Diels-Alder reaction–triggered bioorthogonal protein decaging in living cells

Diels-Alder chemistry is widely used for bioconjugations, and one variant of the reaction can ‘deprotect’ a small molecule via spontaneous elimination. This activation chemistry is now demonstrated on biomolecules in cells at high yields in 10 minutes. Small molecules that specifically activate an intracellular protein of interest are highly desirable. A generally applicable strategy, however, remains elusive. Herein we describe a small molecule–triggered bioorthogonal protein decaging technique that relies on the inverse electron-demand Diels-Alder reaction for eliminating a chemically caged protein side chain within living cells. This method permits the efficient activation of a given protein (for example, an enzyme) in its native cellular context within minutes.