title: Biosensor-Based Evolution and Elucidation of a Biosynthetic Pathway in Escherichia coli.
abstract: The successful evolution of metabolite-producing microbes requires a high-throughput screening method to obtain the desired properties within a short time. In this study, we developed a transcription-factor-driven device that combines a metabolite-responsive element and a selection module. This device was able to specifically sense intracellular l-phenylalanine (l-Phe) and convert this signal into an observable phenotype. Applying this device, we successfully improved l-Phe production by screening hyperproducing phenotypes from a ribonucleotide binding site library and a random mutagenesis library. In addition, several site mutations introduced by random mutagenesis were identified and elucidated to facilitate the improvement of l-Phe production. Our results present a paradigm for screening of compounds that are not easily observable to raise the yield of targeted compounds from a large candidate library. This approach may guide further applications in rewiring metabolic circuits and facilitate the directed evolution of recombinant strains.
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.
authors.forename: Yongfei
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.
authors.forename: Yinyin
authors.initials: Y
authors.lastname: Zhuang
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.
authors.forename: Dongqin
authors.initials: D
authors.lastname: Ding
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.
authors.forename: Yiran
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.
authors.forename: Jibin
authors.initials: J
authors.lastname: Sun
authors.affiliation: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.|Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences , Tianjin 300308, China.
authors.forename: Dawei
authors.identifier: 0000-0002-5857-394X
authors.initials: D
authors.lastname: Zhang
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D053898:Biosynthetic Pathways / Q000235:genetics / Q000502:physiology; D019020:Directed Molecular Evolution / Q000379:methods; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods; D010649:Phenylalanine / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D014157:Transcription Factors; D010649:Phenylalanine
keywords: TyrR; biosensor; high-throughput screening; l-phenylalanine; transcription factor