title: Design and application of a lactulose biosensor.
abstract: In this study the repressor of Escherichia coli lac operon, LacI, has been engineered for altered effector specificity. A LacI saturation mutagenesis library was subjected to Fluorescence Activated Cell Sorting (FACS) dual screening. Mutant LacI-L5 was selected and it is specifically induced by lactulose but not by other disaccharides tested (lactose, epilactose, maltose, sucrose, cellobiose and melibiose). LacI-L5 has been successfully used to construct a whole-cell lactulose biosensor which was then applied in directed evolution of cellobiose 2-epimerase (C2E) for elevated lactulose production. The mutant C2E enzyme with ~32-fold enhanced expression level was selected, demonstrating the high efficiency of the lactulose biosensor. LacI-L5 can also be used as a novel regulatory tool. This work explores the potential of engineering LacI for customized molecular biosensors which can be applied in practice.
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.|University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Jieyuan
authors.initials: J
authors.lastname: Wu
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Peixia
authors.initials: P
authors.lastname: Jiang
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.|University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Dandan
authors.initials: D
authors.lastname: Xiong
authors.affiliation: Beijing Key Laboratory of Plant Resources Research and Development, Beijing Technology and Business University, Beijing 100048, China.
authors.forename: Linglan
authors.initials: L
authors.lastname: Huang
authors.affiliation: Core Facility for Protein Research, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Junying
authors.initials: J
authors.lastname: Jia
authors.affiliation: Core Facility for Protein Research, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Beijing Key Laboratory of Plant Resources Research and Development, Beijing Technology and Business University, Beijing 100048, China.
authors.forename: Jian-Ming
authors.initials: JM
authors.lastname: Jin
authors.affiliation: CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Shuang-Yan
authors.initials: SY
authors.lastname: Tang
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation*; D018360:Crystallography, X-Ray; D004867:Equipment Design; D005818:Genetic Engineering; D007763:Lac Operon / Q000235:genetics; D056968:Lac Repressors / Q000235:genetics; D007792:Lactulose / Q000032:analysis*; D016296:Mutagenesis / Q000235:genetics; D009154:Mutation / Q000235:genetics
chemical_list: D056968:Lac Repressors; D007792:Lactulose