Department of Pathology & Toxicology, Shenzhen Prevention and Treatment Center for Occupational Diseases, Shenzhen, China.
National Key Clinical Specialty of Occupational Diseases, Shenzhen Prevention and Treatment Center for Occupational Diseases, Shenzhen, China.
Appl Microbiol Biotechnol. 2021 Aug;105(14-15):6087-6102. doi: 10.1007/s00253-021-11441-5. Epub 2021 Jul 22.
During the last few decades, whole-cell biosensors have attracted increasing attention for their enormous potential in monitoring bioavailable heavy metal contaminations in the ecosystem. Visual and measurable output signals by employing natural pigments have been demonstrated to offer another potential choice to indicate the existence of bioavailable heavy metals in recent years. The biosynthesis of the blue pigment indigoidine has been achieved in E. coli following heterologous expression of both BpsA (a single-module non-ribosomal peptide synthetase) and PcpS (a PPTase to activate apo-BpsA). Moreover, we demonstrated herein the development of the indigoidine-based whole-cell biosensors to detect bioavailable Hg(II) and Pb(II) in water samples by employing metal-responsive transcriptional regulator MerR and PbrR as the sensory elements, and the indigoidine biosynthesis gene cluster as a reporter element. The resulting indigoidine-based biosensors presented a good selectivity and high sensitivity to target metal ions. High concentration of target metal exposure could be clearly recognized by the naked eye due to the color change by the secretion of indigoidine, and quantified by measuring the absorbance of the culture supernatants at 600 nm. Dose-response relationships existed between the exposure concentrations of target heavy metals and the production of indigoidine. Although fairly good linear relationships were obtained in a relatively limited concentration range of the concentrations of heavy metal ions, these findings suggest that genetically controlled indigoidine biosynthesis triggered by the MerR family transcriptional regulator can enable a sensitive, visual, and qualitative whole-cell biosensor for bioindicating the presence of bioaccessible heavy metal in environmental water samples. KEY POINTS: • Biosynthesis pathway of indigoidine reconstructed in a high copy number plasmid in E. coli. • Visual and colorimetric detection of Hg(II) and Pb(II) by manipulation of indigoidine biosynthesis through MerR family metalloregulator. •Enhanced detection sensitivity toward Hg(II) and Pb(II) achieved using novel pigment-based whole-cell biosensors.
在过去的几十年中，全细胞生物传感器因其在监测生态系统中生物可利用重金属污染方面的巨大潜力而受到越来越多的关注。近年来，利用天然色素产生可见和可测量的输出信号已被证明是另一种潜在的选择，可以表明生物可利用重金属的存在。在大肠杆菌中，通过异源表达 BpsA（一种单模块非核糖体肽合成酶）和 PcpS（一种激活 apo-BpsA 的 PPTase），实现了蓝色色素靛蓝的生物合成。此外，我们在此展示了基于靛蓝的全细胞生物传感器的开发，该传感器通过使用金属反应转录调节剂 MerR 和 PbrR 作为感应元件，以及靛蓝生物合成基因簇作为报告元件，用于检测水样中的生物可利用 Hg(II)和 Pb(II)。所得到的基于靛蓝的生物传感器对目标金属离子表现出良好的选择性和高灵敏度。由于靛蓝的分泌导致颜色变化，高浓度的目标金属暴露可以用肉眼清楚地识别，并通过测量培养上清液在 600nm 处的吸光度进行定量。目标重金属暴露浓度与靛蓝产量之间存在剂量-反应关系。尽管在重金属离子浓度的相对有限浓度范围内获得了相当好的线性关系，但这些发现表明，通过 MerR 家族转录调节剂触发的遗传控制的靛蓝生物合成可以实现一种敏感、可见和定性的全细胞生物传感器，用于生物指示环境水样中生物可利用的重金属的存在。关键点：• 在大肠杆菌中以高拷贝数质粒重建靛蓝的生物合成途径。• 通过 MerR 家族金属调节剂对靛蓝生物合成的操纵进行 Hg(II)和 Pb(II)的可视化和比色检测。• 使用新型色素全细胞生物传感器实现对 Hg(II)和 Pb(II)的检测灵敏度提高。
