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Protein glutathiolation by nitric oxide: an intracellular mechanism regulating redox protein modification
Author(s) -
West Matthew B.,
Hill Bradford G.,
Xuan Yu-Ting,
Bhatnagar Aruni
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.06-5843fje
Subject(s) - glutathione , heat shock protein , nitric oxide synthase type iii , nitric oxide synthase , nitrotyrosine , chemistry , nitric oxide , cytosol , glutathione disulfide , biochemistry , biopterin , enos , biology , endocrinology , tetrahydrobiopterin , enzyme , gene
This study was designed to examine whether NO regulates protein glutathiolation. Exposure to NO donors increased protein glutathiolation in COS‐7 or rat aortic smooth muscle cells as detected by anti‐protein glutathione (GSH) antibodies. This process was reversible and saturable. Stimulation with acetylcholine (ACh) increased protein glutathiolation in isolated rat aortic rings. This was prevented by inhibiting endothelial NO synthase (eNOS). In ACh‐treated rings, proteins showing positive immunoreactivity with the anti‐PSSG antibody (Ab) were identified by matrix assisted laser desorption‐time‐of‐flight mass spectrometry to be actin, vimentin, and heat shock protein 70. Purified actin was more readily glutathiolated by Snitrosoglutathione than by oxidized GSH as determined by electrospray‐ionization mass spectrometry, and nitrosylated actin was glutathiolated by reduced GSH. Relative to wild‐type (WT) mice, increased protein glutathiolation was observed in hearts of mice with cardiac‐specific expression of inducible NO synthase (iNOS). Proteins immunoprecipitated from transgenic hearts revealed GSH‐adducted peptides corresponding to adenine nucleotide translocator and the α‐subunit of F 1 F 0 ATPase. These data suggest that exogenous NO or NO generated by eNOS or iNOS regulates protein adduction with GSH. This could be due to a direct reaction of proteins with S‐nitrosoglutathione or denitrosylation of S‐nitrosylated proteins by reduced GSH. Glutathiolation of cytoskeletal and mitochondrial proteins may be a significant feature of NO bioreactivity.—West, M. B., Hill, B. G., Xuan, Y.‐T., Bhatnagar, A. Protein glutathiolation by nitric oxide: An intracellular mechanism regulating redox protein modification FASEB J. 20, E1049–E1060 (2006)