Development and optimization of a useful assay for determining Hsp90's inherent ATPase activity.
The Hsp90 molecular chaperone is responsible for the conformational maturation of nascent polypeptides and the rematuration of denatured proteins. Inhibition of Hsp90 represents a promising approach towards the treatment of cancer because numerous signaling cascades can be simultaneously targeted by disruption of the Hsp90-mediated process. Hsp90's ATPase activity is essential to the Hsp90-mediated protein folding process, consequently, a coupled assay was developed and optimized for determination of Hsp90's inherent ATPase activity. Using maltose phosphorylase, glucose oxidase, and horseradish peroxidase as components of this assay, a highly reproducible assay with a Z-factor of 0.87 has been produced.
Avila
Christopher
C
The Department of Medicinal Chemistry, The University of Kansas, Malott 4070, Lawrence, 66045-7563, USA.
Kornilayev
Boris A
BA
Blagg
Brian S J
BS
D000251:Adenosine Triphosphatases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D002384:Catalysis; D005949:Glucose Oxidase / Q000378:metabolism; D005964:Glucosyltransferases / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000032:analysis / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D006735:Horseradish Peroxidase / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology; D015394:Molecular Structure; D015203:Reproducibility of Results; D013696:Temperature
D018841:HSP90 Heat-Shock Proteins; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide; D005949:Glucose Oxidase; D006735:Horseradish Peroxidase; D005964:Glucosyltransferases; C021535:maltose phosphorylase; D000251:Adenosine Triphosphatases