Chemoproteomic Profiling Reveals the Mechanism of Bile Acid Tolerance in Bacteria

Bile acids (BAs) are a class of endogenous metabolites
with important
functions. As amphipathic molecules, BAs have strong antibacterial
effects, preventing overgrowth of the gut microbiota and defending
the invasion of pathogens. However, some disease-causing pathogens
can survive the BA stress and knowledge is limited about how they
develop BA tolerance. In this work, we applied a quantitative chemoproteomic
strategy to profile BA-interacting proteins in bacteria, aiming to
discover the sensing pathway of BAs. Using a clickable and photo-affinity
BA probe with quantitative mass spectrometry, we identified a list
of histidine kinases (HKs) of the two-component systems (TCS) in bacteria
as the novel binding targets of BA. Genetic screening revealed that
knocking out one specific HK, EnvZ, renders bacteria with significant
sensitivity to BA. Further biochemical and genetic experiments demonstrated
that BA binds to a specific pocket in EnvZ and activates a downstream
signaling pathway to help efflux of BA from bacteria, resulting in
BA tolerance. Collectively, our data revealed that EnvZ is a novel
sensor of BA in bacteria and its associated TCS signaling pathway
plays a critical role in mediating bacterial BA tolerance, which opens
new opportunities to combat BA-tolerating pathogens.

胆汁酸（Bile acids, BAs）是一类具有重要生理功能的内源性代谢物。作为两亲性分子，胆汁酸具备强效抗菌活性，可抑制肠道菌群过度增殖并抵御病原体入侵。然而，部分致病病原体能够耐受胆汁酸压力，目前学界对其形成胆汁酸耐受的具体机制仍知之甚少。本研究采用定量化学蛋白质组学策略，对细菌中与胆汁酸相互作用的蛋白质进行谱学表征，旨在揭示细菌感知胆汁酸的信号通路。借助可点击光亲和胆汁酸探针结合定量质谱技术，我们鉴定出细菌双组分系统（Two-Component Systems, TCS）中的一系列组氨酸激酶（Histidine Kinases, HKs）为胆汁酸的新型结合靶点。遗传筛选实验发现，敲除特定组氨酸激酶EnvZ会使细菌对胆汁酸的敏感性显著升高。进一步的生化与遗传实验证实，胆汁酸可结合至EnvZ的特定口袋结构，并激活下游信号通路以促进细菌对胆汁酸的外排，最终介导细菌的胆汁酸耐受。综上，本研究揭示EnvZ是细菌中一类新型胆汁酸感受器，其关联的双组分信号通路在介导细菌胆汁酸耐受过程中发挥关键作用，这一发现为对抗胆汁酸耐受型病原体提供了全新的研究方向。