i3S, Instituto de Investigacao e Inovacao em Saude, Universidade do Porto, Porto, Portugal.
Department Macromolecular Structures, Cryo-EM Facility, Centro Nacional de Biotecnología-CSIC, Madrid, Spain.
J Biol Chem. 2022 Nov;298(11):102553. doi: 10.1016/j.jbc.2022.102553. Epub 2022 Oct 6.
The unfoldase ClpC1 is one of the most exciting drug targets against tuberculosis. This AAA+ unfoldase works in cooperation with the ClpP1P2 protease and is the target of at least four natural product antibiotics: cyclomarin, ecumicin, lassomycin, and rufomycin. Although these molecules are promising starting points for drug development, their mechanisms of action remain largely unknown. Taking advantage of a middle domain mutant, we determined the first structure of Mycobacterium tuberculosis ClpC1 in its apo, cyclomarin-, and ecumicin-bound states via cryo-EM. The obtained structure displays features observed in other members of the AAA+ family and provides a map for further drug development. While the apo and cyclomarin-bound structures are indistinguishable and have N-terminal domains that are invisible in their respective EM maps, around half of the ecumicin-bound ClpC1 particles display three of their six N-terminal domains in an extended conformation. Our structural observations suggest a mechanism where ecumicin functions by mimicking substrate binding, leading to ATPase activation and changes in protein degradation profile.
解旋酶 ClpC1 是抗结核药物最令人兴奋的靶点之一。这种 AAA+解旋酶与 ClpP1P2 蛋白酶协同作用，是至少四种天然产物抗生素的靶标：环马菌素、依库霉素、拉索霉素和鲁福霉素。尽管这些分子是药物开发的有前途的起点，但它们的作用机制在很大程度上仍然未知。利用一个中间结构域突变体，我们通过 cryo-EM 确定了结核分枝杆菌 ClpC1 在其apo、环马菌素和依库霉素结合状态下的第一个结构。获得的结构显示了在其他 AAA+家族成员中观察到的特征，并为进一步的药物开发提供了一个图谱。虽然 apo 和环马菌素结合结构无法区分，并且各自的 EM 图谱中均无法看到 N 端结构域，但在依库霉素结合的 ClpC1 颗粒中，约有一半显示出三种 N 端结构域呈伸展构象。我们的结构观察表明，依库霉素的作用机制是通过模拟底物结合，导致 ATP 酶激活和蛋白质降解谱的变化。
