Puri Aaron W, Lupardus Patrick J, Deu Edgar, Albrow Victoria E, Garcia K Christopher, Bogyo Matthew, Shen Aimee
Department of Chemical and Systems Biology, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, California 94305, USA.
Chem Biol. 2010 Nov 24;17(11):1201-11. doi: 10.1016/j.chembiol.2010.09.011.
Clostridium difficile is a leading cause of nosocomial infections. The major virulence factors of this pathogen are the multi-domain toxins TcdA and TcdB. These toxins contain a cysteine protease domain (CPD) that autoproteolytically releases a cytotoxic effector domain upon binding intracellular inositol hexakisphosphate. Currently, there are no known inhibitors of this protease. Here, we describe the rational design of covalent small molecule inhibitors of TcdB CPD. We identified compounds that inactivate TcdB holotoxin function in cells and solved the structure of inhibitor-bound protease to 2.0 Å. This structure reveals the molecular basis of CPD substrate recognition and informed the synthesis of activity-based probes for this enzyme. The inhibitors presented will guide the development of therapeutics targeting C. difficile, and the probes will serve as tools for studying the unique activation mechanism of bacterial toxin CPDs.
艰难梭菌是医院感染的主要病因。该病原体的主要毒力因子是多结构域毒素TcdA和TcdB。这些毒素含有一个半胱氨酸蛋白酶结构域（CPD），该结构域在与细胞内肌醇六磷酸结合后会自蛋白水解释放出一个细胞毒性效应结构域。目前，尚无已知的该蛋白酶抑制剂。在此，我们描述了TcdB CPD共价小分子抑制剂的合理设计。我们鉴定出了能使细胞中TcdB全毒素功能失活的化合物，并将抑制剂结合蛋白酶的结构解析至2.0 Å。该结构揭示了CPD底物识别的分子基础，并为该酶的基于活性的探针的合成提供了依据。所展示的抑制剂将指导针对艰难梭菌的治疗药物的开发，而这些探针将作为研究细菌毒素CPD独特激活机制的工具。
