An N-Allylglyoxylamide Based Bioorthogonal Nitrone Trap

Aldehydes are attractive bioorthogonal coupling partners. The ease of manipulation of aldehydes and their orthogonality to other classes of bioorthogonal reactions has inspired the exploration of chemistries which generate irreversible conjugates. Similarly, nitrones have been shown to be potent 1,3 dipoles in bioorthogonal reactions when paired with strained alkynes. Here we combine the reactivity of nitrones with the simplicity of aldehydes using an N-allyl glyoxylamide, in a cascade reaction with an N-alkylhydroxylamine to produce a bicyclic isoxazolidine. The reaction is found to be catalyzed by 5-methoxyanthranilic acid and proceeds at pH 7 with favorable kinetics. Using the HaloTag7 protein bearing a hydroxylamine we show the reaction to be bioorthogonal in a complex cell lysate and to proceed well at the surface of a HEK293 cell. Furthermore, the reaction is orthogonal to a typical strain-promoted alkyne-azide click reaction. The characteristics of this reaction suggest it will be a useful addition to the pallet of bioorthogonal reactions revolutionizing chemical biology.