Fluorogenic Bifunctional <i>trans</i>‐Cyclooctenes as Efficient Tools for Investigating Click‐to‐Release Kinetics
Mark A. R. de Geus, Elmer Maurits, Alexi J. C. Sarris, Thomas Hansen, Max S. Kloet, Kiki Kamphorst, Wolter ten Hoeve, Marc S. Robillard, Andrea Pannwitz, Sylvestre Bonnet, Jeroen D. C. Codée, Dmitri V. Filippov, Herman S. Overkleeft, Sander I. van Kasteren
Abstract
The inverse electron demand Diels-Alder pyridazine elimination reaction between tetrazines and allylic substituted trans-cyclooctenes (TCOs) is a key player in bioorthogonal bond cleavage reactions. Determining the rate of elimination of alkylamine substrates has so far proven difficult. Here, we report a fluorogenic tool consisting of a TCO-linked EDANS fluorophore and a DABCYL quencher for accurate determination of both the click and release rate constants for any tetrazine at physiologically relevant concentrations.