434630 Development of Biosensors to Identify New Chemicals Against Dengue Fever Vectors

Thursday, November 12, 2015: 2:10 PM
151A/B (Salt Palace Convention Center)
Miriam Shakalli Tang, Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH and David W. Wood, Department of Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH

The development of a bacterial biosensor containing a nuclear hormone receptor (NHR) has proven to be an effective alternative to current compound screening strategies that rely on hormone-mimicking properties. NHRs comprise one of the largest classes of protein targets because they can bind to a wide variety of endocrine disruptors in addition to their native hormones.  Our allosteric biosensors can detect compounds that target NHRs and distinguish between their positive or negative effects on the receptor’s activity.  When this design is extended from human to insect NHRs, an opportunity arises to screen compounds for their potential role as insecticides against infectious disease vectors such as Aedes aegypti, the dengue fever mosquito. This project seeks to construct a nuclear hormone binding sensor in E. coli to favor the identification of those compounds that disrupt the endocrine system of this target insect. By incorporating the ligand-binding domain of  Aedes aegypti’s NHR 96 into our E. coli biosensing plasmid, we are able to detect its response to CITCO, a chemical compound known to be the NHR96 native ligand. After a thorough optimization of parameters such as incubation time, temperature, and dilution in a full factorial experimental setup, we seek to attain a reproducible growth phenotype assay for this novel biosensing system. The  use of domain deletion, linker engineering, and point mutations can help improve the biosensor’s ligand-response ability. Furthermore, the translation of the biosensing protein in an E. coli cell-free system will allow the test compounds to come directly in contact with the biosensing protein as it is made, further increasing the sensitivity and efficiency of the assay.

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