Journal Article

Structural Insights into Bortezomib-Induced Activation of the Caseinolytic Chaperone-Protease System in Mycobacterium tuberculosis

Huang, Xiaodong, Li, Zimu, Wang, Jingjing, Wang, Xiyue, Xiong, Xiaoli, Yuan, Hang, He, Jun, Zhang, Han, Zhao, Yi, Burston, Steven G., Oliveira, A. Sofia. F., Chen, Xinwen, Zhou, Biao, Gao, Yamin, Hu, Jinxing, Mulholland, Adrian J., Spencer, James, Zhao, Heyu, Liu, Banghui, Zhang, Tianyu, Su, Ning, Fang, Cuiting

2025

Overview

The caseinolytic protease (Clp) system has recently emerged as a promising anti-tuberculosis target. The anti-cancer drug bortezomib exhibits potent anti-mycobacterial activity and binds to Mycobacterium tuberculosis ( Mtb ) Clp protease complexes. We determine cryo-EM structures of Mtb ClpP1P2, ClpC1P1P2 and ClpXP1P2 complexes bound to bortezomib in different conformations. Structural and biochemical data indicate that sub-stoichiometric binding by bortezomib to the protease active sites orthosterically activates the Mtb ClpP1P2 complex. Bortezomib activation of Mtb ClpP1P2 induces structural changes promoting the recruitment of the chaperone-unfoldases, Mtb ClpC1 or Mtb ClpX, facilitating holoenzyme formation. The structures of the Mtb ClpC1P1P2 holoenzyme indicate that Mtb ClpC1 motion, induced by ATP rebinding at the Mtb ClpC1 spiral seam, translocates the substrate. In the Mtb ClpXP1P2 holoenzyme structure, we identify a specialized substrate channel gating mechanism involving the Mtb ClpX pore-2 loop and Mtb ClpP2 N-terminal domains. Our results provide insights into the intricate regulation of the Mtb Clp system and suggest that bortezomib can disrupt this regulation by sub-stoichiometric binding at the Mtb Clp protease sites. The study reveals how bortezomib activates Mycobacterium tuberculosis Clp protease complexes. Cryo-EM structures reveal how sub-stoichiometric bortezomib binding triggers structural changes, mediates holoenzyme formation and disrupts Clp system regulation.

• Publisher: Nature Publishing Group UK,Nature Publishing Group,Nature Portfolio
• Subject: 631/326/41/2536 / 631/45/607/468 / 631/535/1258/1259 / Antineoplastic drugs / Bacterial Proteins - chemistry / Bacterial Proteins - genetics / Bacterial Proteins - metabolism / Bacterial Proteins - ultrastructure / Binding / Bortezomib / Bortezomib - chemistry / Bortezomib - pharmacology / Catalytic Domain / Channel gating / Cryoelectron Microscopy / Endopeptidase Clp - chemistry / Endopeptidase Clp - genetics / Endopeptidase Clp - metabolism / Endopeptidase Clp - ultrastructure / Humanities and Social Sciences / Models, Molecular / Molecular Chaperones - chemistry / Molecular Chaperones - metabolism / multidisciplinary / Mycobacterium tuberculosis / Mycobacterium tuberculosis - drug effects / Mycobacterium tuberculosis - enzymology / Mycobacterium tuberculosis - metabolism / Protease / Protein Binding / Proteinase / Science / Science (multidisciplinary) / Stoichiometry / Tuberculosis