Feng Zhaohang, Yashiro Yuka, Tomita Kozo
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba 277-8562, Japan.
Nucleic Acids Res. 2025 Jan 7;53(1). doi: 10.1093/nar/gkae735.
Contact-dependent growth inhibition (CDI) is a bacterial competition mechanism, wherein the C-terminal toxin domain of CdiA protein (CdiA-CT) is transferred from one bacterium to another, impeding the growth of the toxin recipient. In uropathogenic Escherichia coli 536, CdiA-CT (CdiA-CTEC536) is a tRNA anticodon endonuclease that requires a cysteine biogenesis factor, CysK, for its activity. However, the mechanism underlying tRNA recognition and cleavage by CdiA-CTEC536 remains unresolved. Here, we present the cryo-EM structure of the CysK:CdiA-CTEC536:tRNA ternary complex. The interaction between CdiA-CTEC536 and CysK stabilizes the CdiA-CTEC536 structure and facilitates tRNA binding and the formation of the CdiA-CTEC536 catalytic core structure. The bottom-half of the tRNA interacts exclusively with CdiA-CTEC536 and the α-helices of CdiA-CTEC536 engage with the minor and major grooves of the bottom-half of tRNA, positioning the tRNA anticodon loop at the CdiA-CTEC536 catalytic site for tRNA cleavage. Thus, CysK serves as a platform facilitating the recognition and cleavage of substrate tRNAs by CdiA-CTEC536.
接触依赖性生长抑制（CDI）是一种细菌竞争机制，其中CdiA蛋白的C端毒素结构域（CdiA-CT）从一种细菌转移到另一种细菌，从而抑制毒素受体的生长。在尿路致病性大肠杆菌536中，CdiA-CT（CdiA-CTEC536）是一种tRNA反密码子内切酶，其活性需要半胱氨酸生物合成因子CysK。然而，CdiA-CTEC536识别和切割tRNA的潜在机制仍未得到解决。在这里，我们展示了CysK:CdiA-CTEC536:tRNA三元复合物的冷冻电镜结构。CdiA-CTEC536与CysK之间的相互作用稳定了CdiA-CTEC536的结构，并促进了tRNA的结合以及CdiA-CTEC536催化核心结构的形成。tRNA的下半部分仅与CdiA-CTEC536相互作用，并且CdiA-CTEC536的α螺旋与tRNA下半部分的小沟和大沟结合，将tRNA反密码子环定位在CdiA-CTEC536催化位点以进行tRNA切割。因此，CysK作为一个平台，促进CdiA-CTEC536对底物tRNA的识别和切割。
