Fluorophore Labeling of Proteins: a Versatile Trigger-Release-Conjugation Platform Based on the Quinone Methide Chemistry.

conjugation of fluorescent molecules to biomolecules such as proteins under spatiotemporal control offers a powerful means for studying biological systems. For that purpose, the -quinone methide chemistry involving a sequence of the trigger-release-conjugation (TRC) process provides a versatile conjugation method. We have developed a new TRC platform bearing a quaternary ammonium salt for the release process, which can be structurally modified and readily synthesized from commonly used aryl alcohol-type organic fluorophores under environmentally benign conditions. We show that different aryl alcohol fluorophores containing the (morpholinium)methyl group for the release process allow efficient fluorophore labeling of proteins under both light- and chemical-triggering conditions. The bioconjugation in cells as well as in tissues was further demonstrated with an (morpholinium)methyl analogue containing a triggering group sensitive to reactive oxygen species. The new TRC system thus provides a versatile and unique platform for fluorophore labeling of proteins in biological systems under spatiotemporal control.