title: Optimization of a genetically encoded biosensor for cyclin B1-cyclin dependent kinase 1.
abstract: Fluorescent protein (FP)-based biosensors have revolutionized the ability of researchers to monitor enzyme activities in live cells. While the basic design principles for FP-based biosensors are well established, first-generation biosensor constructs typically suffer from relatively low fluorescence responses that limit their general applicability. The protein engineering efforts required to substantially improve the biosensor responses are often both labour and time intensive. Here we report the application of a high throughput bacterial colony screen for improving the response of kinase biosensors. This effort led to the development of a second-generation cyclin B1-CDK1 biosensor with a 4.5-fold greater response than the first-generation biosensor.
authors.affiliation: Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2, Canada. robert.e.campbell@ualberta.ca.
authors.forename: Ahmed Saied F
authors.initials: AS
authors.lastname: Belal
authors.forename: Brittney R
authors.initials: BR
authors.lastname: Sell
authors.forename: Hiofan
authors.initials: H
authors.lastname: Hoi
authors.forename: Michael W
authors.initials: MW
authors.lastname: Davidson
authors.forename: Robert E
authors.initials: RE
authors.lastname: Campbell
mesh_terms: D015374:Biosensing Techniques; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism*; D056744:Cyclin B1 / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D006367:HeLa Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008938:Mitosis; D015202:Protein Engineering
chemical_list: D056744:Cyclin B1; D008164:Luminescent Proteins; D016203:CDC2 Protein Kinase