蛋白质磷酸酶荧光活性探针的设计与合成
Design and synthesis of fluorescent activity probes for protein phosphatases.
作者信息
Casey Garrett R, Beck Jon R, Stains Cliff I
机构信息
Department of Chemistry and Nebraska Center for Integrated Biomolecular Communication, University of Nebraska-Lincoln, Lincoln, NE, United States.
Department of Chemistry and Nebraska Center for Integrated Biomolecular Communication, University of Nebraska-Lincoln, Lincoln, NE, United States; Cancer Genes and Molecular Recognition Program, Fred & Pamela Buffet Cancer Center, University of Nebraska Medical Center, Omaha, NE, United States.
出版信息
Methods Enzymol. 2019;622:29-53. doi: 10.1016/bs.mie.2019.02.002. Epub 2019 Mar 8.
Protein phosphatases act in concert with protein kinases to regulate and maintain the phosphoproteome. However, the catalog of chemical tools to directly monitor the enzymatic activity of phosphatases has lagged behind their kinase counterparts. In this chapter, we provide protocols for repurposing the phosphorylation-sensitive sulfonamido-oxine fluorophore known as Sox to afford direct activity probes for phosphatases. With validated activity probes in-hand, inhibitor screens can be conducted with recombinant enzyme and the role of phosphatases in cell signaling can be investigated in unfractionated cell lysates.
蛋白质磷酸酶与蛋白激酶协同作用，以调节和维持磷酸化蛋白质组。然而，直接监测磷酸酶酶活性的化学工具目录落后于其激酶同类工具。在本章中，我们提供了重新利用被称为Sox的磷酸化敏感磺酰胺氧基荧光团的方案，以提供用于磷酸酶的直接活性探针。有了经过验证的活性探针，就可以用重组酶进行抑制剂筛选，并在未分级的细胞裂解物中研究磷酸酶在细胞信号传导中的作用。
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