A novel assay based on pre-equilibrium titration curves for the determination of enzyme inhibitor binding kinetics

Bernard Noppen, Anouk Vanbelle, Alan W Stitt, Marc Vanhove

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Abstract

Selection of pharmacological agents based on potency measurements performed at equilibrium fail to incorporate the kinetic aspects of the drug-target interaction. Here we describe a method for screening or characterization of enzyme inhibitors that allows the concomitant determination of the equilibrium inhibition constant in unison with rates of complex formation and dissociation. The assay is distinct from conventional enzymatic assays and is based on the analysis of inhibition curves recorded prior to full equilibration of the system. The methodology is illustrated using bicyclic peptide inhibitors of the serine protease plasma kallikrein.
Original languageEnglish
JournalEuropean biophysics journal : EBJ
Early online date22 Jun 2021
DOIs
Publication statusEarly online date - 22 Jun 2021

Keywords

  • Enzyme inhibition assay
  • Pre-equilibrium inhibition curves
  • Tight-binding inhibitors

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