Modulating Sensitivity of an Erythromycin Biosensor for Precise High-Throughput Screening of Strains with Different Characteristics

生物传感器 合成生物学 高通量筛选 代谢工程 单元格排序 调节器 分析物 生物 计算生物学 纳米技术 材料科学 化学 分子生物学 生物信息学 生物化学 流式细胞术 色谱法 基因
作者
Yan Wang,Shixin Li,Ning Xue,Lixian Wang,Xuemei Zhang,Longqian Zhao,Yanmei Guo,Yue Zhang,Meng Wang
出处
期刊:ACS Synthetic Biology [American Chemical Society]
卷期号:12 (6): 1761-1771 被引量:9
标识
DOI:10.1021/acssynbio.3c00059
摘要

Genetically encoded biosensors are powerful tools for product-driven high-throughput screening in synthetic biology and metabolic engineering. However, most biosensors can only properly function in a limited concentration cutoff, and the incompatible performance characteristics of biosensors will lead to false positives or failure in screening. The transcription factor (TF)-based biosensors are usually organized in modular architecture and function in a regulator-depended manner, whose performance properties can be fine-tuned by modifying the expression level of the TF. In this study, we modulated the performance characteristics, including sensitivity and operating range, of an MphR-based erythromycin biosensor by fine-adjusting regulator expression levels via ribosome-binding site (RBS) engineering and obtained a panel of biosensors with varied sensitivities by iterative fluorescence-assisted cell sorting (FACS) in Escherichia coli to accommodate different screening purposes. To exemplify their application potential, two engineered biosensors with 10-fold different sensitivities were employed in the precise high-throughput screening by microfluidic-based fluorescence-activated droplet sorting (FADS) of Saccharopolyspora erythraea mutant libraries with different starting erythromycin productions, and mutants representing as high as 6.8 folds and over 100% of production improvements were obtained starting from the wild-type strain and the high-producing industrial strain, respectively. This work demonstrated a simple strategy to engineer biosensor performance properties, which was significant to stepwise strain engineering and production improvement.
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