Bimodal detection of proteins by $^{129}$Xe NMR and fluorescence spectroscopy

The full understanding of biological phenomena involves sensitive and non-invasive detection. Here we report the optimization of a probe for intracellular proteins that combines the advantages of fluorescence and hyperpolarized $^{129}$Xe NMR detection. The fluorescence detection part is composed of six residues containing a tetracysteine tag (-CCXXCC-) genetically incorporated into the protein of interest and of a small organic molecule, CrAsH. CrAsH becomes fluorescent when it binds to the tetracysteine tag. The part of the biosensor that enables $^{129}$Xe NMR detection, linked to the CrAsH moiety by a spacer, is based on a cryptophane core fully suited to reversibly host xenon. We benchmark three different peptides containing the tetracysteine tag and four organic biosensors of different stereochemistry to propose the best couple, fully suited for the $in\ vitro$ detection of proteins.