Detection of the ATPase activity of the molecular chaperones Hsp90 and Hsp72 using the TranscreenerTM ADP assay kit.
J Biomol Screen
; 15(3): 279-86, 2010 Mar.
Article
en En
| MEDLINE
| ID: mdl-20147598
ABSTRACT
The molecular chaperone heat shock protein 90 (Hsp90) is required for the correct folding and stability of a number of client proteins that are important for the growth and maintenance of cancer cells. Heat shock protein 72 (Hsp72), a co-chaperone of Hsp90, is also emerging as an attractive cancer drug target. Both proteins bind and hydrolyze adenosine triphosphate (ATP), and ATPase activity is essential for their function. Inhibition of Hsp90 ATPase activity leads to the degradation of client proteins, resulting in cell growth inhibition and apoptosis. Several small-molecule inhibitors of the ATPase activity of Hsp90 have been described and are currently being evaluated clinically for the treatment of cancer. A number of methods for the measurement of ATPase activity have been previously used, but not all of these are ideally suited to screening cascades in drug discovery projects. The authors have evaluated the use of commercial reagents (Transcreener ADP) for the measurement of ATPase activity of both yeast and human Hsp90 (ATP K(m) approximately 500 microM) and human Hsp72 (ATP K(m) ~1 microM). The low ATPase activity of human Hsp90 and its stimulation by the co-chaperone Aha1 was measured with ease using reduced incubation times, generating robust data (Z' = 0.75). The potency of several small-molecule inhibitors of both Hsp90 and Hsp72 was determined using the Transcreener reagents and compared well to that determined using other assay formats.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Juego de Reactivos para Diagnóstico
/
Adenosina Trifosfatasas
/
Proteínas HSP90 de Choque Térmico
/
Proteínas del Choque Térmico HSP72
/
Pruebas de Enzimas
Tipo de estudio:
Diagnostic_studies
Límite:
Humans
Idioma:
En
Revista:
J Biomol Screen
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2010
Tipo del documento:
Article
País de afiliación:
Reino Unido