期刊
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
卷 1850, 期 9, 页码 1891-1897出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbagen.2015.05.022
关键词
Trypanothione reductase; Sulfenic acid; Electron transfer; O-2 reduction; Glutathione reductase; Promiscuous enzymes
资金
- Sapienza University of Rome Progetto d'Ateneo
- CNR
Background: Leishmania infantum is a protozoan of the trypanosomatid family causing visceral leishmaniasis. Leishmania parasites are transmitted by the bite of phlebotomine sand flies to the human host and are phagocyted by macrophages. The parasites synthesize N1-N8-bis(glutationyl)-spermidine (trypanothione, TS2), which furnishes electrons to the tryparedoxin-typaredoxin peroxidase couple to reduce the reactive oxygen species produced by macrophages. Trypanothione is kept reduced by typanothione reductase (TR), a FAD-containing enzyme essential for parasite survival. Methods: The enzymatic activity has been studied by stopped-flow, absorption spectroscopy, and amperometric measurements. Results: The study reported here demonstrates that the steady-state parameters change as a function of the order of substrates addition to the TR-containing solution. In particular, when the reaction is carried out by adding NADPH to a solution containing the enzyme and trypanothione, the Km for NADPH decreases six times compared to the value obtained by adding TS2 as last reagent to start the reaction (1.9 vs. 12 mu M). More importantly, we demonstrate that TR is able to catalyze the oxidation of NADPH also in the absence of trypanothione. Thus, TR catalyzes the reduction of O-2 to water through the sequential formation of C(4a)-(hydro)peroxyflavin and sulfenic acid intermediates. This NADPH:O-2 oxidoreductase activity is shared by Saccharomyces cerevisiae glutathione reductase (GR). Conclusions: TR and GR, in the absence of their physiological substrates, may catalyze the electron transfer reaction from NADPH to molecular oxygen to yield water. General significance: TR and GR are promiscuous enzymes. (C) 2015 Elsevier B.V. All rights reserved.
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