Protein S-thiolation in hepatocytes stimulated by t-butyl hydroperoxide, menadione, and neutrophils.
In order to examine potentially important S-thiolated proteins, 35S-labeled hepatocytes were exposed to oxidative stress. A similar group of S-thiolated proteins including carbonic anhydrase III was observed in cells treated with t-butyl hydroperoxide, menadione, or stimulated neutrophils. The radioactive thiols bound to hepatocyte proteins were identified by HPLC and more than 85% was glutathione. In menadione-treated hepatocytes, proteins were gradually S-thiolated over 30 min and 25% of the cellular glutathione pool became protein-bound. In t-butyl hydroperoxide-treated cells, S-thiolation was more transient and 11% of the glutathione was protein-bound. Neutrophil-treated hepatocytes had nearly the same amount of protein S-thiolation (8% after 25 min). Two major proteins that were S-thiolated in untreated hepatocytes did not increase during any form of oxidative stress. In neutrophil-treated hepatocytes protein S-thiolation was not accompanied by either formation of glutathione disulfide or a measurable change in the total amount of glutathione. In both t-butyl hydroperoxide-and menadione-treated cells there was extensive formation of glutathione disulfide and in menadione-treated cells a significant increase in the total hepatocyte glutathione pool was observed. This result suggests that protein S-thiolation may occur by mechanisms that do not result from thiol/disulfide exchange between glutathione disulfide and protein sulfhydryls. It is suggested that a thiyl radical intermediate is important in neutrophil-mediated protein S-thiolation.
Chai
Y C
YC
Department of Biochemistry, Iowa State University, Ames 50011.
Hendrich
S
S
Thomas
J A
JA
D000818:Animals; D002256:Carbonic Anhydrases / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D009504:Neutrophils / Q000378:metabolism; D010545:Peroxides / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D014812:Vitamin K / Q000494:pharmacology; D020122:tert-Butylhydroperoxide
D004220:Disulfides; D010545:Peroxides; D011506:Proteins; D013438:Sulfhydryl Compounds; D014812:Vitamin K; D020122:tert-Butylhydroperoxide; D002256:Carbonic Anhydrases; D005978:Glutathione