Department of Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, University of Tübingengrid.10392.39, Tübingen, Germany.
Cluster of Excellence-Controlling Microbes to Fight Infections, University of Tübingengrid.10392.39, Tübingen, Germany.
mBio. 2022 Dec 20;13(6):e0141322. doi: 10.1128/mbio.01413-22. Epub 2022 Oct 26.
Clp proteases consist of a proteolytic, tetradecameric ClpP core and AAA+ Clp-ATPases. Streptomycetes, producers of a plethora of secondary metabolites, encode up to five different ClpP homologs, and the composition of their unusually complex Clp protease machinery has remained unsolved. Here, we report on the composition of the housekeeping Clp protease in , consisting of a heterotetradecameric core built of ClpP1, ClpP2, and the cognate Clp-ATPases ClpX, ClpC1, or ClpC2, all interacting with ClpP2 only. Antibiotic acyldepsipeptides (ADEP) dysregulate the Clp protease for unregulated proteolysis. We observed that ADEP binds ClpP1, but not ClpP2, thereby not only triggering the degradation of nonnative protein substrates but also accelerating Clp-ATPase-dependent proteolysis. The explanation is the concomitant binding of ADEP and Clp-ATPases to opposite sides of the ClpP1P2 barrel, hence revealing a third, so far unknown mechanism of ADEP action, i.e., the accelerated proteolysis of native protein substrates by the Clp protease. Clp proteases are antibiotic and anticancer drug targets. Composed of the proteolytic core ClpP and a regulatory Clp-ATPase, the protease machinery is important for protein homeostasis and regulatory proteolysis. The acyldepsipeptide antibiotic ADEP targets ClpP and has shown promise for treating multiresistant and persistent bacterial infections. The molecular mechanism of ADEP is multilayered. Here, we present a new way how ADEP can deregulate the Clp protease system. Clp-ATPases and ADEP bind to opposite sides of ClpP, accelerating the degradation of natural Clp protease substrates. We also demonstrate the composition of the major Clp protease complex, a heteromeric ClpP1P2 core with the Clp-ATPases ClpX, ClpC1, or ClpC2 exclusively bound to ClpP2, and the killing mechanism of ADEP in .
Clp 蛋白酶由一个蛋白水解的十四聚体 ClpP 核心和 AAA+ Clp-ATPase 组成。产生大量次级代谢产物的链霉菌编码多达五个不同的 ClpP 同源物，其异常复杂的 Clp 蛋白酶机制的组成仍未解决。在这里，我们报告了组成组成的管家 Clp 蛋白酶，由一个异源十四聚体核心组成，由 ClpP1、ClpP2 和同源的 Clp-ATPases ClpX、ClpC1 或 ClpC2 组成，所有这些都只与 ClpP2 相互作用。抗生素酰基二肽（ADEP）扰乱 Clp 蛋白酶以进行不受调节的蛋白水解。我们观察到 ADEP 结合 ClpP1，但不结合 ClpP2，从而不仅触发非天然蛋白底物的降解，而且加速 Clp-ATPase 依赖性蛋白水解。解释是 ADEP 和 Clp-ATPases 同时结合到 ClpP1P2 桶的相反两侧，因此揭示了 ADEP 作用的第三个，迄今为止未知的机制，即 Clp 蛋白酶对天然蛋白底物的加速蛋白水解。Clp 蛋白酶是抗生素和抗癌药物的靶标。由蛋白水解核心 ClpP 和调节 Clp-ATPase 组成，蛋白酶机制对蛋白质的动态平衡和调节蛋白水解很重要。酰基二肽抗生素 ADEP 靶向 ClpP，并显示出治疗多药耐药和持久细菌感染的潜力。ADEP 的分子机制是多层次的。在这里，我们提出了 ADEP 可以使 Clp 蛋白酶系统失活的一种新方式。Clp-ATPases 和 ADEP 结合到 ClpP 的相反两侧，加速了天然 Clp 蛋白酶底物的降解。我们还证明了主要 Clp 蛋白酶复合物的组成，一个异源 ClpP1P2 核心，与 Clp-ATPases ClpX、ClpC1 或 ClpC2 仅与 ClpP2 结合，以及 ADEP 在 中的杀伤机制。
