Quorum sensing and Chromobacterium violaceum:: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones
Quorum sensing and Chromobacterium violaceum:: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones
复制标题
DOI：
10.1099/00221287-143-12-3703
复制
发表时间：
1997-12-01
期刊：
影响因子：
--
通讯作者：
Williams, P
中科院分区：
文献类型：
--
作者：
McClean, KH;Winson, MK;Williams, P
群体感应依赖于可扩散信号分子与转录激活蛋白的相互作用，以将基因表达与细胞群体密度偶联。在Cram阴性细菌中，这样的信号分子通常是N-酰基高丝氨酸内酯（AHL），其N-酰基侧链的结构不同。紫色色杆菌（Chromobacterium violaceum）是一种常见于土壤和水体中的革兰氏阴性细菌，产生紫色素。以前，作者描述了一种紫色素阴性的C.其中色素生产可以通过与野生型菌株的上清液温育来恢复。将该突变体作为AHLs的通用生物传感器。天然C. violaceun，阿勒分子首次被化学表征。通过溶剂萃取、高效液相色谱和质谱分析，从野生型C. CV 026中不存在的violaceum培养物上清液。由于紫色素的产生构成了用于检测AHL的简单测定，因此我们探索了CV 026对一系列合成阿勒和N-酰基高半胱氨酸硫内酯（ART）类似物的响应能力。在CV 026中，所有阿勒和AHT化合物均可诱导紫色菌素，这些化合物的N-酰基侧链长度为C-4至C-8，敏感性不同。尽管具有C-10至C-14的N-酰基侧链的阿勒化合物不能诱导紫素产生，但如果将活化阿勒（例如HHL）掺入琼脂中，则可以通过其抑制紫素产生的能力来检测这些长链AHL。CV 026在促进阿勒混合物从培养上清液中提取和薄层色谱分离检测的多功能性也得到了证明。因此，这些采用CV 026的简单生物测定大大扩展了检测广谱阿勒信号分子的能力。
Quorum sensing relies upon the interaction of a diffusible signal molecule with a transcriptional activator protein to couple gene expression with cell population density. In Cram-negative bacteria, such signal molecules are usually N-acylhomoserine lactones (AHLs) which differ in the structure of their N-acyl side chains. Chromobacterium violaceum, a Cram-negative bacterium commonly found in soil and water, produces the characteristic purple pigment violacein. Previously the authors described a violacein-negative, mini-Tn5 mutant of C. violaceum (CV026) in which pigment production can be restored by incubation with supernatants from the wild-type strain. To develop this mutant as a general biosensor for AHLs. the natural C. violaceun, AHL molecule was first chemically characterized. By using solvent extraction, HPLC and mass spectrometry, a single AHL, N-hexanoyl-L-homoserine lactone (HHL), was identified in wild-type C. violaceum culture supernatants which was absent from CV026. Since the production of violacein constitutes a simple assay for the detection of AHLs, we explored the ability of CV026 to respond to a series of synthetic AHL and N-acylhomocysteine thiolactone (ART) analogues. In CV026, violacein is inducible by all the AHL and AHT compounds evaluated with N-acyl side chains from C-4 to C-8 in length, with varying degrees of sensitivity. Although AHL compounds with N-acyl side chains from C-10 to C-14 are unable to induce violacein production, if an activating AHL (e.g. HHL) is incorporated into the agar, these long-chain AHLs can be detected by their ability to inhibit violacein production. The versatility of CV026 in facilitating detection of AHL mixtures extracted from culture supernatants and separated by thin-layer chromatography is also demonstrated. These simple bioassays employing CV026 thus greatly extend the ability to detect a wide spectrum of AHL signal molecules.