title: An Escherichia coli system for evolving improved light-controlled DNA-binding proteins.
abstract: Light-switchable proteins offer numerous opportunities as tools for manipulating biological systems with exceptional degrees of spatiotemporal control. Most designed light-switchable proteins currently in use have not been optimised using the randomisation and selection/screening approaches that are widely used in other areas of protein engineering. Here we report an approach for screening light-switchable DNA-binding proteins that relies on light-dependent repression of the transcription of a fluorescent reporter. We demonstrate that the method can be used to recover a known light-switchable DNA-binding protein from a random library.
authors.affiliation: Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Rd. N., Mississauga, Ontario, Canada L5L 1C6.
authors.forename: Mostafizur
authors.initials: M
authors.lastname: Mazumder
authors.affiliation: Department of Chemistry, University of Toronto, 80 Saint George St, Toronto M5S 3H6, Canada.
authors.forename: Katherine E
authors.initials: KE
authors.lastname: Brechun
authors.affiliation: Department of Chemistry, University of Toronto, 80 Saint George St, Toronto M5S 3H6, Canada.
authors.forename: Yongjoo B
authors.initials: YB
authors.lastname: Kim
authors.affiliation: Institute for Biochemistry and Biology, University of Potsdam, Karl-Liebknecht-Str. 24-25, Potsdam, Golm 14476, Germany.
authors.forename: Stefan A
authors.initials: SA
authors.lastname: Hoffmann
authors.affiliation: Department of Chemistry, University of Toronto, 80 Saint George St, Toronto M5S 3H6, Canada.
authors.forename: Yih Yang
authors.initials: YY
authors.lastname: Chen
authors.affiliation: Department of Chemistry, University of Toronto, 80 Saint George St, Toronto M5S 3H6, Canada.
authors.forename: Carrie-Lynn
authors.initials: CL
authors.lastname: Keiski
authors.affiliation: Institute for Biochemistry and Biology, University of Potsdam, Karl-Liebknecht-Str. 24-25, Potsdam, Golm 14476, Germany.
authors.forename: Katja M
authors.initials: KM
authors.lastname: Arndt
authors.affiliation: Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Rd. N., Mississauga, Ontario, Canada L5L 1C6 david.mcmillen@utoronto.ca awoolley@chem.utoronto.ca.
authors.forename: David R
authors.initials: DR
authors.lastname: McMillen
authors.affiliation: Department of Chemistry, University of Toronto, 80 Saint George St, Toronto M5S 3H6, Canada david.mcmillen@utoronto.ca awoolley@chem.utoronto.ca.
authors.forename: G Andrew
authors.initials: GA
authors.lastname: Woolley
mesh_terms: D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D008027:Light; D015202:Protein Engineering; D014158:Transcription, Genetic
chemical_list: D004268:DNA-Binding Proteins
keywords: directed evolution; fluorescent reporter; optogenetics