Structure-function studies of Escherichia coli RnlA reveal a novel toxin structure involved in bacteriophage resistance
Structure-function studies of Escherichia coli RnlA reveal a novel toxin structure involved in bacteriophage resistance
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大肠杆菌 RnlA 的结构功能研究揭示了一种参与噬菌体抗性的新型毒素结构
DOI：
10.1111/mmi.12409
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发表时间：
2013-12-01
影响因子：
3.6
通讯作者：
Dong, Yu-Hui
中科院分区：
文献类型：
--
作者：
Wei, Yong;Gao, Zeng-Qiang;Dong, Yu-Hui
大肠杆菌RNLA-RNLB是新近发现的一种毒素-抗毒素(TA)系统，在噬菌体抵抗中起作用。RNLA是一种具有mRNA内切核酸酶活性的毒素，同源抗毒素RNLB可抑制RNLA对大肠杆菌细胞的毒性。有趣的是，T4噬菌体编码抗毒素DMD，DMD作用于RNLA促进自身繁殖，这表明RNLA-DMD代表了一个新的TA系统。在这里，我们已经确定了精制到2.10埃的RNLA的晶体结构。RNLA由三个独立的结构域组成：NTD(N-末端结构域)、NRD(N重复结构域)和DBD(DMD结合结构域)，这是已知毒素结构中尚未观察到的一个组织。小角X射线散射(SAXS)分析表明，RNLA通过两个单体的DBD之间的相互作用在溶液中形成了二聚体。体外和体内功能研究表明，在这三个结构域中，只有DBD负责DMD的识别和抑制以及RNLA的亚细胞定位。尤其是位于DBD C末端的螺旋在结合DMD中起着至关重要的作用。我们的全面研究揭示了RNLA毒性的关键区域，并为其结构与功能的关系提供了新的见解。
Escherichia coli RnlA-RnlB is a newly identified toxin-antitoxin (TA) system that plays a role in bacteriophage resistance. RnlA functions as a toxin with mRNA endoribonuclease activity and the cognate antitoxin RnlB inhibits RnlA toxicity in E. coli cells. Interestingly, T4 phage encodes the antitoxin Dmd, which acts against RnlA to promote its own propagation, suggesting that RnlA-Dmd represents a novel TA system. Here, we have determined the crystal structure of RnlA refined to 2.10 angstrom. RnlA is composed of three independent domains: NTD (N-terminal domain), NRD (N repeated domain) and DBD (Dmd-binding domain), which is an organization not previously observed among known toxin structures. Small-angle X-ray scattering (SAXS) analysis revealed that RnlA forms a dimer in solution via interactions between the DBDs from both monomers. The in vitro and in vivo functional studies showed that among the three domains, only the DBD is responsible for recognition and inhibition by Dmd and subcellular location of RnlA. In particular, the helix located at the C-terminus of DBD plays a vital role in binding Dmd. Our comprehensive studies reveal the key region responsible for RnlA toxicity and provide novel insights into its structure-function relationship.