Development of a novel fluorescence assay for studying lipid bilayer perturbation induced by amyloidogenic peptides using cell plasma membrane vesicles.
Biochim Biophys Acta Biomembr
; 1865(3): 184118, 2023 03.
Article
em En
| MEDLINE
| ID: mdl-36621762
ABSTRACT
Numerous pathophysiological conditions are associated with the misfolding and aggregation of proteins into insoluble amyloid fibrils. The mechanisms by which this process leads to cellular dysfunction remain elusive, though several hypotheses point toward the perturbation of the cell plasma membrane by pre-fibrillar intermediates and/or amyloid growth. However, current models to study membrane perturbations are largely limited to synthetic lipid vesicles and most of experimental approaches cannot be transposed to complex cell-derived plasma membrane systems. Herein, vesicles originating from the plasma membrane of erythrocytes and ß-pancreatic cells were used to study the perturbations induced by an amyloidogenic peptide, the islet amyloid polypeptide (IAPP). These biologically relevant lipid vesicles displayed a characteristic clustering in the presence of the amyloidogenic peptide, which was able to rupture membranes. By exploiting Förster resonance energy transfer (FRET), a rapid, simple, and potentially high-throughput assay to detect membrane perturbations of intact mammalian cell plasma membrane vesicles was implemented. The FRET kinetics of membrane perturbations closely correlated with the kinetics of thioflavin-T fluorescence associated with amyloid formation. This novel kinetics assay expands the toolbox available to study amyloid-associated membrane damage, bridging the gap between synthetic lipid vesicles and living cells.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Células Secretoras de Insulina
/
Bicamadas Lipídicas
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
Biochim Biophys Acta Biomembr
Ano de publicação:
2023
Tipo de documento:
Article
País de afiliação:
Canadá