Protein structure analysis by robust electron spin resonance at the single molecule level.

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The project aims to demonstrate analysis of structure and dynamics of a protein by single-molecule electron spin spectroscopy. This ambitious goal has come within reach by recent joint work of the applicants: ESR detection of a single protein, labeled by a stable organic electron spin, using a nearby Nitrogen-Vacancy (NV) color center in diamond as a sensor [F. Shi, et al., Science 347, 1135 (2015)]. The project will overcome two major problems encountered in this proof-of-principle experiment: low stability of the spin labels involved and inefficient detection. This shall be accomplished by two approaches: resonant single-photon and two-photon excitation of the NV sensor at cryogenic temperature; use of more resilient labels and nano-photonic structures for readout enhancement at room temperature.The project will equally develop protocols for preparation of single site-specifically labeled proteins on the sensor surface and structure determination by multidimensional spectroscopy. It will finally combine all these developments to enable stable and efficient spectroscopy of single molecules by electron spin resonance, in particular three-dimensional structure analysis of multiply labeled proteins.