RESUMO
Lewy bodies and Lewy neurites, neuropathological hallmarks of several neurological diseases, are mainly made of filamentous assemblies of α-synuclein. However, other macromolecules including Tau, ubiquitin, glyceraldehyde-3-phosphate dehydrogenase, and glycosaminoglycans are routinely found associated with these amyloid deposits. Glyceraldehyde-3-phosphate dehydrogenase is a glycolytic enzyme that can form fibrillar aggregates in the presence of acidic membranes, but its role in Parkinson disease is still unknown. In this work, the ability of heparin to trigger the amyloid aggregation of this protein at physiological conditions of pH and temperature is demonstrated by infrared and fluorescence spectroscopy, dynamic light scattering, small angle x-ray scattering, circular dichroism, and fluorescence microscopy. Aggregation proceeds through the formation of short rod-like oligomers, which elongates in one dimension. Heparan sulfate was also capable of inducing glyceraldehyde-3-phosphate dehydrogenase aggregation, but chondroitin sulfates A, B, and C together with dextran sulfate had a negligible effect. Aided with molecular docking simulations, a putative binding site on the protein is proposed providing a rational explanation for the structural specificity of heparin and heparan sulfate. Finally, it is demonstrated that in vitro the early oligomers present in the glyceraldehyde-3-phosphate dehydrogenase fibrillation pathway promote α-synuclein aggregation. Taking into account the toxicity of α-synuclein prefibrillar species, the heparin-induced glyceraldehyde-3-phosphate dehydrogenase early oligomers might come in useful as a novel therapeutic strategy in Parkinson disease and other synucleinopathies.
Assuntos
Amiloide/química , Gliceraldeído-3-Fosfato Desidrogenases/química , Heparina/química , Multimerização Proteica , alfa-Sinucleína/química , Amiloide/metabolismo , Animais , Sulfatos de Condroitina/química , Sulfatos de Condroitina/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Heparitina Sulfato/química , Heparitina Sulfato/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Doença de Parkinson/metabolismo , Coelhos , alfa-Sinucleína/metabolismoRESUMO
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional enzyme related with Huntington's, Parkinson's and Alzheimer's diseases. The ability of negatively charged membranes to induce a rapid formation of GAPDH amyloid fibrils has been demonstrated, but the mechanisms by which GAPDH reaches the fibrillar state remains unclear. In this report, we describe the structural changes undergone by GAPDH at physiological pH and temperature conditions right from its interaction with acidic membranes until the amyloid fibril is formed. According to our results, the GAPDH-membrane binding induces a beta-structuring process along with a loss of quaternary structure in the enzyme. In this way, experimental evidences on the initial steps of GAPDH amyloid fibrils formation pathway are provided.
Assuntos
Amiloide/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Membranas Artificiais , Amiloide/ultraestrutura , Gliceraldeído-3-Fosfato Desidrogenases/ultraestrutura , Humanos , Microscopia Eletrônica de Transmissão , Multimerização Proteica , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
Inhibition or reversion of protein self-aggregation has been suggested as a possible preventive mechanism against amyloid diseases, and many efforts are underway to found out molecules capable to restrain the protein aggregation process. In this paper, the inhibitory effects of thyroid hormone analogues on heat-induced fibrillation process of serum albumin are reported. Among the analogues tested, 3,5,3',5'-tetraiodothyroacetic and 3,5,3'-triiodothyroacetic acid showed the most important inhibitory effects on amyloid formation. Thyroxine exhibits a lesser protective effect, while 3,5,3'-triiodothyronine showed no significant inhibition. The gaining of a negative charge together with a size reduction of the hormone molecule could play an essential role in the inhibition of fibrils formation. According to infrared spectroscopy results, the thyroid hormones analogues protective effects proceed via the stabilization of the protein native structure. The current work demonstrates the effectiveness of naturally occurring molecules in the inhibition of albumin fibril formation.
Assuntos
Albumina Sérica/metabolismo , Tiroxina/análogos & derivados , Tri-Iodotironina/análogos & derivados , Amiloide/metabolismo , Animais , Bovinos , Relação Dose-Resposta a Droga , Desenho de Fármacos , Temperatura Alta , Cinética , Ligação Proteica/efeitos dos fármacos , Estrutura Secundária de Proteína/efeitos dos fármacos , Albumina Sérica/química , Espectroscopia de Infravermelho com Transformada de Fourier , Tiroxina/farmacologia , Tri-Iodotironina/farmacologiaRESUMO
The muscle-type nicotinic receptor has two pharmacologically distinguishable acetylcholine binding sites at the alpha-gamma and alpha-delta subunit interfaces; alpha-conotoxins can bind them selectively. As reported, alpha-conotoxin MI has greater affinity for the site near the alpha-delta interface of the BC(3)H1 cell receptor but, in the case of the Torpedo californica receptor, displays greater affinity for that near the alpha-gamma interface. To further investigate ligand selectivity, we study the conotoxin MI-Torpedo marmorata receptor interaction. In this work, we show the binding of alpha-conotoxin MI to the T. marmorata receptor and the influence of the antagonist alpha-Bungarotoxin and the agonist carbamylcholine on such binding; in addition, and contrasting with the results for the Torpedo californica receptor, we identify the alpha-delta subunit interface as the high affinity binding site. This is the first work describing different characteristics of the interaction between alpha-conotoxin MI and receptors from different species of the same genus.