Synthesis of Novel Caspase Inhibitors for Characterization of the Active Caspase Proteome in Vitro and in Vivo

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• 作者：Alexander J. Henzing ; Helen Dodson ; Joel M. Reid ; Scott H. Kaufmann ; Robert L. Baxter ; William C. Earnshaw
• 刊名：Journal of Medicinal Chemistry
• 出版年：2006
• 出版时间：December 28, 2006
• 年：2006
• 卷：49
• 期：26
• 页码：7636 - 7645
• 全文大小：338K
• 年卷期：v.49,no.26(December 28, 2006)
• ISSN：1520-4804

文摘

Caspases are cysteine proteases that are essential for cytokine maturation and apoptosis. To facilitate thedissection of caspase function in vitro and in vivo, we have synthesized irreversible caspase inhibitors withbiotin attached via linker arms of various lengths (12a-d) and a 2,4-dinitrophenyl labeled inhibitor (13).Affinity labeling of apoptotic extracts followed by blotting reveals that these affinity probes detect activecaspases. Using the strong affinity of avidin for biotin, we have isolated affinity-labeled caspase 6 fromapoptotic cytosolic extracts of cells overexpressing procaspase 6 by treatment with 12c, which containsbiotin attached to the N^{mages/entities/epsiv.gif">}-lysine of the inhibitor by a 22.5 Å linker arm, followed by affinity purification onmonomeric avidin-sepharose beads. Compound 13 has proven sufficiently cell permeable to rescue cellsfrom apoptotic execution. These novel caspase inhibitors should provide powerful probes for the study ofthe active caspase proteome during apoptosis both in vitro and in vivo.