Elsevier

Nitric Oxide

Volume 94, 1 January 2020, Pages 69-72
Nitric Oxide

Influence of hemoglobin and albumin on the NO donation effect of tetranitrosyl iron complex with thiosulfate

https://doi.org/10.1016/j.niox.2019.10.010Get rights and content

Highlights

  • Hemoglobin and albumin stabilize nitrosyl iron complex with thiosulphate ligands.

  • Thiosulfate DNIC interacts with positively charged amino acid residues of Hb.

  • DNIC with thiosulphate ligands decays 27 times slower in the presence of BSA.

Abstract

The effects of deoxyhemoglobin (Hb) and albumin on the NO-donor activity of the anionic tetranitrosyl iron complex with thiosulfate ligands (1) were studied for the first time. It was shown that Hb significantly stabilizes complex 1; in its presence, NO generation from the complex proceeds at a noticeably slower rate. A similar effect is observed when complex 1 is bound to albumin, in which case complex 1 decomposes 27 times slower than in the absence of albumin in the solution. The observed effects provide a prolonged action of complex 1 as NO-donor, which may enhance its potential pharmacological efficacy.

Introduction

Structural synthetic analogs of the active center of non-heme iron-sulfur proteins, nitrosyl iron complexes (DNICs) with functional sulfur-containing ligands, are of great interest as new generation drugs for the treatment of cardiovascular, oncological and other socially significant diseases [1,2].

Anionic binuclear DNIC with thiosulfate ligands (complex 1) Na2 [Fe2(S2O3)2(NO)4]4H2O (Fig. 1) is one of the most promising representatives of this compounds class [3]. It has been shown that complex 1 is a more effective NO donor compared with the NONOate, 2,2′-(hydroxynitrosohydrazono)bis-ethanimine [4], it exhibits antimetastatic activity and induces the expression of the genes of the protective DNA repair pathways in the cell, including the SOS and SoxRS systems of E. Coli [5], and also completely inhibits the catalytic activity of Ca2+-ATPase and phosphodiesterase at a concentration of 0.01 mM [6].

To understand the chemical and biological mechanisms of complex 1 action as a prodrug, a comprehensive study of its metabolic processes is necessary to conduct. It is assumed that the targets of DNICs as NO donors in vivo can be Fe- and thiol-containing proteins, as well as low molecular weight thiols [7,8]. In this work, the biotransformation of complex 1 was analyzed by the effects of its interaction with deoxyhemoglobin (Hb) and bovine serum albumin (BSA).

According to the literature [9], erythrocyte hemoglobin is one of the most important elements of the biological transformation of NO donors and the NO transport inside the body. It was found that besides the formation of protein-bound complexes with the oxygenated form of the protein (HbO2) [10], DNICs are also able to bind with deoxygenated Hb [11], becoming more prolonged NO donors. As a result of the analysis carried out in Ref. [11], it was concluded that the observed effect of cationic complexes stabilization can occur due to the binding of their positively charged groups with negatively charged amino acid residues on the Hb surface. It was interesting to determine whether a similar effect would be observed in the case of anionic complex 1.

Another no less important objective of this work was to study the effect of BSA on the decay process of complex 1. BSA is a blood transport protein the main function of which is a transfer of various biologically active substances [12]. It has been established [7] that BSA is one of the main targets of the DNICs action, and the binding of DNICs occurs with cysteine (Cys 34) or histidine residues in the hydrophobic pocket of the molecule to form protein-bound nitrosyl complexes [13].

According to experimental data obtained in animals [14], high-molecular complexes formed by the interaction of DNICs with various proteins are more stable than low-molecular complexes, and as a result, most of the DNICs in cells and tissues have a protein form.

Section snippets

Materials

In the work, tris(hydroxymethyl)aminomethane (“Serva”, Germany), sulfanilamide (SA) (“Sigma”, USA), N-(1-Naphthyl)ethylenediamine dihydrochloride (NEDA) (“MP Biomedicals”, Germany) and bovine serum albumin, fraction 5 (BioClot, GmbH) were used. Water was distilled in a “Bi/Duplex” distiller (Germany). Binuclear tetranitrosyl iron complex 1 has been obtained using techniques [3]. A homogeneous solution of HbO2 was isolated from fresh blood of experimental animals by the known method [15]. To

Results

Hb is very convenient for the spectrophotometric recording of the NO generation kinetics from complex 1 because it is a trap for NO: the rate of NO binding to the Hb heme is close to diffusion, the equilibrium constant is 3∙1010 M−1 [17]. In addition, formed nitrosyl hemoglobin (HbNO) has a characteristic absorption spectrum: one Hb absorption maxima (556 nm) splits into two (545 nm and 575 nm) during HbNO formation, which is convenient to use to determine the depth of the reaction. Thus, the

Discussion

The results of this work demonstrate a high stabilizing ability of proteins towards the complex 1. As was shown earlier, the albumin-bound complex is formed by the rapid reaction of free cysteine and histidine groups with DNIC [13]. Hemoglobin, from this point of view, is a unique protein that can bind DNICs and their decay products in various ways [10,23]. In arterial blood, where HbO2 predominates, the binding of NO and DNICs occurs with the free SH group (Cys 93). At the same time, when the

Acknowledgements

The work was supported by the Ministry of Education and Science of the Russian Federation, state task No 0089-2019-0014 and the Presidium Program of the Russian Academy of Sciences for 2018–2020. “Nanostructures: physics, chemistry, biology, fundamentals of technology”, the project “Development of ways to optimize the pharmacological properties of biologically active compounds through inclusion in molecular nanostructures”.

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