Selective On/Off-Nitroxides as Radical Probes to Investigate Non-radical Enzymatic Activity by Electron Paramagnetic Resonance

Indranil Duttagupta (1) , Natacha Jugniot (2) , Gérard Audran (1) , Jean-Michel Franconi (2) , Sylvain R.A. Marque (1, 3) , Philippe Massot (2) , Philippe Mellet (2) , Elodie Parzy (2) , Eric Thiaudiere (2) , Nicolas Vanthuyne (4)

1 ICR - Institut de Chimie Radicalaire
2 CRMSB - Centre de résonance magnétique des systèmes biologiques
3 RAS, SB, NN - Vorozhtsov Novosibirsk Institute of Organic Chemistry
4 ISM2 - Institut des Sciences Moléculaires de Marseille

A nitroxide carrying a peptide specific to the binding pocket of the serine proteases chymotrypsin and cathepsin G is prepared. This peptide is attached as an enol ester to the nitroxide. Upon enzymatic hydrolysis of the peptide, the enol ester moiety is transformed into a ketone moiety. This transformation affords a difference of 5G in phosphorus hyperfine coupling constant between the electronic paramagnetic resonance (EPR) signals of each nitroxide. This property is used to monitor the enzymatic activity of chymotrypsin and cathepsin G by EPR. Michaelis constants were determined and match those reported for conventional optical probes.

Mots clés: Nitroxides; Selectivity; EPR spectroscopy; Enzymatic activity; Michaelis constants

Domaines: Chimie; Chimie organique; Biochimie [q-bio.BM]

Indranil Duttagupta, Natacha Jugniot, Gérard Audran, Jean-Michel Franconi, Sylvain R.A. Marque, et al.. Selective On/Off-Nitroxides as Radical Probes to Investigate Non-radical Enzymatic Activity by Electron Paramagnetic Resonance. Chemistry - A European Journal, 2018, 24 (30), pp.7615-7619. ⟨10.1002/chem.201800866⟩. ⟨hal-02091903⟩