High-Throughput Differential Scanning Fluorimetry of GFP-Tagged Proteins.
Methods Mol Biol
; 2089: 69-85, 2020.
Article
en En
| MEDLINE
| ID: mdl-31773648
Differential scanning fluorimetry is useful for a wide variety of applications including characterization of protein function, structure-activity relationships, drug screening, and optimization of buffer conditions for protein purification, enzyme activity, and crystallization. A limitation of classic differential scanning fluorimetry is its reliance on highly purified protein samples. This limitation is overcome through differential scanning fluorimetry of GFP-tagged proteins (DSF-GTP). DSF-GTP specifically measures the unfolding and aggregation of a target protein fused to GFP through its proximal perturbation effects on GFP fluorescence. As a result of this unique principle, DSF-GTP can specifically measure the thermal stability of a target protein in the presence of other proteins. Additionally, the GFP provides a unique in-assay quality control measure. Here, we describe the workflow, steps, and important considerations for executing a DSF-GTP experiment in a 96-well plate format.
Palabras clave
Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Rastreo Diferencial de Calorimetría
/
Proteínas Fluorescentes Verdes
/
Ensayos Analíticos de Alto Rendimiento
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Fluorometría
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2020
Tipo del documento:
Article
País de afiliación:
Australia