Thioflavin-S staining of bacterial inclusion bodies for the fast, simple, and inexpensive screening of amyloid aggregation inhibitors.
Amyloid aggregation is linked to a large number of human disorders, from neurodegenerative diseases as Alzheimer's disease (AD) or spongiform encephalopathies to non-neuropathic localized diseases as type II diabetes and cataracts. Because the formation of insoluble inclusion bodies (IBs) during recombinant protein production in bacteria has been recently shown to share mechanistic features with amyloid self-assembly, bacteria have emerged as a tool to study amyloid aggregation. Herein we present a fast, simple, inexpensive and quantitative method for the screening of potential anti-aggregating drugs. This method is based on monitoring the changes in the binding of thioflavin-S to intracellular IBs in intact Eschericchia coli cells in the presence of small chemical compounds. This in vivo technique fairly recapitulates previous in vitro data. Here we mainly use the Alzheimer's related β-amyloid peptide as a model system, but the technique can be easily implemented for screening inhibitors relevant for other conformational diseases simply by changing the recombinant amyloid protein target. Indeed, we show that this methodology can be also applied to the evaluation of inhibitors of the aggregation of tau protein, another amyloidogenic protein with a key role in AD.
S
S
Pouplana
A
A
Espargaro
C
C
Galdeano
E
E
Viayna
I
I
Sola
S
S
Ventura
D
D
Muñoz-Torrero
Departament de Fisicoquimica, Facultat de Farmacia, and Institut de Nanociencia i Nanotecnologia (IN2UB), and Laboratori de Quimica Farmaceutica (Unitat Associada al CSIC), Facultat de Farmacia, and Institut de Biomedicina (IBUB), Universitat de Barcelona, Av. Joan XXIII, 27-31, 08028-Barcelona, Spain. rsabate@ub.edu.
R
R
Sabate
D016229:Amyloid beta-Peptides / Q000032:analysis* / Q000378:metabolism; D052160:Benzothiazoles; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D002479:Inclusion Bodies / Q000737:chemistry* / Q000378:metabolism; D013844:Thiazoles / Q000737:chemistry*; D013997:Time Factors
D016229:Amyloid beta-Peptides; D052160:Benzothiazoles; D013844:Thiazoles; C009462:thioflavin T