Department of Environmental Health Science, Konkuk University, Seoul 05029, Republic of Korea.
School of Biological Sciences and Technology, Chonnam National University, Gwangju 61186, Republic of Korea.
J Microbiol Biotechnol. 2020 May 28;30(5):681-688. doi: 10.4014/jmb.1911.11053.
Bacterial cell-based biosensors, or whole-cell bioreporters (WCBs), are an alternative tool for the quantification of hazardous materials. Most WCBs share similar working mechanisms. In brief, the recognition of a target by sensing domains induces a biological event, such as changes in protein conformation or gene expression, providing a basis for quantification. WCBs targeting heavy metal(loid)s employ metalloregulators as sensing domains and control the expression of genes in the presence of target metal(loid) ions, but the diversity of targets, specificity, and sensitivity of these WCBs are limited. In this study, we genetically engineered the metal-binding loop (MBL) of ZntR, which controls the znt-operon in . In the MBL of ZntR, three Cys sites interact with metal ions. Based on the crystal structure of ZntR, MBL sequences were modified by sitedirected mutagenesis. As a result, the metal-sensing properties of WCBs differed depending on amino acid sequences and the new selectivity to Cr or Pb was observed. Although there is room for improvement, our results support the use of currently available WCBs as a platform to generate new WCBs to target other environmental pollutants including metal(loid)s.
基于细菌细胞的生物传感器，或全细胞生物报告器（WCB），是定量危险物质的另一种工具。大多数 WCB 具有相似的工作机制。简而言之，感应域对目标的识别会引发生物事件，例如蛋白质构象或基因表达的变化，为定量提供基础。针对重金属（类）的 WCB 采用金属调控器作为感应域，并在存在目标金属（类）离子的情况下控制基因的表达，但这些 WCB 的目标多样性、特异性和灵敏度有限。在这项研究中，我们对控制 中 znt 操纵子的 ZntR 的金属结合环（MBL）进行了基因工程改造。在 ZntR 的 MBL 中，三个 Cys 位点与金属离子相互作用。基于 ZntR 的晶体结构，通过定点突变修饰了 MBL 序列。结果，WCB 的金属感应特性因氨基酸序列而异，并观察到对 Cr 或 Pb 的新选择性。尽管还有改进的空间，但我们的结果支持将现有的 WCB 用作平台来生成针对其他环境污染物（包括金属（类））的新型 WCB。
