Integrative X-ray Structure and Molecular Modeling for the Rationalization of Procaspase-8 Inhibitor Potency and Selectivity.,ACS Chemical Biology

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Integrative X-ray Structure and Molecular Modeling for the Rationalization of Procaspase-8 Inhibitor Potency and Selectivity.
ACS Chemical Biology ( IF 3.5 ) Pub Date : 2020-01-23 , DOI: 10.1021/acschembio.0c00019
Janice H Xu , Jerome Eberhardt , Brianna Hill-Payne ₁ , Gonzalo E González-Páez , José Omar Castellón ₁ , Benjamin F Cravatt , Stefano Forli , Dennis W Wolan , Keriann M Backus ₁

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Caspases are a critical class of proteases involved in regulating programmed cell death and other biological processes. Selective inhibitors of individual caspases, however, are lacking, due in large part to the high structural similarity found in the active sites of these enzymes. We recently discovered a small-molecule inhibitor, 63-R, that covalently binds the zymogen, or inactive precursor (pro-form), of caspase-8, but not other caspases, pointing to an untapped potential of procaspases as targets for chemical probes. Realizing this goal would benefit from a structural understanding of how small molecules bind to and inhibit caspase zymogens. There have, however, been very few reported procaspase structures. Here, we employ X-ray crystallography to elucidate a procaspase-8 crystal structure in complex with 63-R, which reveals large conformational changes in active-site loops that accommodate the intramolecular cleavage events required for protease activation. Combining these structural insights with molecular modeling and mutagenesis-based biochemical assays, we elucidate key interactions required for 63-R inhibition of procaspase-8. Our findings inform the mechanism of caspase activation and its disruption by small molecules and, more generally, have implications for the development of small molecule inhibitors and/or activators that target alternative (e.g., inactive precursor) protein states to ultimately expand the druggable proteome.

中文翻译：

用于合理化 Procaspase-8 抑制剂效力和选择性的综合 X 射线结构和分子建模。

半胱天冬酶是一类关键的蛋白酶，参与调节程序性细胞死亡和其他生物过程。然而，缺乏单个半胱天冬酶的选择性抑制剂，这在很大程度上是由于这些酶的活性位点中发现的高度结构相似性。我们最近发现了一种小分子抑制剂 63-R，它与 caspase-8 的酶原或无活性前体（前体）共价结合，但不与其他 caspase 结合，这表明 procaspase 作为化学探针靶标的潜力尚未开发。。实现这一目标将受益于对小分子如何结合并抑制半胱天冬酶原的结构理解。然而，有关天冬氨酸蛋白酶原结构的报道非常少。在这里，我们采用 X 射线晶体学来阐明与 63-R 复合的 procaspase-8 晶体结构，这揭示了活性位点环中的巨大构象变化，这些变化适应了蛋白酶激活所需的分子内裂解事件。将这些结构见解与分子建模和基于诱变的生化测定相结合，我们阐明了 63-R 抑制 procaspase-8 所需的关键相互作用。我们的研究结果揭示了半胱天冬酶激活及其被小分子破坏的机制，更一般地说，对针对替代（例如，非活性前体）蛋白质状态以最终扩展可成药蛋白质组的小分子抑制剂和/或激活剂的开发具有影响。

更新日期：2020-01-24

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