Design of spiro[2.3]hex-1-ene, a genetically encodable double-strained alkene for superfast photoclick chemistry

Reactive yet stable alkene reporters offer a facile route to studying fast biological processes via the cycloaddition-based bioorthogonal reactions. Here, we report the design and synthesis of a strained spirocyclic alkene, spiro[2.3]hex-1-ene (Sph), for an accelerated photoclick chemistry, and its site-specific introduction into proteins via amber codon suppression using the wild-type pyrrolysyl-tRNA synthetase/tRNA_CUA pair. Because of its high ring strain and reduced steric hindrance, Sph exhibited fast reaction kinetics (k₂ up to 34 000 M^-1 s^-1) in the photoclick chemistry and afforded rapid (<10 s) bioorthogonal protein labeling.