Objectives: to determine which is the ratio in the mixture of A?py3-42 and A?1-42, where A?py3-42 has the major influence on the aggregation kinetics, secondary structure, morphology and toxicity of A?1-42. Methods: We characterized the cell toxicity by analysis of the deregulation of the calcium; aggregation kinetics by turbidity assay; conformational analysis by Circular Dichroism; morphological characterization by TEM and AFM; secondary structure determination by NanoIR. Results: The different mixtures and peptides alone were differently effective in inducing free Ca2+ increase in the exposed cells, but the 5% mixture showed the higher increase of fluorescence. The conformational analysis showed that the 5% mixture has an intermediate curve trend between A?1-42 and A?py3-42 alone. The turbidity assay showed faster aggregation kinetics for A?py3-42 and slower aggregation rates for A?1-42, the 5% mixture showed the aggregation kinetics more similar to those of A?py3-42. Morphological analysis revealed in A?py3-42 and 50% mixture big aggregates and few fibers, in the other mixtures and A?1-42 short protofibrils and small spheroidal aggregates and in the 5% mixture large spheroidal aggregates. The NanoIR showed about A?1-42 aggregates only in ?-helix and in ?-helix and ?-sheet, about A?py3-42 exclusively in ?-sheet and about 5% mixture in random coil and in a mixture of ?-helix and ?-sheet with more ?-sheet. Conclusions: here we showed that A?py3-42 revealed its maximum effective in the 5% mixture. This finding correlates with the presence of low concentrations of A?py3-42 in the core of aggregates extracted from cerebral deposits
PYROGLUTAMYLATED AMYLOID-beta AFFECTS THE STRUCTURE, MORPHOLOGY AND TOXICITY OF THE FULL LENGTH BY SMALL QUANTITIES
Galante Denise;Angelo Perico;Simona Tomaselli;Cristina D'Arrigo
2015
Abstract
Objectives: to determine which is the ratio in the mixture of A?py3-42 and A?1-42, where A?py3-42 has the major influence on the aggregation kinetics, secondary structure, morphology and toxicity of A?1-42. Methods: We characterized the cell toxicity by analysis of the deregulation of the calcium; aggregation kinetics by turbidity assay; conformational analysis by Circular Dichroism; morphological characterization by TEM and AFM; secondary structure determination by NanoIR. Results: The different mixtures and peptides alone were differently effective in inducing free Ca2+ increase in the exposed cells, but the 5% mixture showed the higher increase of fluorescence. The conformational analysis showed that the 5% mixture has an intermediate curve trend between A?1-42 and A?py3-42 alone. The turbidity assay showed faster aggregation kinetics for A?py3-42 and slower aggregation rates for A?1-42, the 5% mixture showed the aggregation kinetics more similar to those of A?py3-42. Morphological analysis revealed in A?py3-42 and 50% mixture big aggregates and few fibers, in the other mixtures and A?1-42 short protofibrils and small spheroidal aggregates and in the 5% mixture large spheroidal aggregates. The NanoIR showed about A?1-42 aggregates only in ?-helix and in ?-helix and ?-sheet, about A?py3-42 exclusively in ?-sheet and about 5% mixture in random coil and in a mixture of ?-helix and ?-sheet with more ?-sheet. Conclusions: here we showed that A?py3-42 revealed its maximum effective in the 5% mixture. This finding correlates with the presence of low concentrations of A?py3-42 in the core of aggregates extracted from cerebral depositsFile | Dimensione | Formato | |
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