Toward an understanding of protein glycation in Parkinson's disease: in vitro and in vivo studies of folding, aggregation and degradation pathways of alpha-synuclein, synphilin-1 and parkin under glycation conditions.

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Insulin glycation by methylglyoxal results in native-like aggregation and inhibition of fibril formation

Publication . Oliveira, Luis MA; Lages, Ana; Gomes, Ricardo A; Neves, Henrique; Família, Carlos; Coelho, Ana V; Quintas, Alexandre

Background: Insulin is a hormone that regulates blood glucose homeostasis and is a central protein in a medical condition termed insulin injection amyloidosis. It is intimately associated with glycaemia and is vulnerable to glycation by glucose and other highly reactive carbonyls like methylglyoxal, especially in diabetic conditions. Protein glycation is involved in structure and stability changes that impair protein functionality, and is associated with several human diseases, such as diabetes and neurodegenerative diseases like Alzheimer’s disease, Parkinson’s disease and Familiar Amyloidotic Polyneuropathy. In the present work, methylglyoxal was investigated for their effects on the structure, stability and fibril formation of insulin.

2011Journal article

Open access

Extracellular Alpha-Synuclein Oligomers Modulate Synaptic Transmission and Impair LTP Via NMDA-Receptor Activation

Publication . Diógenes, Maria José; Dias, Raquel B.; Rombo, Diogo M.; Miranda, Hugo Vicente; Maiolino, Francesca; Guerreiro, Patrícia; Näsström, Thomas; Franquelim, Henri G.; Oliveira, Luis M. A.; Castanho, Miguel A. R. B.; Lannfelt, Lars; Bergström, Joakim; Ingelsson, Martin; Quintas, Alexandre; Sebastião, Ana M.; Lopes, Luísa V.; Outeiro, Tiago Fleming

Parkinson’s disease (PD) is the most common representative of a group of disorders known as synucleinopathies, in which misfolding and aggregation of -synuclein (a-syn) in various brain regions is themajorpathological hallmark. Indeed, themotorsymptomsinPDare causedby a heterogeneous degeneration of brain neurons not only in substantia nigra pars compacta but also in other extrastriatal areas of the brain. In addition to the well known motor dysfunction in PD patients, cognitive deficits and memory impairment are also an important part of the disorder, probably due to disruption of synaptic transmission and plasticity in extrastriatal areas, including the hippocampus. Here, we investigated the impact of a-syn aggregation onAMPAandNMDAreceptor-mediated rat hippocampal (CA3-CA1) synaptic transmission and long-term potentiation (LTP), the neurophysiological basis for learning and memory. Our data show that prolonged exposure to a-syn oligomers, but not monomers or fibrils, increases basal synaptic transmission through NMDA receptor activation, triggering enhanced contribution of calcium-permeable AMPA receptors. Slices treated with a-syn oligomers were unable to respond with further potentiation to theta-burst stimulation, leading to impaired LTP. Prior delivery of a low-frequency train reinstated the ability to express LTP, implying that exposuretoa-synoligomersdrivestheincreaseofglutamatergicsynaptictransmission,preventingfurtherpotentiationbyphysiologicalstimuli. Our novel findings provide mechanistic insight on how a-syn oligomers may trigger neuronal dysfunction and toxicity in PD and other synucleinopathies.

2012-08-22Journal article

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Glycation potentiates α-synuclein-associated neurodegeneration in synucleinopathies

Publication . Miranda, Hugo Vicente; Szegő, Éva M.; Oliveira, Luís M. A.; Breda, Carlo; Darendelioglu, Ekrem; Oliveira, Rita M. de; Ferreira, Diana G.; Gomes, Marcos A.; Rott, Ruth; Oliveira, Márcia; Munari, Francesca; Enguita, Francisco J.; Simões, Tânia; Rodrigues, Eva F.; Heinrich, Michael; Martins, Ivo C.; Zamolo, Irina; Riess, Olaf; Cordeiro, Carlos; Ponces-Freire, Ana; Santos, Nuno C.; Lopes, Luisa V.; Xiang, Wei; Jovin, Thomas M.; Penque, Deborah; Engelender, Simone; Zweckstetter, Markus; Klucken, Jochen; Giorgini, Flaviano; Quintas, Alexandre; Outeiro, Tiago F.

α-Synuclein misfolding and aggregation is a hallmark in Parkinson’s disease and in several other neurodegenerative diseases known as synucleinopathies. The toxic properties of α-synuclein are conserved from yeast to man, but the precise underpinnings of the cellular pathologies associated are still elusive, complicating the development of effective therapeutic strategies. Combining molecular genetics with target-based approaches, we established that glycation, an unavoidable age-associated post-translational modification, enhanced α-synuclein toxicity in vitro and in vivo, in Drosophila and in mice. Glycation affected primarily the N-terminal region of α-synuclein, reducing membrane binding, impaired the clearance of α-synuclein, and promoted the accumulation of toxic oligomers that impaired neuronal synaptic transmission. Strikingly, using glycation inhibitors, we demonstrated that normal clearance of α-synuclein was re-established, aggregation was reduced, and motor phenotypes in Drosophila were alleviated. Altogether, our study demonstrates glycation constitutes a novel drug target that can be explored in synucleinopathies as well as in other neurodegenerative conditions.

2017-05Journal article

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Insights into the molecular mechanism of protein native-like aggregation upon methylglyoxal glycation

Publication . Oliveira, Luís M. A.; Gomes, Ricardo A.; Yang, Dennis; Família, Carlos; Lages, Ana; Coelho, Ana V.; Murphy, Regina M.; Quintas, Alexandre

"Protein glycation induces structural and stability changes that impair protein function, and is associated with several human neurodegenerative diseases, such as Alzheimer’s disease, Parkinson’s disease and Familial Amyloidotic Polyneuropathy. Recently we have shown that methylglyoxal induces and stabilizes the formation of small native-like aggregates in the amyloidogenic protein insulin and the same was previously shown for α-synuclein. However, the fundamental biophysical mechanism underlying such methylglyoxal-induced protein aggregation is not yet fully understood. In this study, we used the model protein cytochrome c to characterize the specific glycation targets and to investigate the glycation effects on protein structure, stability and aggregation. Methylglyoxal was found modify cytochrome c in a single residue and to induce the formation of cytochrome c native-like aggregates. Additionally, it is shown that methylglyoxal glycation of cytochrome c also results in the formation of a partially unfolded species. Interestingly, the formation of this partially unfolded species is not implicated in the aggregation process, a clear difference from amyloid fibril mechanisms that involve partially or totally unfolded intermediates. Equilibrium-unfolding experiments using guanidinium hydrochloride shows that glycation strongly reduces cytochrome c conformational stability. This reduction is balanced by aggregation that increases conformational stability. The data collected from analytical and spectroscopic techniques along with kinetic analysis based on least-squares parameter fitting and statistical model discrimination permitted the proposal of a comprehensive thermodynamic and kinetic model for native-like aggregation of methylglyoxal glycated cytochrome c."

2013-06Journal article

Open access