Trypanosoma Cruzi trypanothione reductase is inactivated by peroxidase-generated phenothiazine cationic radicals

• Published in: Free radical research 2001-01, Vol.34 (4), p.363-378
• Main Authors: Gutierrez-Correa, Jose, Fairlamb, Alan H., Stoppani, Andres O.M.
• Format: Article
• Language: English
• Subjects: Animals; cationic radicals; Cations, Monovalent; Electron Spin Resonance Spectroscopy; Enzyme Inhibitors - pharmacology; Free Radicals; horseradish peroxidase; Horseradish Peroxidase - metabolism; Horseradish Peroxidase - pharmacology; Hydrogen Peroxide - metabolism; Hydrogen Peroxide - pharmacology; Kinetics; myeloperoxidase; myoglobin; Myoglobin - metabolism; Myoglobin - pharmacology; NADH, NADPH Oxidoreductases - drug effects; Peroxidase - metabolism; Peroxidase - pharmacology; phenothiazine; Phenothiazines - metabolism; Phenothiazines - pharmacology; Reactive Oxygen Species - metabolism; Recombinant Proteins - antagonists & inhibitors; Structure-Activity Relationship; Sulfhydryl Compounds - pharmacology; Time Factors; Trypanosoma cruzi; Trypanothione reductase
• ISSN: 1071-5762; 1029-2470
• DOI: 10.1080/10715760100300311

Trypanosoma cruzi trypanothione reductase (TR) was irreversibly inhibited by peroxidase/H2O2/phenothiazine (PTZ) systems. TR inactivation depended on (a) time of incubation with the phenothiazine system; (b) the peroxidase nature and (c) the PTZ structure and concentration. With the most effective systems, TR inactivation kinetics were biphasic, with a relatively fast initial phase during which about 75% of the enzyme activity was lost, followed by a slower phase leading to total enzyme inactivation. GSH prevented TR inactivation by the peroxidase/H2O2/PTZ+· systems. Production of PTZ+· cation radicals by PTZ peroxidation was essential for TR inactivation. Horseradish peroxidase, leukocyte myeloperoxidase (MPO) and the pseudo-peroxidase myoglobin (Mb) were effective catalysts of PTZ+· production. Promazine, thioridazine, chlorpromazine, propionylpromazine prochlorperazine, perphenazine and trimeprazine were effective constituents of the HRP/H2O2/PTZ system. The presence of substituents at the PTZ nucleus position 2 exerted significant influence on PTZ activity, as shown by the different effects of 2-trifluoromethyl and 2-H or 2-chlorophenothiazines. The PTZ+· cation radicals disproportionation regenerated the non-radical PTZ molecule and produced the PTZ sulfoxide that was inactive on TR. Thiol compounds including GSH interacted with PTZ+· cation radicals transferring an electron from the sulfide anion to the PTZ+·, thus nullifying the PTZ+· biological and chemical activities.