Structure-function analyses reveal the molecular architecture and neutralization mechanism of a bacterial HEPN-MNT toxin-antitoxin system
Structure-function analyses reveal the molecular architecture and neutralization mechanism of a bacterial HEPN-MNT toxin-antitoxin system
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结构-功能分析揭示了细菌HEPN-MNT毒素-抗毒素系统的分子结构和中和机制
DOI：
10.1074/jbc.ra118.002421
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发表时间：
2018-05-04
影响因子：
4.8
通讯作者：
Zhang, Heng
中科院分区：
文献类型：
--
作者：
Jia, Xuanyan;Yao, Jianyun;Zhang, Heng
毒素-抗毒素（TA）基因座是细菌中调节各种细胞活动（包括细胞生长和死亡）的小遗传模块。已预测编码HEPN（高等真核生物和原核生物核苷酸结合）结构域和同源MNT（最小核苷酸转移酶）结构域的双基因模块代表古细菌和细菌中普遍存在的新型II型TA系统。然而，TA家族的中和机制和细胞靶点仍不清楚。具有HEPN结构域的毒素SO_3166及其具有MNT结构域的同源抗毒素SO_3165构成了典型的II型TA系统，其调节细菌希瓦氏菌中的细胞运动性并赋予质粒稳定性。在这里，我们报告的晶体结构和溶液构象的SO_3166-SO_3165对，代表第一个复杂的结构，在这个TA家庭。结构表明，SO_3165和SO_3166形成紧密的杂八聚体（以2：6的比例），这是在其他TA系统中非常罕见的组织。我们还观察到，SO_3166二聚化使得能够在HEPN结构域界面处形成深裂缝，该裂缝具有复合RX 4 - 6 H活性位点，该复合RX 4 - 6 H活性位点作为RNA切割RNA酶发挥作用。SO_3165与SO_3166的结合主要是通过其双螺旋（2和4），起分子识别元件的作用.此外，它们插入SO_3166裂缝空间阻断RX 4 - 6 H位点或缩小裂缝以抑制RNA底物结合。基于结构的突变证实了这些-螺旋在SO_3166结合和抑制中的重要作用。我们的结构-功能分析首次深入了解了HEPN-MNT TA家族的中和机制。
Toxin-antitoxin (TA) loci in bacteria are small genetic modules that regulate various cellular activities, including cell growth and death. The two-gene module encoding a HEPN (higher eukaryotes and prokaryotes nucleotide-binding) domain and a cognate MNT (minimal nucleotidyltransferase) domain have been predicted to represent a novel type II TA system prevalent in archaea and bacteria. However, the neutralization mechanism and cellular targets of the TA family remain unclear. The toxin SO_3166 having a HEPN domain and its cognate antitoxin SO_3165 with an MNT domain constitute a typical type II TA system that regulates cell motility and confers plasmid stability in the bacterium Shewanella oneidensis. Here, we report the crystal structure and solution conformation of the SO_3166-SO_3165 pair, representing the first complex structures in this TA family. The structures revealed that SO_3165 and SO_3166 form a tight heterooctamer (at a 2:6 ratio), an organization that is very rare in other TA systems. We also observed that SO_3166 dimerization enables the formation of a deep cleft at the HEPN-domain interface harboring a composite RX4-6H active site that functions as an RNA-cleaving RNase. SO_3165 bound SO_3166 mainly through its two -helices (2 and 4), functioning as molecular recognition elements. Moreover, their insertion into the SO_3166 cleft sterically blocked the RX4-6H site or narrowed the cleft to inhibit RNA substrate binding. Structure-based mutagenesis confirmed the important roles of these -helices in SO_3166 binding and inhibition. Our structure-function analysis provides first insights into the neutralization mechanism of the HEPN-MNT TA family.