Thioredoxin catalyzes the denitrosation of low-molecular mass and protein S-nitrosothiols.
While most proteins have critical thiols whose oxidation affects their activity, it has been suggested that S-nitrosation and denitrosation of cellular thiols are fundamental processes similar to protein phosphorylation and dephosphorylation, respectively. However, understanding the biosynthesis and catabolism of S-nitrosothiols has proven to be difficult, in part because of the low stability of this class of metabolites. Herein, we report that thioredoxin catalyzes the denitrosation of a series of S-nitrosoamino acids and S-nitrosoproteins derived from HepG2 cells. Notably, all S-nitrosoproteins with a molecular mass of 23-30 kDa were catabolized by thioredoxin. Experimental evidence is presented which shows that both glutathione and reduced human thioredoxin denitrosate S-nitrosothioredoxin, which has been suggested to act as an anti-apoptotic factor via trans-S-nitrosation of caspase 3. In HepG2 cells, we observed that S-nitrosocysteine ethyl ester impedes the activity of caspase 3. However, a subsequent incubation of the cells in nitrosothiol-free medium resulted in reconstitution of the enzymatic activity, most likely due to endogenous denitrosation of S-nitrosocaspase 3. The latter process was markedly inhibited in thioredoxin reductase-deficient HepG2 cells, suggesting that the thioredoxin/thioredoxin reductase system tends to maintain intracellular caspase 3 in a reduced, SH state. The data obtained are discussed within the general reaction mechanisms encompassing the cellular homeostasis of S-nitrosothiols.
Sengupta
Rajib
R
Department of Surgery, The University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, USA.
Ryter
Stefan W
SW
Zuckerbraun
Brian S
BS
Tzeng
Edith
E
Billiar
Timothy R
TR
Stoyanovsky
Detcho A
DA
D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D002384:Catalysis; D045744:Cell Line, Tumor; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D024101:Mitochondrial Proteins / Q000378:metabolism; D008970:Molecular Weight; D015538:Nitrosation; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D026403:S-Nitrosothiols / Q000378:metabolism*; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism; D013879:Thioredoxins / Q000737:chemistry / Q000378:metabolism*
D000596:Amino Acids; D024101:Mitochondrial Proteins; D011506:Proteins; D026403:S-Nitrosothiols; C467002:TXN2 protein, human; D013879:Thioredoxins; D013880:Thioredoxin-Disulfide Reductase; D053148:Caspase 3; D005978:Glutathione