In-brain synthesis of receptor-based protease sensors by coupling ligand-directed chemistry and click chemistry

The chemical modification of natural proteins in living systems is highly desirable as cutting-edge research at the chemistry–biology interface. Recent advances in bio-orthogonal protein modification have enabled the production of chemically functionalized proteins in cultured cell systems. However, a limited number of methods are applicable in vivo because of the complexity of the three-dimensional constructs of living systems with diverse, heterogeneous cell populations and flow systems filled with tissue fluids. Here we report a genetic-engineering-free method to modify receptor proteins with various probes in the living mouse brain by combining in-brain ligand-directed chemistry with bio-orthogonal click chemistry, and propose a chemical guideline for the reaction design. The rapid and selective tethering of a set of fluorescent peptides to AMPA-type glutamate receptors allowed the synthesis of receptor-based fluorescent sensors. These probes enabled mapping of the activity of matrix metalloproteinase-9 proximal to AMPA-type glutamate receptors in the living brain to be realized with high spatial resolution. The complexity of living systems makes methods for chemically functionalizing proteins in vivo rare. Here, a genetic-engineering-free method to modify receptor proteins with various probes in the live mouse brain is reported by combining ligand-directed chemistry with click chemistry. This method enables the synthesis of receptor-based fluorescent sensors.