Abstract 2126 Construction of a functional bacterial-based estrogen biosensor based utilizing dimerization-dependent Red Fluorescent Protein

The presence of estrogenic compounds (endocrine-disruptors, EDCs) in the water supply raises concerns about human and aquatic health. In recent years, there has been increasing amounts of these steroid hormones leaching into water and soil particularly from livestock farms and sewage plants. Current methods for detection of these EDCs remains expensive and time-consuming, requiring techniques such as liquid chromatography-mass spectrometry and high-performance liquid chromatography. Previously reported estrogen biosensors required multiple cloning and transformation steps for successful detection in bacteria. Advances in Synthetic biology allow for the modulization of gene parts and construction of genetic devises composed fragments modified to be interchangeable and provide novel functions. Building on recent approaches using rational engineering to express dimerization dependent Red Fluorescent Proteins (ddRFP) to detect protein-protein interactions, we present a ddRFP-based Estrogen biosensor. We have engineered plasmids to express two monomeric weakly fluorescent RFP proteins derived from mutant dTomato (Campbell et al, 2021) fused to the Estrogen Receptor alpha protein (ERalpha) in bacteria cells. These are strongly fluorescent upon dimerization. These RFP constructs are brought together in the cell when ERalpha dimerizes in the presence of estrogen. We present here a construction strategy and data demonstrating the efficiency of this dual-functional biosensor to detect estradiols in an aqueous environment. Putative quantification methods will be demonstrated. NSF Research Experiences for Undergraduates (REU): REU in Synthetic Biology.