Peroxynitrite alters the catalytic activity of rodent liver proteasome in vitro and in vivo

• Published in: Hepatology (Baltimore, Md.) Vol. 40; no. 3; pp. 574 - 582
• Main Authors: Osna, Natalia A., Haorah, James, Krutik, Viatcheslav M., Donohue, Terrence M.
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01-09-2004; Wiley
• Subjects: Animals; Biological and medical sciences; Catalysis; Cysteine Endopeptidases - drug effects; Cysteine Endopeptidases - metabolism; Dose-Response Relationship, Drug; Fundamental and applied biological sciences. Psychology; Liver - enzymology; Liver. Bile. Biliary tracts; Mice; Mice, Inbred C57BL; Molsidomine - analogs & derivatives; Molsidomine - pharmacology; Multienzyme Complexes - drug effects; Multienzyme Complexes - metabolism; Peroxynitrous Acid - pharmacology; Proteasome Endopeptidase Complex; Rats; S-Nitroso-N-Acetylpenicillamine - pharmacology; Vertebrates: digestive system; Human; Peptides; Rat; Digestive system; Enzyme; Liver; Rodentia; In vitro; Biological activity; Protein; Prevention; Assay; In vivo; Peptidases; Signal transduction; Vertebrata; Mammalia; Donor; Mouse; Animal; Preparation; Hydrolases
• ISSN: 0270-9139; 1527-3350
• DOI: 10.1002/hep.20352

The proteasome is an important multicatalytic enzyme complex that degrades misfolded and oxidized proteins, signal transduction factors, and antigenic peptides for presentation. We investigated the in vitro effects of peroxynitrite (PN) on the peptidase activity of both crude 20S and 26S and purified 20S proteasome preparations from rat liver as well as proteasome activity in Hep G2 cells and in mouse liver. Crude and purified proteasome preparations were exposed to PN or to the PN donor, 3‐morpholinosydnonimine hydrochloride (SIN‐1), and then assayed for chymotrypsin‐like activity. For in vivo experiments, mice were treated with molsidomine, which is metabolized to SIN‐1 in liver. PN and SIN‐1 dose‐dependently modulated the chymotrypsin‐like activity of the 20S proteasome: lower concentrations enhanced proteasome activity, and higher concentrations caused its decline. The NO donor S‐nitroso‐N‐acetylpenicillamine (SNAP), at all concentrations, suppressed 20S proteasome activity. We observed similar results when liver soluble fractions (S‐100) were treated with PN, SIN‐1, or SNAP, except that enzyme activity in S‐100 fractions was less sensitive than the purified enzymes to these agents. Treatment of Hep G2 cells with 0.01 or 0.1 mmol/L SIN‐1 stimulated in situ proteasome activity in these cells, while 1 mmol/L SIN‐1 suppressed it. SNAP treatment did not affect proteasome activity in Hep G2 cells. Mice treated with molsidomine had enhanced liver proteasome activity 6 hours after treatment, but after 24 hours enzyme activity declined below control levels. In conclusion, PN dose‐dependently modulated proteasome activity, regulating protein degradation by the proteasome in liver cells. (HEPATOLOGY 2004;40:574–582.)