First evidence for an LDL- and HDL-associated nitratase activity that denitrates albumin-bound nitrotyrosine--physiological consequences.
In the systemic circulation, LDL occurs in the form of a weakly nitrated LDL-albumin complex (LAC). The question here is whether LAC (or HDL) is able to denitrate the albumin-bound 3-NO(2)-tyrosine (3NT). Nitrated albumin was incubated in the presence of lipoprotein fraction (LPF) to be tested, with or without Ca(2+). After precipitation and centrifugation, supernatants (SNs) and protein pellets (PP) were collected. HCl proteolysis was carried out with deuterated 3NT as an internal standard, and amino acids were derivatized for GC-MS analysis, whereas SNs were used for NO(2) (-)/NO(3) (-)-fluorimetric assays. A loss of 3NT, higher with albumin-low LDL than with albumin-rich LDL or HDL, was found in PP only in the presence of Ca(2+). gamma-Tocopherol loading of LPF inhibited 3NT loss. 3NT loss was found for the first time to be stoichiometrically equivalent to NO(3) (-), proving that the 3NT loss must be ascribed to a 3NT-denitrating nitratase activity. 3NT loss and NO(3) (-) production that clearly cannot be attributed to PON-1 were impaired by D-penicillamine and phenylacetate, inhibitor, and substrate of PON-1, respectively, leading to speculate on the active site. Finally, nitratase activity and albumin contribute to beneficially convert peroxynitrite (ONOO(-)) into nonbioactive NO(3) (-). But, in inflammatory conditions, xanthine oxidoreductase is expressed leading to detrimentally reduce O(2) and NO(3) (-) into O(2) (*) (-) and NO(*) that may interact, reconstituting the ONOO(-) pool. The real consequence of nitratase activity and the physiological significance of nitration/denitration processes remain to be explored.
Léger
Claude L
CL
EA 2993 Nutrition Humaine et Athérogénèse, Institut de Biologie, Boulevard Henri IV, Cedex 2, Montpellier, France. claude.leger@univ-montp1.fr
Torres-Rasgado
Enrique
E
Fouret
Gilles
G
Carbonneau
Marie-Annette
MA
D002118:Calcium / Q000494:pharmacology; D006801:Humans; D008075:Lipoproteins, HDL / Q000378:metabolism*; D008077:Lipoproteins, LDL / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism*; D030421:Peroxynitrous Acid / Q000378:metabolism; D012709:Serum Albumin / Q000378:metabolism*; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
D008075:Lipoproteins, HDL; D008077:Lipoproteins, LDL; D012709:Serum Albumin; D030421:Peroxynitrous Acid; C002744:3-nitrotyrosine; D014443:Tyrosine; D010088:Oxidoreductases; C045258:nitratase; D002118:Calcium