Enhanced Raman Imaging of Ligand-Receptor Recognition Abstract: The goal of this proposal is to identify molecular interactions relevant to drug targeting and chemical signaling in living cells. A critical bottleneck in the development of drugs is the identification of off-target effects. Methods that can identify the molecular interactions associated with proteins recognizing and binding to drug candidates in cellular and other live models can be used to understand and minimize unwanted side effects and complications. Identifying these effects at earlier stages of drug screening is important to avoid late stage drug failure. We are developing technologies that take advantage of the plasmonic properties of metallic nanoparticles to enable chemical-specific spectroscopic studies of ligand-receptor binding in living cells. These investigations will provide new approaches to probing the receptor’s chemical residues that bind peptide antagonists and provide insights into molecular interactions that regulate the proteins involved in signaling and drug targeting. Our approach combines enhanced Raman scattering from both nanoparticles (Surface enhanced Raman scattering, or SERS) and scanning probes (tip enhanced Raman scattering, or TERS), nanoparticle tracking microscopy, non-standard applications of static and dynamic quantum chemical calculations, and super- resolution SERS imaging to characterize chemical interactions that regulate binding to protein receptors. Information present in the enhanced Raman scattered response provides molecular level detail of the interactions governing recognition by the protein. In concert, our methodologies provide a new approach to monitoring protein binding to putative drugs in living cells, and to characterize targeting specificity. The specific aims of this proposal are: 1) Screen the targeting specificity of peptide-functionalized nanoparticles in live cells. 2) Develop super-resolution SERS imaging to improve binding specificity studies and identify particle location in cells. 3) Combine non-standard quantum and numerical simulations with experiments to identify key motifs in amino acid conformation related to peptide binding. Overall, the technology and platform we propose will address the challenge of obtaining chemical information from ligands binding to receptor proteins in intact cells. These studies will provide new insights into the molecular interactions that regulate signaling pathways and how anomalies in these interactions are associated with disease and treatment.