Dou Chao, Long Zhaolin, Li Shoujie, Zhou Dan, Jin Ying, Zhang Li, Zhang Xuan, Zheng Yanhui, Li Lin, Zhu Xiaofeng, Liu Zheng, He Siyu, Yan Weizhu, Yang Lulu, Xiong Jie, Fu Xianghui, Qi Shiqian, Ren Haiyan, Chen She, Dai Lunzhi, Wang Binju, Cheng Wei
Division of Respiratory and Critical Care Medicine, Respiratory Infection and Intervention Laboratory of Frontiers Science Center for Disease-related Molecular Network, State Key Laboratory of Biotherapy, West China Hospital of Sichuan University, Chengdu, Sichuan, China.
State Key Laboratory of Structural Chemistry of Solid Surface and Fujian Provincial Key Laboratory of Theoretical and Computational Chemistry, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen, Fujian, China.
Cell Res. 2022 Mar;32(3):302-314. doi: 10.1038/s41422-022-00620-2. Epub 2022 Feb 2.
Methanobactins (Mbns) are a family of copper-binding peptides involved in copper uptake by methanotrophs, and are potential therapeutic agents for treating diseases characterized by disordered copper accumulation. Mbns are produced via modification of MbnA precursor peptides at cysteine residues catalyzed by the core biosynthetic machinery containing MbnB, an iron-dependent enzyme, and MbnC. However, mechanistic details underlying the catalysis of the MbnBC holoenzyme remain unclear. Here, we present crystal structures of MbnABC complexes from two distinct species, revealing that the leader peptide of the substrate MbnA binds MbnC for recruitment of the MbnBC holoenzyme, while the core peptide of MbnA resides in the catalytic cavity created by the MbnB-MbnC interaction which harbors a unique tri-iron cluster. Ligation of the substrate sulfhydryl group to the tri-iron center achieves a dioxygen-dependent reaction for oxazolone-thioamide installation. Structural analysis of the MbnABC complexes together with functional investigation of MbnB variants identified a conserved catalytic aspartate residue as a general base required for MbnBC-mediated MbnA modification. Together, our study reveals the similar architecture and function of MbnBC complexes from different species, demonstrating an evolutionarily conserved catalytic mechanism of the MbnBC holoenzymes.
甲烷菌素（Mbns）是一类与甲烷营养菌摄取铜有关的铜结合肽，是治疗以铜蓄积紊乱为特征疾病的潜在治疗剂。Mbns是通过由含MbnB（一种铁依赖性酶）和MbnC的核心生物合成机制催化的MbnA前体肽在半胱氨酸残基处的修饰而产生的。然而，MbnBC全酶催化的机制细节仍不清楚。在此，我们展示了来自两个不同物种的MbnABC复合物的晶体结构，揭示底物MbnA的前导肽与MbnC结合以募集MbnBC全酶，而MbnA的核心肽位于由MbnB-MbnC相互作用形成的催化腔中，该催化腔含有独特的三铁簇。底物巯基与三铁中心的连接实现了用于恶唑酮-硫代酰胺安装的双氧依赖性反应。MbnABC复合物的结构分析以及MbnB变体的功能研究确定了一个保守的催化天冬氨酸残基是MbnBC介导的MbnA修饰所需的通用碱。总之，我们的研究揭示了来自不同物种的MbnBC复合物的相似结构和功能，证明了MbnBC全酶的进化保守催化机制。
