Interaction of antimony tartrate with the tripeptide glutathione

Abstract

The tripeptide glutathione (g-l-Glu-l-Cys-Gly, GSH) is thought to play an important role in the biological processing of antimony drugs. We have studied the complexation of the antileishmanial drug potassium antimony(III) tartrate to GSH in both aqueous solution and intact red blood cells by NMR spectroscopy and electrospray ionization mass spectrometry. The deprotonated thiol group of the cysteine residue is shown to be the only binding site for Sb (III), and a complex with the stoichiometry [Sb(GS) 3 ] is formed. The stability constant for [Sb(GS) 3 ] was determined to be log K 25 (I 0.1 m, 298 K) based on a competition reaction between tartrate and GSH at different pH* values. In spite of being highly thermodynamically stable, the complex is kinetically labile. The rate of exchange of GSH between its free and Sb-bound form is pH-dependent, ranging from slow exchange on the 1 H-NMR timescale at low pH (2 s 21 at pH 3.2) to relatively rapid exchange at biological pH (. 440 s 21 ). Such facile exchange may be important in the transport of Sb(III) in various biofluids and tissues in vivo. Our spin±echo 1 H-NMR data show that Sb(III) rapidly entered red blood cell walls and was complexed by intracellular glutathione.Keywords: antimony; glutathione; nuclear magnetic resonance; red blood cells.Leishmaniasis is a parasitic infection caused by various species of the protozoan Leishmania. It remains one of the largest killers in the tropical regions of America, Africa and Asia, with approximately 1.5 billion people at risk and 20 million cases being reported annually. Among the drugs used to treat this condition, several antimony-containing complexes, such as N-methylglucamine antimonate(V) (Glucantimew), sodium stibogluconate (Pentostamw), sodium antimony(III) gluconate (Triostamw) and potassium antimony(III) tartrate (Tartar emeticw), are currently the first choice [1±3], despite their toxicity [4]. Antimony complexes with mannan appear to be promising for further improving the stability and solubility of these drugs [5]. Despite extensive clinical use for several decades, the molecular basis for the selective antileishmanial action of these drugs remains unclear. Furthermore, clinical resistance to these drugs has been increasingly reported in recent years [6±8]. It is generally believed that relatively nontoxic Sb(V) complexes may be prodrugs that could be reduced to the more toxic and active trivalent Sb(III) at or near the site of action (e.g. amastigotes) [9,10].Glutathione (g-l-Glu-l-Cys-Gly, GSH) is a potentially polydentate ligand, and is widely used as a model system for the binding of metal ions by peptides and proteins. It is present in many cells at millimolar concentration and generally is the most abundant nonprotein thiol. In cells, one of its roles is as a substrate for glutathione peroxidase, an enzyme capable of both removing hydrogen peroxide from cells and repairing peroxidatively damaged membranes. It is also involved in metal detoxification, and an ATP-dependent glutathione S-conjugate pump has ...

Mechanisms underlying the toxic effects of antimony species in human embryonic kidney cells (HEK-293) and their comparison with arsenic species

“…Although a significant increase in the DCF fluorescence by the treatment of Sb was not observed, the data show a dose-dependent correlation. The increasing trend in the generation of ROS in the presence of a high concentration of Sb or As exposure can be explained by studies of several cell lines, which have shown the formation of oxidative stress at high concentrations of As(III) (Alarifi et al, 2013;Selvaraj et al, 2013) and the depletion of glutathione in complex with Sb(III) or As(III) (Burford et al, 2005) (Sun et al, 2000;Kobayashi et al, 2005), resulting in the disturbance of the intracellular redox balance.…”

“…Protein aggregates in yeast, even those of chaperone proteins, were suggested to the interact using the sulfhydryl groups of Cys (Jacobson et al, 2012). In the case of Sb, no study clearly addresses that its protein aggregates in living cells exist, although some studies for the interaction of Sb with biomolecules, such as glutathione, adenine and guanine nucleosides, obtained by 13 C and 1 H NMR, ESI-MS and circular dichroism (Burford et al, 2005;Demicheli et al, 2002;Sun et al, 2000) have been reported. Although the hitherto study mentioned that the toxicological mechanisms of Sb may be quite similar to those of As because of the physicochemical similarities of Sb to As (Beyersmann and Hartwig, 2008), the results obtained in this study show some toxicological differences between Sb and As.…”

“…Regarding the interactions of inorganic Sb with biomolecules, other groups have determined the existence of Sb-S complexes with zinc finger peptides (Demicheli et al, 2008;Frézard et al, 2012), glutathione (Burford et al, 2005;Sun et al, 2000), guanine and adenosine nucleosides (Demicheli et al, 2002), but the direct interaction of Sb with proteins has not been described.…”

Dual Action of Antimonial Drugs on Thiol Redox Metabolism in the Human Pathogen Leishmania donovani

“…The precise mechanism of efflux remains uncertain, but previous studies on plasma membrane vesicles have shown that Leishmania cell lines actively extrude 73 As (as arsenite) in the presence of ATP and either glutathione or trypanothione at equal rates (19). Based on the fact that Sb III inhibits this activity (18), coupled with the fact that Sb III forms a complex with either glutathione (38) or trypanothione (39), it has been postulated that Sb III is effluxed in a similar fashion. Although this is an attractive hypothesis, direct demonstration of cotransport of antimony and thiol is lacking.…”

Drug Resistance in Leishmaniasis

“…Spontaneous formation of Sb(III) complexed with either glutathione, trypanothione or both has been demonstrated by proton nuclear magnetic resonance spectroscopy (140,162) and by mass spectrometry (107). Since glutathione S-transferase (GST) is elevated in mammalian cells selected for resistance to arsenite (97), it has been proposed that formation of the metalloid-thiol pump substrates in Leishmania spp.…”