Development of high-throughput assays based on fluorescence polarization for inhibitors of the polo-box domains of polo-like kinases 2 and 3
Development of high-throughput assays based on fluorescence polarization for inhibitors of the polo-box domains of polo-like kinases 2 and 3
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DOI：
10.1016/j.ab.2009.08.031
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发表时间：
2009-12-15
影响因子：
2.9
通讯作者：
Berg, Thorsten
中科院分区：
文献类型：
--
作者：
Reindl, Wolfgang;Graeber, Martin;Berg, Thorsten
丝氨酸/苏氨酸激酶Plk1、Plk2和Plk3含有一个被称为polo-box结构域（PBD）的蛋白-蛋白相互作用结构域。最近，作为atp竞争配体抑制激酶的一种替代方法，人们成功地探索了对癌症靶点Plk1的PBD的抑制。然而，由于Plk1、Plk2和Plk3的PBD具有非常相似的最佳结合基序，Plk1的PBD相对于Plk2和Plk3的PBD的绝对特异性对小分子来说也可能是一个很大的挑战。为了帮助研究Plk PBD抑制剂的活性谱，并确定选择性小分子，以揭示抑制Plk2和Plk3 PBD的细胞后果，我们开发了基于荧光偏振对Plk2和Plk3 PBD的高通量分析。该方法对时间和10%二甲亚砜稳定，Z值>= 0.7，非常适合高通量筛选。此外，我们的数据提供了Plk2和Plk3的pbd结合偏好的见解。(C) 2009爱思唯尔公司版权所有。
The serine/threonine kinases Plk1, Plk2, and Plk3 harbor a protein-protein interaction domain dubbed polo-box domain (PBD). Recently, the inhibition of the PBD of the cancer target Plk1 has been successfully explored as an alternative to the inhibition of the kinase by ATP-competitive ligands. However, because the PBDs of Plk1, Plk2, and Plk3 have very similar optimal binding motifs, absolute specificity for the PBD of Plk1 over the PBDs of Plk2 and Plk3 may also represent a big challenge for a small molecule. To aid in the activity profiling of Plk PBD inhibitors, and to identify selective small molecules that will reveal the cellular consequences of inhibiting the PBDs of Plk2 and Plk3, we have developed high-through put assays based on fluorescence polarization against the PBDs of Plk2 and Plk3. The assays are stable with regard to time and 10% dimethyl sulfoxide and have Z' values >= 0.7, making them well-suited for high-throughput screening. Moreover, our data provide insights into the binding preferences of the PBDs of Plk2 and Plk3. (C) 2009 Elsevier Inc. All rights reserved.