A Simple Growth Test of a Saccharomyces cerevisiae Cu, Zn-Superoxide Dismutase-Deficient Mutant in Hypertonic Medium for Biological Evaluation of Antioxidants

Abstract

Osmotic stress hampers the growth of Saccharomyces cerevisiae, and this effect is mediated by oxidative stress. A simple test for the biological evaluation of antioxidants was developed on the basis of recovery of growth delay of Cu, Zn-superoxide dismutase-lacking yeast (SOD1Δ) under hyperosmosis. The SOD1Δ strain suffered from growth inhibition in a medium containing 1.7 M sorbitol. Protein oxidation in SOD1Δ cells was greatly increased by exposure to a hypertonic medium, indicating the accumulation of reactive oxygen species followed by oxidation of cellular materials. The hyperosmosis-induced growth arrest of SOD1Δ was abolished by the addition of L-ascorbic acid to the medium; the antioxidant effect depended on the concentration, ranging from 1 to 10 mM. Cysteine, N-acetyl-cysteine, and glutathione were also able to restore the growth of SOD1Δ cells. High concentrations of these thiol compounds proved less effective, probably due to adverse effects of an excess of these antioxidants. No growth restoration was seen for typical polyphenol antioxidants, including curcumin, quercetin and catechin.

Suppressive Effects of Natural Compounds on Methionine Auxotrophy of a Cu,Zn-Superoxide Dismutase-Deficient Mutant of <i>Saccharomyces cerevisiae</i>

“…These antioxidants have been demonstrated to have effective antioxidant activities in various in vitro assays, and most compounds are ingredients in commercially available dietary supplements (Bouayed and Bohn, 2010;Bergström et al, 2012;Sadowska-antioxidants (AsA and thiol antioxidants) can suppress Met auxotrophy of the sod1∆ strain by exogenous supplementation. The same pattern of antioxidant action was found when studying the restoration of sod1∆ from oxidative stress-sensitive phenotypes (Lewinska et al, 2004;Kozioł et al, 2005;Tamura et al, 2010;Kwolek-Mirek et al, 2012) and a mutator phenotype (Nagira et al, 2013).…”

“…The sod1∆ strain used here was obtained from the collection of the Saccharomyces Genome Deletion Project, and exhibited Met auxotrophy under atmospheric oxygen levels, as well as sensitivity to oxygen and hypertonic stress, and a mutator phenotype (Tamura et al, 2010;Nagira et al, 2013). In the present study, we modified the method of Zyracka et al (2005a;2005b) and thrombosis (Wierzbicki, 2007;Cattaneo, 1999).…”

“…These detrimental phenotypes of sod1∆ could be alleviated by the addition of vitamin C (ascorbic acid, AsA), suggesting the possibility that the yeast mutant can be used as a biosensor for low molecular weight antioxidants (Zyracka et al, 2005b;Tamura et al, 2010;Nagira et al, 2013).…”

Ascorbic Acid and Thiol Antioxidants Suppress Spontaneous Mutagenesis in a Cu,Zn-superoxide Dismutase-deficient Mutant of Saccharomyces cerevisiae

“…Replacement of the SOD1 gene with the KanMX cassette in the mutant strain was veriˆed by PCR. Defect of SOD1 in the SOD1D strain was previously demonstrated by a zymographic assay for SOD activity in a crude cell extract (18). Spontaneous mutation frequencies were measured in SOD1D and parental strains by the CAN1 forward-mutation assay.…”

“…These obvious SOD1D phenotypes could be ameliorated by supplementing AsA at physiological levels, indicating the potent ability of AsA to scavenge endogenous ROS (10,13,14). Therefore, the yeast SOD1D mutant can be used as a biosensor for low molecular weight antioxidants on the basis of its ability to relieve the oxidative damage-induced phenotype (15)(16)(17)(18).…”