Reduction of dehydroascorbate to ascorbate by the selenoenzyme thioredoxin reductase.
Recycling of ascorbate from its oxidized forms is essential to maintain stores of the vitamin in human cells. Whereas reduction of dehydroascorbate to ascorbate is thought to be largely GSH-dependent, we reconsidered the possibility that the selenium-dependent thioredoxin system might contribute to ascorbate regeneration. We found that purified rat liver thioredoxin reductase functions as an NADPH-dependent dehydroascorbate reductase, with an apparent Km of 2. 5 mM for dehydroascorbate, and a kcat of 90 min-1. Addition of 2.8 microM purified rat liver thioredoxin lowered the apparent Km to 0.7 mM, without affecting the turnover (kcat of 71 min-1). Since thioredoxin reductase requires selenium, we tested the physiologic importance of this enzyme for dehydroascorbate reduction in livers from control and selenium-deficient rats. Selenium deficiency lowered liver thioredoxin reductase activity by 88%, glutathione peroxidase activity by 99%, and ascorbate content by 33%, but did not affect GSH content. NADPH-dependent dehydroascorbate reductase activity due to thioredoxin reductase, on the basis of inhibition by aurothioglucose, was decreased 88% in dialyzed liver cytosolic fractions from selenium-deficient rats. GSH-dependent dehydroascorbate reductase activity in liver cytosol was variable, but typically 2-3-fold that of NADPH-dependent activity. These results show that the thioredoxin system can reduce dehydroascorbate, and that this function is required for maintenance of liver ascorbate content.
May
J M
JM
Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-6303, USA. james.may@mcmail.vanderbilt.edu
Mendiratta
S
S
Hill
K E
KE
Burk
R F
RF
D000818:Animals; D001205:Ascorbic Acid / Q000096:biosynthesis*; D003683:Dehydroascorbic Acid / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D051381:Rats; D012643:Selenium / Q000172:deficiency; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism*; D013879:Thioredoxins / Q000378:metabolism
D013879:Thioredoxins; D009249:NADP; D013880:Thioredoxin-Disulfide Reductase; D012643:Selenium; D001205:Ascorbic Acid; D003683:Dehydroascorbic Acid