生物传感器
酶
生物化学
合成生物学
大肠杆菌
细胞内
检出限
高通量筛选
化学
定向进化
生物
计算生物学
色谱法
基因
突变体
作者
Shengwei Sun,Kailin Peng,Sen Sun,Mengxi Wang,Yuting Shao,Longxiang Li,Jiahui Xiang,Rita-Cindy Aye-Ayire Sedjoah,Zhihong Xin
标识
DOI:10.1021/acssynbio.3c00036
摘要
Although a variety of whole-cell-based biosensors have been developed for different applications in recent years, most cannot meet practical requirements due to insufficient sensing performance. Here, we constructed two sets of modular genetic circuits by serial and parallel modes capable of significantly amplifying the input/output signal in Escherichia coli. The biosensors are engineered using σ54-dependent phenol-responsive regulator DmpR as a sensor and enhanced green fluorescent protein as a reporter. Cells harboring serial and parallel genetic circuits displayed nearly 9- and 16-fold higher sensitivity than the general circuit. The genetic circuits enabled rapid detection of six phenolic contaminants in 12 h and showed the low limit of detection of 2.5 and 2.2 ppb for benzopyrene (BaP) and tetracycline (Tet), with a broad detection range of 0.01-1 and 0.005-5 μM, respectively. Furthermore, the positive rate was as high as 73% when the biosensor was applied to screen intracellular enzymes with ester-hydrolysis activity from soil metagenomic libraries using phenyl acetate as a phenolic substrate. Several novel enzymes were isolated, identified, and biochemically characterized, including serine peptidases and alkaline phosphatase family protein/metalloenzyme. Consequently, this study provides a new signal amplification method for cell-based biosensors that can be widely applied to environmental contaminant assessment and screening of intracellular enzymes.
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