We have harnessed photo-induced electron transfer (PET) as a mechanism for a new photochemically-driven protein ligation reaction. We demonstrated that this could be achieved by pairing the inherently photo- and redox-lability of Tryptophan (Trp) residues by with an N-carbamoyl pyridinium salt that simultaneously couples photo-induced electron transfer (PET) with Trp to a radical fragmentation/recombination process to carbamylate Trp residues site-selectively in high conversion and yields. We also demonstrated that the photophysical and electrochemical properties of the pyridinium scaffold can be readily tuned such that either [Trp]*-pyridinium PET (Trp is a photoreductant) or Trp->[pyridinium]* PET (Trp is a reductive quencher) can be invoked to drive the labelling chemistry.
We have further refined the pyridinium scaffold to include a donor-acceptor relationship between the two aryl groups. This relationship enables photoconjugation at Trp using visible light as a trigger, and at concentrations of protein and probe that are an order of magnitude more dilute than those used with our first-generation probes without sacrificing reaction times (10-60 min). These mild conditions enabled us to conduct global chemoproteomic profiling of Trp residues in live cell culture. Beyond demonstrating biocompatibility, this experiment enabled us directly label and identify >200 hyper-reactive Trp residues in their native environment. These types of data sets can then be used as a basis for discovering “hotspots” that are of biochemical significance, as well to find new targets and sites for drug discovery.
