Hinnebusch J, Cherepanov P, Du Y, Rudolph A, Dixon J D, Schwan T, Forsberg A
Rocky Mountain Laboratories, Hamilton, MT, USA.
Int J Med Microbiol. 2000 Oct;290(4-5):483-7. doi: 10.1016/S1438-4221(00)80070-3.
Purified murine toxin (Ymt) of Yersinia pestis is highly toxic for mice and rats but less active in other animals such as guinea pigs, rabbits, dogs and monkeys. This suggested that Ymt contributes to the very low infectious dose of Y. pestis in mice. The gene encoding Ymt (ymt) is localised on the 100-kb plasmid pFra, which is unique for Y. pestis. Sequence analysis revealed that Ymt showed homology to proteins of the phospholipase D (PLD) superfamily of proteins. Y. pestis strains expressing Ymt possessed PLD activity whereas strains carrying deletions in the ymt gene showed no detectable PLD activity. Western blot analysis showed that Ymt was associated with bacteria under normal growth conditions, and immunogold EM revealed that Ymt was mainly localised in the bacterial cytoplasm. Ymt was purified to homogeneity, and the purified toxin showed a dose-dependent PLD activity. Substitution of amino acids in the PLD consensus motif of Ymt essentially abolished the enzymatic activity and these variants of the toxin were no longer toxic to mice. Interestingly, an in-frame deletion mutant of ymt in the Y pestis strain KIM was not significantly attenuated for mouse virulence. Together with the observation that expression of Ymt was higher at room temperature compared to 37 degrees C this prompted us to investigate the role of Ymt in the flea vector. Fleas were infected with isogenic ymt+ or ymt- mutant strains of Y. pestis. Preliminary results suggest that Ymt is important for survival of Y. pestis in the flea and thereby also for the flea-borne route of infection.
鼠疫耶尔森菌的纯化鼠毒素（Ymt）对小鼠和大鼠具有高毒性，但在豚鼠、兔子、狗和猴子等其他动物中活性较低。这表明Ymt促成了鼠疫耶尔森菌在小鼠中极低的感染剂量。编码Ymt的基因（ymt）定位于100 kb的质粒pFra上，这是鼠疫耶尔森菌所特有的。序列分析表明，Ymt与磷脂酶D（PLD）超家族的蛋白质具有同源性。表达Ymt的鼠疫耶尔森菌菌株具有PLD活性，而在ymt基因中携带缺失的菌株则未检测到PLD活性。蛋白质印迹分析表明，Ymt在正常生长条件下与细菌相关，免疫金电镜显示Ymt主要定位于细菌细胞质中。Ymt被纯化至同质，纯化后的毒素显示出剂量依赖性的PLD活性。Ymt的PLD共有基序中的氨基酸替换基本上消除了酶活性，并且这些毒素变体对小鼠不再有毒性。有趣的是，鼠疫耶尔森菌菌株KIM中ymt的框内缺失突变体对小鼠毒力没有明显减弱。结合Ymt在室温下的表达高于37℃这一观察结果，这促使我们研究Ymt在跳蚤载体中的作用。用鼠疫耶尔森菌的同基因ymt +或ymt-突变菌株感染跳蚤。初步结果表明，Ymt对鼠疫耶尔森菌在跳蚤中的存活很重要，因此对跳蚤传播的感染途径也很重要。
