Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
Frontier Science Center for Synthetic Biology, Key Laboratory of Systems Bioengineering (MOE), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
Biotechnol Bioeng. 2020 Dec;117(12):3820-3834. doi: 10.1002/bit.27525. Epub 2020 Sep 18.
Cell surface display of heavy metal-binding proteins has been used to enhance the adsorption capacity of heavy metals and the engineered microbial cells can be potentially used for the bioremediation of heavy metals. In this study, the proteins PbrR, PbrR691, and PbrD from the Cupriavidus metallidurans strain CH34 were displayed on the extracellular membrane of Escherichia coli BL21 cells, with the N-domain of ice-nucleation protein as the anchor protein to achieve specific adsorption of lead ions (Pb ) and bioremediation of lead in the soil. The localization of fusion proteins was confirmed by western blot analysis. We investigated the effects of fusion pattern, expression level, heavy metal concentration, and the presence of other heavy metal ions on the adsorption of Pb by these engineered bacteria, and the optimal linker peptide (flexible linker) and inducer concentration (0.5 mM) were obtained. The engineered bacteria showed specific selectivity and strong adsorption capacity for Pb . The maximum Pb adsorption capacity of strains displaying the three proteins (PbrR, PbrR691, and PbrD) were 942.1-, 754.3-, and 864.8-μmol/g cell dry weight, respectively, which was the highest reported to date. The engineered E. coli bacteria were also applied to Pb -contaminated soil and the detoxification effects were observed via the seed germination test and the growth of Nicotiana benthamiana in comparison with the control BL21, which provides the proof-of-concept for in situ remediations of Pb -contaminated water or soil.
已经利用重金属结合蛋白的细胞表面展示来增强重金属的吸附能力，并且经过工程改造的微生物细胞可潜在用于重金属的生物修复。在这项研究中，来自铜绿假单胞菌 CH34 菌株的 PbrR、PbrR691 和 PbrD 蛋白被展示在大肠杆菌 BL21 细胞的细胞外膜上，冰核蛋白的 N 结构域作为锚定蛋白，以实现对铅离子 (Pb ) 的特异性吸附和对土壤中铅的生物修复。融合蛋白的定位通过 Western blot 分析得到确认。我们研究了融合模式、表达水平、重金属浓度以及其他重金属离子的存在对这些工程菌吸附 Pb 的影响，并获得了最佳的连接肽（柔性接头）和诱导剂浓度（0.5 mM）。这些工程菌对 Pb 表现出特异性选择性和强大的吸附能力。展示这三种蛋白（PbrR、PbrR691 和 PbrD）的菌株的最大 Pb 吸附容量分别为 942.1、754.3 和 864.8 μmol/g 细胞干重，这是迄今为止报道的最高值。这些工程大肠杆菌菌也被应用于 Pb 污染土壤中，并通过种子发芽试验和对 Nicotiana benthamiana 的生长与对照 BL21 进行比较观察到了解毒效果，这为 Pb 污染水或土壤的原位修复提供了概念验证。
