Schaefer Amy L, Harwood Caroline S, Greenberg E Peter
Department of Microbiology, University of Washington, Seattle, WA, USA.
Methods Mol Biol. 2018;1673:35-47. doi: 10.1007/978-1-4939-7309-5_3.
Many Proteobacteria synthesize acyl-homoserine lactone (AHL) molecules for use as signals in cell density-dependent gene regulation known as quorum sensing (QS) and response. AHL detection protocols are essential to QS researchers and several techniques are available, including a C-AHL radiolabel assay. This assay is based on the uptake of radiolabeled methionine by living cells and conversion of the radiolabel into S-adenosylmethionine (SAM). The radiolabeled SAM is then incorporated into AHL signal by an AHL synthase enzyme. Here we describe a methodology to perform the AHL radiolabel assay, which is unbiased, relatively fast, and very sensitive compared to other AHL detection protocols.
许多变形菌门细菌合成酰基高丝氨酸内酯(AHL)分子,用作细胞密度依赖性基因调控(称为群体感应(QS)和应答)中的信号。AHL检测方案对QS研究人员至关重要,并且有几种技术可供使用,包括C-AHL放射性标记测定法。该测定法基于活细胞对放射性标记的甲硫氨酸的摄取以及将放射性标记转化为S-腺苷甲硫氨酸(SAM)。然后,放射性标记的SAM通过AHL合酶掺入AHL信号中。在这里,我们描述了一种进行AHL放射性标记测定的方法,与其他AHL检测方案相比,该方法无偏差、相对快速且非常灵敏。