Staurosporine-based binding assay for testing the affinity of compounds to protein kinases

被引：10

作者：

Iyer, Granesh H. ^{[1
]}

Taslimi, Paul ^{[1
]}

Pazhanisamy, S. ^{[1
]}

机构：

[1] Vertex Pharmaceut Inc, Cambridge, MA 02139 USA

来源：

ANALYTICAL BIOCHEMISTRY | 2008年 / 373卷 / 02期

关键词：

staurosporine; binding assay; competitive ligand displacement; fluorescence; dissociation constant; protein kinase;

D O I：

10.1016/j.ab.2007.11.004

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Staurosporine is a broad-spectrum inhibitor of both tyrosine and serine/threonine protein kinases. Excitation of staurosporine and its analogues at 296 nm results in major emission bands centered at 378 and 396 nm. The intensity of the emission bands is enhanced oil binding to the adenosine triphosphate (ATP) site of many protein kinases. This property was used to develop a competitive displacement assay for evaluating the binding affinity of small molecules to protein kinases. The assay was validated in both cuvette and plate formats for several phosphorylated and non-phosphorylated protein kinases. The throughput of the assay is high enough to be used in drug discovery for screening as well as lead optimization. (C) 2007 Elsevier Inc. All rights reserved.

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页码：197 / 206

页数：10

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