Catalytic Mechanism of Cruzain from Trypanosoma cruzi As Determined from Solvent Kinetic Isotope Effects of Steady-State and Pre-Steady-State Kinetics.

ABSTRACT: Cruzain, an important drug target for Chagas disease, is a member of clan CA of the cysteine proteases. Understanding the catalytic mechanism of cruzain is vital to the design of new inhibitors. To this end, we have determined pH-rate profiles for substrates and affinity agents and solvent kinetic isotope effects in pre-steady-state and steady-state modes using three substrates: Cbz-Phe-Arg-AMC, Cbz-Arg-Arg-AMC, and Cbz-Arg-Ala-AMC. The pH-rate profile of k_cat/ K_m for Cbz-Arg-Arg-AMC indicated p K₁ = 6.6 (unprotonated) and p K₂ ∼ 9.6 (protonated) groups were required for catalysis. The temperature dependence of the p K = 6.2-6.6 group exhibited a Δ H_ion value of 8.4 kcal/mol, typical of histidine. The pH-rate profile of inactivation by

SUBMITTER: Zhai X 

PROVIDER: S-EPMC10569748 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature