Institut de Biotecnologia i de Biomedicina and Departament de Bioquímica i Biologia Molecular, Universitat Autònoma de Barcelona, E-08193 Bellaterra, Spain
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Department of Pathology, School of Medicine, Stanford University, CA-94025 Stanford, USA
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Mol. BioSyst., 2011,7, 1121-1128
23 Nov 2010,
19 Dec 2010
First published online
14 Jan 2011
Protein aggregation and amyloid formation lie behind an increasing number of human diseases. Here we describe the application of an “aggregation reporter”, in which the test protein is fused to dihydrofolate reductase, as a general method to assess the intracellular solubility of amyloidproteins in eukaryotic background. Because the aggregation state of the target protein is linked directly to yeast cells survival in the presence of methotrexate, protein solubility can be monitored in vivo without the requirement of a functional assay for the protein of interest. In addition, the approach allows the in vivo visualization of the cellular location and aggregated state of the target protein. To demonstrate the applicability of the assay in the screening of genes or compounds that modulate amyloidprotein aggregation in living cells, we have used as models the Alzheimer's amyloid β peptide, polyglutamine expansions of huntingtin, α-synuclein and non-aggregating variants thereof. Moreover, the anti-aggregational properties of small molecules and the effects of the yeast protein quality control machinery have also been evaluated using this method.
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