Fig. 1.

Concept and design of LUPIN. a The emission spectrum of NLuc (blue) overlaps very well with the absorption spectrum of the ruthenium photocatalyst (red), suggesting that efficient BRET should be possible. b The fusion protein SNAP-Pro30-NanoLuc (NLuc)-cpDHFR is linked via self-labeling SNAP to the synthetic linker containing the ruthenium photocatalyst and methotrexate (purple ball, DHFR ligand), positioning the ruthenium in close proximity to the NLuc. Free methotrexate (green ball) can push the sensor into the open conformation, thus turning off the BRET to the photocatalyst. c By installing a PNA next to the ruthenium catalyst, complementary pyridinium substrates can bind and undergo photoreductive cleavage by the ruthenium, unmasking the effector molecule