Modulation of lysozyme function and degradation after nitration with peroxynitrite.
BACKGROUND
Peroxynitrite (PN) is formed from superoxide and nitric oxide, both of which are increased during hepatic ethanol metabolism. Peroxynitrite forms adducts with proteins, causing structural and functional alterations. Here, we investigated PN-induced alterations in lysozyme structure and function, and whether they altered the protein's susceptibility to proteasome-catalyzed degradation.


METHODS
Hen egg lysozyme was nitrated using varying amounts of either PN or the PN donor, 3-morpholinosydnonimine (SIN-1). The activity, nitration status and the susceptibility of lysozyme to proteasome-catalyzed degradation were assessed.


RESULTS
Lysozyme nitration by PN or SIN-1 caused dose-dependent formation of 3-nitrotyrosine-lysozyme adducts, causing decreased catalytic activity, and enhanced susceptibility to degradation by the 20S proteasome. Kinetic analyses revealed an increased affinity by the 20S proteasome toward nitrated lysozyme compared with the native protein.


CONCLUSIONS
Lysozyme nitration enhances the affinity of the modified enzyme for degradation by the proteasome, thereby increasing its susceptibility to proteolysis.


CONCLUSIONS
Increased levels of peroxynitrite have been detected in tissues of ethanol-fed animals. The damaging effects from excessive peroxynitrite in the cell increase hepatotoxicity and cellular death by protein modification due to nitration. Cellular defenses against such changes include enhanced proteolysis by the proteasome in order to maintain protein quality control.
Curry-McCoy
Tiana V
TV
Liver Study Unit, Omaha Veterans Affairs Medical Center, USA. tcurrymccoy@unmc.edu
Osna
Natalia A
NA
Donohue
Terrence M
TM
D000818:Animals; D055162:Biocatalysis; D002460:Cell Line; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D005470:Fluorometry; D006801:Humans; D009113:Muramidase / Q000378:metabolism*; D030421:Peroxynitrous Acid / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D013057:Spectrum Analysis; D013997:Time Factors
D030421:Peroxynitrous Acid; D009113:Muramidase; D046988:Proteasome Endopeptidase Complex