Alpha-Ketoglutarate: A Potential Inner Mitochondrial and Cytosolic Protector against Peroxynitrite and Peroxynitrite-Induced Nitration?
| Autor: | Reinhold Wintersteiger, Ralf Herwig, Joachim Greilberger, Klaus Zangger, Michaela Greilberger |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
inorganic chemicals
Physiology Clinical Biochemistry RM1-950 Biochemistry Article law.invention Luminol chemistry.chemical_compound Alpha ketoglutarate law Nitration Nitrite Bovine serum albumin Tyrosine Molecular Biology Chemiluminescence Chromatography biology reactive oxygen and nitrogen species (RONS) Chemistry Cell Biology peroxynitrite (ONOO−) alpha-ketoglutarate (αKG) biology.protein cardiovascular system Therapeutics. Pharmacology Peroxynitrite |
| Zdroj: | Antioxidants Volume 10 Issue 9 Antioxidants, Vol 10, Iss 1501, p 1501 (2021) |
| ISSN: | 2076-3921 |
| Popis: | The generation of peroxynitrite (ONOO−) is associated with several diseases, including atherosclerosis, hypertension, neurodegeneration, cancer, inflammation, and sepsis. Alpha-ketoglutarate (αKG) is a known potential highly antioxidative agent for radical oxidative species such as peroxides. The question arises as to whether αKG is also a potential scavenger of ONOO− and a potential protector against ONOO−-mediated nitration of proteins. NMR studies of 1 mM αKG in 100 mM phosphate-buffered saline at pH 7.4 and pH 6.0 were carried out in the presence or absence of a final concentration of 2 mM ONOO−. An ONOO−–luminol-induced chemiluminescence reaction was used to measure the scavenging function of several concentrations of αKG quantification of αKG was performed via spectrophotometric enzymatic assay of αKG in the absence or presence of 0, 1, or 2 mM ONOO−. The nitration of tyrosine residues on proteins was measured on ONOO−-treated bovine serum albumin (BSA) in the presence or absence of 0–24 mM αKG by an ELISA technique using a specific anti-IgG against nitro-tyrosine. The addition of ONOO− to αKG led to the formation of succinic acid and nitrite at pH 7.0, but not at pH 6.0, as αKG was stable against ONOO−. The absorbance of enzymatically estimated αKG at the time point of 30 min was significantly lower in favour of ONOO− (1 mM: 0.21 ± 0.03, 2 mM: 0.12 ± 0.05 vs. 0 mM: 0.32 ± 0.02 p < 0.001). The luminol technique showed an inverse logarithmic correlation of the ONOO− and αKG concentrations (y = −2 × 105 ln(x) + 1 × 106 r2 = 0.99). The usage of 4 mM αKG showed a significant reduction by nearly half in the chemiluminescence signal (284,456 ± 29,293 cps, p < 0.001) compared to the control (474,401 ± 18,259) for 20 and 200 mM αKG, there were further reductions to 163,546 ± 26,196 cps (p < 0.001) and 12,658 ± 1928 cps (p < 0.001). Nitrated tyrosine residues were estimated using the ELISA technique. A negative linear correlation was obtained by estimating nitrated tyrosine residues in the presence of αKG (r2 = 0.94): a reduction by half of nitrated tyrosine was estimated using 12 mM αKG compared to the control (326.1 ± 39.6 nmol vs. 844.5 ± 128.4 nmol 0.001). |
| Databáze: | OpenAIRE |
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