Site-directed spin labeling EPR spectroscopy in vivo

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Site-directed spin labeling (SDSL) EPR spectroscopy has emerged as a powerful technique to study the structure and conformational dynamics of biomolecules in vitro; however, the application of this technique in live cells still remains challenging. The goal of this proposal is the development and application of SDSL EPR spectroscopy in vivo, which allows addressing problems of cell biology not accessible by in vitro experiments. Orthogonal spin labeling techniques including enzymatic labeling and click chemistry approaches, which have been developed in the preceding funding period, will be applied to study the interaction and conformational changes of the bacterial toxin colicin A and the corresponding immunity protein Cai in live Escherichia coli cells.