Fluorescence Linked Enzyme Chemoproteomic Strategy for Discovery of a Potent and Selective DAPK1 and ZIPK Inhibitor

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• 作者：David A. Carlson ; Aaron S. Franke ; Douglas H. Weitzel ; Brittany L. Speer ; Philip F. Hughes ; Laura Hagerty ; Christopher N. Fortner ; James M. Veal ; Thomas E. Barta ; Bartosz J. Zieba ; Avril V. Somlyo ; Cindy Sutherland ; Jing Ti Deng ; Michael P. Walsh ; Justin A. MacDonald ; Timothy A. J. Haystead
• 刊名：ACS Chemical Biology
• 出版年：2013
• 出版时间：December 20, 2013
• 年：2013
• 卷：8
• 期：12
• 页码：2715-2723
• 全文大小：503K
• 年卷期：v.8,no.12(December 20, 2013)
• ISSN：1554-8937

文摘

DAPK1 and ZIPK (also called DAPK3) are closely related serine/threonine protein kinases that regulate programmed cell death and phosphorylation of non-muscle and smooth muscle myosin. We have developed a fluorescence linked enzyme chemoproteomic strategy (FLECS) for the rapid identification of inhibitors for any element of the purinome and identified a selective pyrazolo[3,4-d]pyrimidinone (HS38) that inhibits DAPK1 and ZIPK in an ATP-competitive manner at nanomolar concentrations. In cellular studies, HS38 decreased RLC20 phosphorylation. In ex vivo studies, HS38 decreased contractile force generated in mouse aorta, rabbit ileum, and calyculin A stimulated arterial muscle by decreasing RLC20 and MYPT1 phosphorylation. The inhibitor also promoted relaxation in Ca²⁺-sensitized vessels. A close structural analogue (HS43) with 5-fold lower affinity for ZIPK produced no effect on cells or tissues. These findings are consistent with a mechanism of action wherein HS38 specifically targets ZIPK in smooth muscle. The discovery of HS38 provides a lead scaffold for the development of therapeutic agents for smooth muscle related disorders and a chemical means to probe the function of DAPK1 and ZIPK across species.