Efficiency of superoxide anions in the inactivation of selected dehydrogenases
Introduction
The superoxide anion radical demonstrated considerably smaller reactivity with proteins in comparison to the OH radical, and therefore in some papers its reaction with proteins is neglected (Bielski and Cabelli, 1995, Bielski et al., 1985, Schüssler and Puchala, 2004, Gebicki, 2006). The lower reactivity of O2− is connected with its longer half-life time in comparison to the hydroxyl radical. In effect, it can diffuse at great distances and react with amino acid residues, which are the most important for protein functioning. If these residues are situated inside or near the protein’s active site, the superoxide radical can also cause significant inactivation of the enzyme.
Moore et al. (2000) in the work on the radiolysis of protein A estimated that superoxide is as efficient as the hydroxyl radical in this protein inactivation. Similar observations regarding alcohol dehydrogenase were made by Badiello et al. (1974).
The aim of this paper was to estimate the effectiveness of the superoxide radical in the inactivation of selected enzymes in comparison to the effectiveness of other products of water radiolysis. It was also attempted to find a connection between the structure of the examined enzymes and their sensitivity to free-radical inactivation.
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Materials
Alcohol dehydrogenase (ADH) (alcohol:NAD+ oxidoreductase, EC 1.1.1.1), molecular weight of 147 kDa from baker’s yeast, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (d-glyceraldehyde-3-phosphate: NAD+ oxidoreductase, EC 1.2.1.12), molecular weight of 143 kDa from a rabbit muscle, lactate dehydrogenase (LDH) (l-lactate:NAD+ oxidoreductase, EC 1.1.1.27), molecular weight of 140 kDa from a rabbit muscle, xanthine oxidase (XO) (xanthine:oxygen oxidoreductase, EC 1.17.3.2) from bovine milk, xanthine
Efficiency of radiation-generated radicals in inactivation of dehydrogenases
In this paper, the radiation inactivation of three enzymes: GAPDH, ADH and LDH was determined. The solutions of enzymes were irradiated under air with doses of up to 120 Gy in four systems:
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in the absence of other compounds;
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in the presence of mannitol;
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in the presence of mannitol and SOD and
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in the presence of mannitol, SOD and catalase.
The dependence of log (% activity) on the radiation dose for the studied systems is presented in Fig. 1, Fig. 2, Fig. 3. In all the cases, these dependences are
Discussion
The superoxide anion radical is the most ubiquitous reactive form of oxygen in all aerobic organisms. Its main sources are: mitochondria, neutrophils, hemoglobin autooxidation processes and reactions of numerous enzymes including xanthine oxidase. It is believed that superoxide anion radical shows low reactivity and in the oxidative stress it is regarded mainly as an initiator of more reactive species such as OH and ONOO− (Gebicki, 2006). Several papers devoted to the direct influence of O2− on
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