecular docking and all-atom molecular dynamics (MD) simulations were used. Docking results between aggregation-prone region (APR) of hen egg-white lysozyme (HEWL) and either of ligands showed that they interact with different residues of the APR (amyloid fibril nucleus). According to MD results, bis(indolyl)-2-methyl-phenyl-methene made a distance between the two cores, which was 1.5 times greater than that bis(indolyl)-2-chloro-phenyl-methene made. Analysis of RMSD/RMSF values revealed that bis(indolyl)-2-methyl-phenyl-methene stabilized strands of A and B, while destabilized strands C and D. The hydrophobic 'methyl' functional group in bis(indolyl)-2-methyl-phenyl-methene facilitate its deep penetration between core nuclei, via destabilizing outer strands of C and D. Considering this fact that results of this study are in agreement with experimental findings, details of the discovered mechanism of interaction between ligands and HEWL's APR would be inspiring for further anti-fibrillation drug designs.Communicated by Ramaswamy H. Sarma.
Asunto(s)
Amiloide , Amiloidosis , Amiloide/química , Amiloidosis/tratamiento farmacológico , Humanos , Indoles/química , Indoles/farmacología , Ligandos , Simulación del Acoplamiento Molecular , Simulación de Dinámica MolecularRESUMEN
Amyloid or similar protein aggregates are the hallmarks of many disorders, including Alzheimer's, Parkinson's, Huntington's diseases and amyloidoses. The inhibition of the formation of these aberrant species by small molecules is a promising strategy for disease treatment. However, at present, all such diseases lack an appropriate therapeutic approach based on small molecules. In this work we have evaluated five bis(indolyl)phenylmethane derivatives to reduce amyloid fibril formation by hen egg white lysozyme (HEWL) and its associated cytotoxicity. HEWL is a widely used model system to study the fundamentals of amyloid fibril formation and is heterologous to human lysozyme, which forms amyloid fibrils in a familial form of systemic amyloidosis. HEWL aggregation was tested in the presence and absence of the five compounds, under conditions in which the protein is partially unfolded. To this purpose, various techniques were used, including Congo red and Thioflavin T binding assays, atomic force microscopy, Fourier-Transform Infrared spectroscopy and cell-based cytotoxicity assays, such as the MTT reduction test and the trypan blue test. It was found that all compounds inhibited the formation of amyloid fibrils and their associated toxicity, diverging the aggregation process towards the formation of large, morphologically amorphous, unstructured, nontoxic aggregates, thus resembling class I molecules defined previously. In addition, the five compounds also appeared to disaggregate pre-formed fibrils of HEWL, which categorizes them into class IA. The half maximal inhibitory concentration (IC50) was found to be ca 12.3 ± 1.0 µM for the forefather compound.
Asunto(s)
Amiloide/química , Indoles/química , Indoles/farmacología , Muramidasa/química , Agregado de Proteínas/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Indoles/toxicidad , Cinética , Células MCF-7RESUMEN
Lipid-binding domains regulate positioning of the membrane proteins via specific interactions with phospholipid's head groups. Spinal cord injury (SCI) diminishes the integrity of neural fiber membranes at nanoscopic level. In cases that the ruptured zone size is beyond the natural resealing ability, there is a need for reinforcing factors such as polymers (e.g. Polyethylene glycol) to patch the dismantled axoplasm. Certain conserved sequential and structural patterns of interacting residues specifically bind to PEGs. It is also found that PEG600, PEG400 and PEG200 share the strongest interaction with the lipid-binding domains even more successful than phospholipid head groups. The alpha helix structure composed of hydrophobic, neutral and acidic residues prepares an opportunity for PEG400 to play an amphipathic role in the interaction with injured membrane. This in-silico study introduces a mechanism for PEG restorative ability at the molecular level. It is believed that PEG400 interrelates the injured membrane to their underneath axoplasm while retaining the integrity of ruptured membrane via interaction with ENTH domains of membrane proteins. This privilege of PEG400 in treating injured membrane must be considered in designing of polymeric biomaterials that are introduced for SCI repair.
Asunto(s)
Lípidos de la Membrana/metabolismo , Proteínas de la Membrana/metabolismo , Polietilenglicoles/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Axones/metabolismo , Simulación por Computador , Modelos Biológicos , Recuperación de la FunciónRESUMEN
Immediate membrane sealing after spinal cord injury (SCI) can prevent further degradation and result in ultimate functional recovery. It has been reported that polyethylene glycol (PEG) can repair membrane damage caused by mechanical insults to the spinal cord. Furthermore, membrane fluidity and its sealing process vary at different temperatures. Here, we have assessed the possible synergistic effects of PEG and temperature on the repair of neural membranes in an SCI model. The effects of PEGs (400, 1,000 and 2,000 Da) were studied at different temperatures (25, 37 and 40 °C) by means of compound action potential (CAP) recovery and a lactate dehydrogenase (LDH) assay. Isolated spinal cords were mounted in a double sucrose gap chamber, where the amplitude and area of CAPs were recorded after implementing injury, in the presence and absence of PEG. Moreover, the LDH assay was used to assess the effects of PEG on membrane resealing. Data showed that the least CAP recovery occurred at 25 °C, followed by 37 and 40 °C, in all treated groups. Moreover, maximum CAP amplitude recovery, 65.46 ± 5.04 %, was monitored in the presence of PEG400 at 40 °C, followed by 41.49 ± 2.41 % in PEG1000 and 37.36 ± 1.62 % in PEG2000. Furthermore, raising the temperature from 37 to 40 °C significantly increased CAP recovery in the PEG2000 group. Similar recovery patterns were obtained by CAP area measurements and LDH assay. The results suggest that application of low-molecular weight PEG (PEG400) in mild hyperthermia conditions (40 °C) provides the optimum condition for membrane sealing in SCI model.