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Titolo:
INFLAMMATION AND NOX-INDUCED NITRATION - ASSAY FOR 3-NITROTYROSINE BYHPLC WITH ELECTROCHEMICAL DETECTION
Autore:
SHIGENAGA MK; LEE HH; BLOUNT BC; CHRISTEN S; SHIGENO ET; YIP H; AMES BN;
Indirizzi:
UNIV CALIF BERKELEY,DIV BIOCHEM & MOL BIOL BERKELEY CA 94720
Titolo Testata:
Proceedings of the National Academy of Sciences of the United Statesof America
fascicolo: 7, volume: 94, anno: 1997,
pagine: 3211 - 3216
SICI:
0027-8424(1997)94:7<3211:IANN-A>2.0.ZU;2-3
Fonte:
ISI
Lingua:
ENG
Soggetto:
NITRIC-OXIDE SYNTHASE; L-ARGININE; PEROXYNITRITE FORMATION; NITROGEN-DIOXIDE; SUPEROXIDE; PROTEINS; INHIBITION; INJURY; NITROTYROSINE; NITROSATION;
Keywords:
PEROXYNITRITE; NITRIC OXIDE SYNTHASE; REACTIVE NITROGEN OXIDE SPECIES; ZYMOSAN;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
36
Recensione:
Indirizzi per estratti:
Citazione:
M.K. Shigenaga et al., "INFLAMMATION AND NOX-INDUCED NITRATION - ASSAY FOR 3-NITROTYROSINE BYHPLC WITH ELECTROCHEMICAL DETECTION", Proceedings of the National Academy of Sciences of the United Statesof America, 94(7), 1997, pp. 3211-3216

Abstract

The identification of N-15-labeled 3-nitrotyrosine (NTyr) by gas chromatography/mass spectroscopy in protein hydrolyzates from activated RAW 264.7 macrophages incubated with N-15-L-arginine confirms that nitric oxide synthase (NOS) is involved In the nitration of protein-bound tyrosine (Tyr), An assay is presented for NTyr that employs HPLC with tandem electrochemical and UV detection. The assay involves enzymatic hydrolysis of protein, acetylation. solvent extraction, O-deacetylation, and dithionite reduction to produce an analyte containing N-acetyl-3-aminotyrosine, an electrochemically active derivative of NTyr, We estimate the level of protein-bound NTyr in normal rat plasma to be approximate to 0-1 residues per 10(6) Tyr with a detection limit of 0.5 per10(7) Tyr when greater than or equal to 100 nmol of Tyr is analyzed and when precautions are taken to limit nitration artifacts, Zymosan-treated RAW 264.7 cells were shown to have an approximate to 6-fold higher level of protein-bound NTyr compared with control cells and cells treated with N-G-monomethyl-L-arginine, an inhibitor of NOS, Intraperitoneal injection of F344 rats with zymosan led to a marked elevation inprotein-bound NTyr to approximate to 13 residues per 10(6) Tyr, an approximate to 40-fold elevation compared with plasma protein of untreated rats; cotreatment with N-G-monomethyl-L-arginine inhibited the formation of NTyr in plasma protein from blood and peritoneal exudate by 69% and 53%, respectively. This assay offers a highly sensitive and quantitative approach for investigating the role of reactive byproducts of nitric oxide in the many pathological conditions and disease states associated with NOx exposure such as inflammation and smoking.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 25/11/20 alle ore 10:15:37