An ultrafast phototrigger of the Trp(5CN)-Trp motif in a beta-hairpin peptide

Phototriggers are useful molecular tools to initiate reactions in enzymes by light for the purpose of photoenzymatic design and mechanistic investigations. Here, we incorporated the non-natural amino acid 5-cyanotryptophan (W-5CN) in a polypeptide scaffold and resolved the photochemical reaction of the W-5CN-W motif using femtosecond transient UV/Vis and mid-IR spectroscopy. We identified a marker band of similar to 2037 cm(-1) from the CN stretch of the electron transfer intermediate W-5CN (center dot-) in the transient IR measurement and found UV/Vis spectroscopic evidence for the W center dot+ radical at 580 nm. Through kinetic analysis, we characterized that the charge separation between the excited W-5CN andWoccurs in 253 ps, with a charge-recombination lifetime of 862 ps. Our study highlights the potential use of theW5CN-W pair as an ultrafast phototrigger to initiate reactions in enzymes that are not light-sensitive, making downstream reactions accessible to femtosecond spectroscopic detection.