Enzyme-dependent ascorbate recycling in human erythrocytes: role of thioredoxin reductase.

Mendiratta S, Qu ZC, May JM
Free Radic Biol Med. 1998 25 (2): 221-8

PMID: 9667500 · DOI:10.1016/s0891-5849(98)00060-4

Human erythrocytes efficiently reduce dehydroascorbic acid (DHA) to ascorbate, which helps to maintain the ascorbate content of blood. Whereas erythrocyte DHA reduction is thought to occur primarily through a direct chemical reaction with GSH, this work addresses the role of enzyme-mediated DHA reduction by these cells. The ability of intact erythrocytes to recycle DHA to ascorbate, estimated as DHA-dependent ferricyanide reduction, was decreased in parallel with GSH depletion by glutathione-S-transferase substrates. In contrast, the sulfhydryl reagent phenylarsine oxide inhibited DHA reduction to a much greater extent than it decreased GSH in intact cells. DHA reduction in excess of that due to a direct chemical reaction with GSH was also observed in freshly prepared hemolysates. Hemolysates likewise showed NADPH-dependent reduction of DHA that appeared due to thioredoxin reductase, because this activity was inhibited 68% by 10 microM aurothioglucose, doubled by 5 microM E. coli thioredoxin, and had an apparent Km for DHA (1.5 mM) similar to that of purified thioredoxin reductase. Additionally, aurothioglucose-sensitive, NADPH-dependent DHA reductase activity was decreased 80% in hemolysates prepared from phenylarsine oxide-treated cells. GSH-dependent DHA reduction in hemolysates was more than 10-fold that of NADPH-dependent reduction. Nonetheless, the ability of phenylarsine oxide to decrease DHA reduction in intact cells with little effect on GSH suggests that enzymes, such as thioredoxin reductase, may contribute more to this activity than previously considered.

MeSH Terms (19)

Arsenicals Ascorbic Acid Aurothioglucose Cell-Free System Cell Membrane Dehydroascorbic Acid Dinitrochlorobenzene Dose-Response Relationship, Drug Enzymes Erythrocytes Ferricyanides Glutathione Glutathione Transferase Hemolysis Humans Ketones NADP Thioredoxin-Disulfide Reductase Vitamin E

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