N-acylhomoserine Lactones undergo lactonolysis in a pH-, temperature-, and acyl chain length-dependent manner during growth of Yersinia pseudotuberculosis and Pseudomonas aeruginosa
N-acylhomoserine Lactones undergo lactonolysis in a pH-, temperature-, and acyl chain length-dependent manner during growth of Yersinia pseudotuberculosis and Pseudomonas aeruginosa
复制标题
DOI：
10.1128/iai.70.10.5635-5646.2002
复制
发表时间：
2002-10-01
影响因子：
3.1
通讯作者：
Williams, P
中科院分区：
文献类型：
--
作者：
Yates, EA;Philipp, B;Williams, P
在革兰氏阴性细菌病原体中，例如铜绿假单胞菌和假结核耶尔森氏菌，通过N-酰基高丝氨酸内酯（阿勒）信号分子的细胞间通讯参与了与毒力相关的基因的细胞群体密度依赖性控制。这种现象，称为群体感应，依赖于AHL的积累到一个阈值浓度，在该浓度下靶结构基因被激活。通过使用能够检测一系列AHLs的生物传感器，我们观察到，在Y。在假结核和铜绿假单胞菌中，AHLs在指数期积累，但在稳定期大量消失。当加入到稳定期后期，各病原体的无细胞培养上清液中时，主要的铜绿假单胞菌[N-丁酰基高丝氨酸内酯（C4-HSL）和N-（3-氧代十二烷酰基）高丝氨酸内酯（3-氧代-C12-HSL）]和Y.假结核[N-（3-氧代己酰基）高丝氨酸内酯（3-氧代-C6-HSL）和N-己酰基高丝氨酸内酯（C6-HSL）] AHL失活。短酰基链化合物（例如，C4-HSL）比长链分子（例如，3-氧代-C12-HSL）。细胞提取物几乎没有发生阿勒失活，也没有明显的特定酶失活的证据。这阿勒营业额被发现是由于pH依赖性的内酯分解。通过将生长培养基酸化至pH 2.0，可以逆转内酯分解。通过使用碳-13核磁共振谱，我们发现，高丝氨酸内酯（HSL），N-丙酰基HSL（C3-HSL），和C4-HSL的开环随着pH值的增加而增加，但随着N-酰基链的延长而减少。在低pH水平下，内酯环闭合，但不是通过开环反应机制的简单逆转。C4-HSL、C6-HSL、3-氧代-C6-HSL和N-辛酰基高丝氨酸内酯（C8-HSL）的开环（如通过水溶液中的pH随时间降低所确定的）对于具有更多供电子的较长侧链的AHL也不太快。将温度从22 ℃升高到37 ℃增加了开环速率。总之，这些数据表明：（i）为了在哺乳动物组织液中在生理条件下发挥功能，AHL需要长度为至少四个碳的N-酰基侧链，和（ii）酰基侧链越长，阿勒信号分子越稳定。
In gram-negative bacterial pathogens, such as Pseudomonas aeruginosa and Yersinia pseudotuberculosis, cell-to-cell communication via the N-acylhomoserine lactone (AHL) signal molecules is involved in the cell population density-dependent control of genes associated with virulence. This phenomenon, termed quorum sensing, relies upon the accumulation of AHLs to a threshold concentration at which target structural genes are activated. By using biosensors capable of detecting a range of AHLs we observed that, in cultures of Y. pseudotuberculosis and P. aeruginosa, AHLs accumulate during the exponential phase but largely disappear during the stationary phase. When added to late-stationary-phase, cell-free culture supernatants of the respective pathogen, the major P. aeruginosa [N-butanoylhomoserine lactone (C4-HSL) and N-(3-oxododecanoyl)homoserine lactone (3-oxo-C12-HSL)] and Y. pseudotuberculosis [N-(3-oxohexanoyl)homoserine lactone (3-oxo-C6-HSL) and N-hexanoylhomoserine lactone (C6-HSL)] AHLs were inactivated. Short-acyl-chain compounds (e.g., C4-HSL) were turned over more extensively than long-chain molecules (e.g., 3-oxo-C12-HSL). Little AHL inactivation occurred with cell extracts, and no evidence for inactivation by specific enzymes was apparent. This AHL turnover was discovered to be due to pH-dependent lactonolysis. By acidifying the growth media to pH 2.0, lactonolysis could be reversed. By using carbon-13 nuclear magnetic resonance spectroscopy, we found that the ring opening of homoserine lactone (HSL), N-propionyl HSL (C3-HSL), and C4-HSL increased as pH increased but diminished as the N-acyl chain was lengthened. At low pH levels, the lactone rings closed but not via a simple reversal of the ring opening reaction mechanism. Ring opening of C4-HSL, C6-HSL, 3-oxo-C6-HSL, and N-octanoylhomoserine lactone (C8-HSL), as determined by the reduction of pH in aqueous solutions with time, was also less rapid for AHLs with more electron-donating longer side chains. Raising the temperature from 22 to 37 C increased the rate of ring opening. Taken together, these data show that (i) to be functional under physiological conditions in mammalian tissue fluids, AHLs require an N-acyl side chain of at least four carbons in length and (ii) that the longer the acyl side chain the more stable the AHL signal molecule.