Biosensors utilizing ligand dependent intein-splicing: Application in a Diagnostic Pacifier

This year’s project will produce a novel biochemical assay for the quantification of a given ligand in solution. To achieve this goal, we will be developing an engineered protein construct that consists of three domains, 1) a modular ligand binding domain, 2) an intein splicing domain, 3) and a split NanoLuc® Luciferase domain. While protein constructs such as this can be applied to a variety of substrates and/or ligands, one chosen application for this novel technology is to detect and quantify the amount of cortisol present in saliva. As cortisol is indicative of the human stress response, we would effectively be able to quantify an individual’s stress at a given time. This protein construct may also be used for diagnostic purposes involving the detection of hormone imbalances. A specific application of this technology is the communication of stress in non-verbal individuals, such as infants. Therefore, we are producing a novel protein construct that links the binding of cortisol to the endogenous human glucocorticoid ligand binding domain to intein splicing together two halves of a split NanoLuc® Luciferase. The splicing event would produce a functional luciferase reporter, and the resulting luminescence would allow for a direct quantification of cortisol levels.