Structural insights into the inhibition mechanism of bacterial toxin LsoA by bacteriophage antitoxin Dmd
Structural insights into the inhibition mechanism of bacterial toxin LsoA by bacteriophage antitoxin Dmd
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噬菌体抗毒素Dmd抑制细菌毒素LsoA机制的结构见解
DOI：
10.1111/mmi.13420
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发表时间：
2016-09-01
影响因子：
3.6
通讯作者：
Dong, Yu-Hui
中科院分区：
文献类型：
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作者：
Wan, Hua;Otsuka, Yuichi;Dong, Yu-Hui
细菌已经获得了多种抗性机制，包括毒素-抗毒素(TA)系统对噬菌体的抗性，而噬菌体也进化成克服细菌抗噬菌体机制。来自T4噬菌体的DMD可以抑制来自大肠杆菌的同源毒素LsoA和RNLA的毒性，这是已知TA系统中第一个针对多种细菌毒素的噬菌体抗毒素的例子。LsoA-DMD复合体的晶体结构表明，DMD被插入到LsoA的N-末端重复结构域(NRD)和DMD结合结构域(DBD)之间的深槽中。当DMD结合时，NRD从封闭的构象显著转变为开放的构象。DMD的定点突变表明，保守残基(W31和N40)是LsoA结合所必需的，通过下拉和细胞毒性试验确定了毒性抑制。进一步的诱变实验证明，LsoA的DMD结合残基(R243、E246和R305)是其毒性所必需的，提示DMD和LsoB可能具有不同的抑制LsoA毒性的机制。我们的结构-功能研究表明，DMD可以识别LsoA，并可能通过底物模仿占据活性部位来抑制其毒性。这些发现为细菌和噬菌体在共同进化中的防御和反防御机制提供了独特的见解。
Bacteria have obtained a variety of resistance mechanisms including toxin-antitoxin (TA) systems against bacteriophages (phages), whereas phages have also evolved to overcome bacterial anti-phage mechanisms. Dmd from T4 phage can suppress the toxicities of homologous toxins LsoA and RnlA from Escherichia coli, representing the first example of a phage antitoxin against multiple bacterial toxins in known TA systems. Here, the crystal structure of LsoA-Dmd complex showed Dmd is inserted into the deep groove between the N-terminal repeated domain (NRD) and the Dmd-binding domain (DBD) of LsoA. The NRD shifts significantly from a 'closed' to an 'open' conformation upon Dmd binding. Site-directed mutagenesis of Dmd revealed the conserved residues (W31 and N40) are necessary for LsoA binding and the toxicity suppression as determined by pull-down and cell toxicity assays. Further mutagenesis identified the conserved Dmd-binding residues (R243, E246 and R305) of LsoA are vital for its toxicity, and suggested Dmd and LsoB may possess different inhibitory mechanisms against LsoA toxicity. Our structure-function studies demonstrate Dmd can recognize LsoA and inhibit its toxicity by occupying the active site possibly via substrate mimicry. These findings have provided unique insights into the defense and counter-defense mechanisms between bacteria and phages in their co-evolution.