Mammalian thioltransferase (glutaredoxin) and protein disulfide isomerase have dehydroascorbate reductase activity.
Homogeneous native and recombinant porcine liver thioltransferase (glutaredoxin), bovine thymus and human placenta thioltransferase (glutaredoxin) were examined for dehydroascorbate reductase activity (EC 1.8.5.1) involving the direct catalytic reduction of dehydroascorbic acid (DHA) by glutathione. Each enzyme had substantial activity with apparent Km and Vmax for dehydroascorbate between 0.2 and 2.2 mM and 6-27 nmol min-1, respectively, and for gluathione between 1.6 and 8.7 mM and 11-30 nmol min-1, respectively. In the presence of purified bovine liver thioredoxin reductase, homogeneous bovine liver thioredoxin failed to reduce DHA to ascorbic acid as measured by NADPH oxidation. Highly purified bovine liver protein disulfide isomerase (PDI) reacted directly with DHA and GSH to catalyze the reduction of DHA to ascorbic acid. The apparent Km for DHA was 1.0 mM and the Vmax was 8 nmol min-1, and for GSH were 3.9 mM and 14 nmol min-1, respectively. These results suggest that thioltransferase and PDI contribute to the regeneration of oxidized ascorbic acid in mammalian cells, and based on their cellular location, thioltransferase is proposed to be the major cytoplasmic activity, whereas interaction of DHA with microsomal membrane PDI may catalyze regeneration of ascorbic acid and initiate oxidation of intralumenal protein thiols to disulfides.
Wells
W W
WW
Department of Biochemistry, Michigan State University, East Lansing 48824.
Xu
D P
DP
Yang
Y F
YF
Rocque
P A
PA
D000818:Animals; D002417:Cattle; D005260:Female; D054477:Glutaredoxins; D006801:Humans; D007535:Isomerases / Q000378:metabolism*; D008099:Liver / Q000201:enzymology; D008970:Molecular Weight; D010088:Oxidoreductases / Q000378:metabolism*; D010920:Placenta / Q000201:enzymology; D011247:Pregnancy; D019704:Protein Disulfide-Isomerases; D011506:Proteins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013552:Swine; D013879:Thioredoxins / Q000302:isolation & purification / Q000378:metabolism; D013950:Thymus Gland / Q000201:enzymology
D054477:Glutaredoxins; D011506:Proteins; D011994:Recombinant Proteins; D013879:Thioredoxins; D010088:Oxidoreductases; C020666:glutathione dehydrogenase (ascorbate); D007535:Isomerases; D019704:Protein Disulfide-Isomerases