A study on the thermal stability of bovine Cu, Zn superoxide dismutase : the effects of ionic strength and solution composition

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Williams, Douglas B.
Johnson, Eric R.
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Thesis (M.S.)
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The following thesis examines the effects of ionic strength and solution composition on the thermal stability and. activity of bovine erythrocyte superoxide dismutase (BESOD). An indirect assay procedure was employed. The oxidation of xanthine, catalyzed by xanthine oxidase, served as a generator of superoxide radicals (02) and ferricytochrome c functioned as a scavenger of these radicals. The assays were monitored and recorded spectrophotometrically. Percent superoxide dismutase activity remaining after a 30 minute heat treatment and 241 hours after the heat treatment were determined. As the concentration and computed ionic strength values of the ionic solutions increased, the percent BESOD activity remaining after 30 minutes at 80 C decreased. Recovery of BESOD activity after 24 hours was minimal in all ionic solutions. BESOD denaturation was apparently similar in the chloride and sulfate solutions, but much greater in the phosphate solutions. Mathematical interpretation confirmed the similar relationships between chloride and sulfate solutions, and the much greater effects of phosphate solutions on thermal denaturation of SOD. At concentrations of 10-1 M or less, thermal denaturation of SOD was greatest, implying the appearance of a denaturation threshold. A specific phosphate effect may be apparent and was described by two models: 1- complexation of Cut + ions with phosphate anions, and 2- conformational alteration due to charge repulsion between molecular strands of the SOD molecule. Both of these models could be possible explanations as to the much greater thermal denaturation observed in the phosphate solutions.