Fluorogenic In Situ Thioacetalization: Expanding the Chemical Space of Fluorescent Probes, Including Unorthodox, Bifurcated, and Mechanosensitive Chalcogen Bonds

Progress with fluorescentflippers, small-molecule probes to imagemembrane tension in living systems, has been limited by the effortneeded to synthesize the twisted push-pull mechanophore. Here,we move to a higher oxidation level to introduce a new design paradigmthat allows the screening of flipper probes rapidly, at best in situ. Late-stage clicking of thioacetals and acetalsallows simultaneous attachment of targeting units and interfacersand exploration of the critical chalcogen-bonding donor at the sametime. Initial studies focus on plasma membrane targeting and developthe chemical space of acetals and thioacetals, from acyclic aminoacids to cyclic 1,3-heterocycles covering dioxanes as well as dithiolanes,dithianes, and dithiepanes, derived also from classics in biologylike cysteine, lipoic acid, asparagusic acid, DTT, and epidithio-diketopiperazines.From the functional point of view, the sensitivity of membrane tensionimaging in living cells could be doubled, with lifetime differencesin FLIM images increasing from 0.55 to 1.11 ns. From a theoreticalpoint of view, the complexity of mechanically coupled chalcogen bondingis explored, revealing, among others, intriguing bifurcated chalcogenbonds.

Automated summary

We have developed a new design paradigm for fluorescent flipper probes, which allows for rapid screening and targeting of membrane tension in living cells.