Mitochondrial NADH-cytochrome b5 reductase plays a crucial role in the reduction of D-erythroascorbyl free radical in Saccharomyces cerevisiae

The relevance of NADH-cytochrome b(5) reductase to the NADH-dependent reduction of D-erythroascorbyl free radical was investigated in Saccharomyces cerevisiae. MCRI. which is known to encode NADH-cytochrome b(5) reductase in S. cerevisiae, was disrupted by the insertion of URA3 gene into the gene of MCRI. In the mcr1 disruptant cells, the activity of NADH-D-erythroascorbyl free radical reductase almost disappeared and the intracellular level of D-erythroascorbic acid was about 11% of that of the congenic wild-type strain. In the transformant cells carrying MCRI in multicopy plasmid, the intracellular level of D-erythroascorbic acid and the activity of NADH-D-erythroascorbyl free radical reductase increased up to 1.7-fold and 2.1-fold, respectively. Therefore, it indicated that the,MCRI product. mitochondrial NADH-cytochrome b(5) reductase, plays a key role in the NADH-dependent reduction of D-erythroascorbyl free radical in S. cerevisiae. On the other hand. the mcr1 disruptant cells were hypersensitive to hydrogen peroxide and menadione, and overexpression of MCRI made the cells more resistant against oxidative stress. These results suggested that the mitochondrial NADH-cytochrome b(5) reductase functions as NADH-D-erythroascorbyl free radical reductase and plays an important role in the response to oxidative damage in S. cerevisiae. (C) 2001 Elsevier Science B.V. All rights reserved.