Kawano Hiroaki, Hirokawa Yasutaka, Mori Hideo
Biofrontier Laboratories, Kyowa Hakko Kogyo Co., Ltd.
Biosci Biotechnol Biochem. 2009 Jan;73(1):117-23. doi: 10.1271/bbb.80531. Epub 2009 Jan 7.
We designed and constructed six major toxin-antitoxin disruptants (DeltachpBIK, DeltadinJ-yafQ, DeltahipBA, DeltamazEF, DeltarelBE, and DeltayefM-yoeB) of Escherichia coli K-12 W3110. On prolonged cultivation of these disruptants with minimal M9 medium, the DeltahipBA cells exhibited a significantly longer life span than that of the other disruptants and of wild-type cells, as analyzed with a LIVE/DEAD BacLight kit (Invitrogen, Carlsbad, CA) in combination with flow cytometry analysis. The gene expression level of hipA in the wild-type cells was highest at the stationary phase of 40 h. The DeltahipBA cells showed higher macromolecular synthesis activity than the wild-type cells at the stationary phase. Stationary phase cells of DeltahipBA and the wild-type strain showed a significantly extended life span under anaerobic conditions. Furthermore, the DeltahipBA cells showed higher resistance to H(2)O(2) than the wild type. These results suggest that HipBA induces cell death with oxidative stress during prolonged cultivation. This is the first report that an E. coli toxin-antitoxin (TA) system affects frequency of survival during the long-term stationary phase.
我们设计并构建了大肠杆菌K-12 W3110的六种主要毒素-抗毒素破坏株（DeltachpBIK、DeltadinJ-yafQ、DeltahipBA、DeltamazEF、DeltarelBE和DeltayefM-yoeB）。在用基本M9培养基对这些破坏株进行长时间培养时，使用LIVE/DEAD BacLight试剂盒（英杰公司，加利福尼亚州卡尔斯巴德）并结合流式细胞术分析，发现DeltahipBA细胞的寿命显著长于其他破坏株及野生型细胞。野生型细胞中hipA的基因表达水平在40小时的稳定期最高。DeltahipBA细胞在稳定期的大分子合成活性高于野生型细胞。在厌氧条件下，DeltahipBA和野生型菌株的稳定期细胞的寿命显著延长。此外，DeltahipBA细胞对H₂O₂的抗性高于野生型。这些结果表明，在长时间培养过程中，HipBA会因氧化应激诱导细胞死亡。这是关于大肠杆菌毒素-抗毒素（TA）系统影响长期稳定期存活频率的首次报道。
