Hydroxyl radical oxidation of guanosine 5'-triphosphate (GTP): requirement for a GTP-Cu(II) complex.
Levels of oxidized guanosine base in DNA have become a hallmark biomarker in assessing oxidative stress implicated in a variety of disease and toxin-induced states. However, there is evidence that the guanosine in the nucleotide triphosphate pool (GTP) is more susceptible to oxidation than guanosine residues incorporated into nucleic acids and this causes a substantial amount of the oxidized product, 8-oxoguanosine 5'-triphosphate (oxo(8)GTP), to accumulate in cell-free and in cell-culture preparations. Electron paramagnetic resonance (EPR) spectroscopy and direct EPR analysis of free radical production by copper sulfate and L-ascorbic acid demonstrates that the hydroxyl radical (HO(*)) is produced via oxidation of Cu(+) to Cu(2+) while in a complex with GTP. This HO(*) production is dependent on the availability of oxygen and the presence of GTP in the reaction milieu. Verification of free radical-mediated production of oxo(8)GTP is presented using HPLC with electrochemical detection and matrix-assisted laser desorption/ionization linear time-of-flight mass spectrometry (MALDI-LTOF-MS). The sum of these results is presented in a novel mechanism of GTP oxidation by Cu(2+) and L-ascorbic acid. A better understanding of the chemistry involved in this oxidative modification of GTP facilitates a more comprehensive understanding of its potential physiological consequences.
Cerchiaro
Giselle
G
Centro de Ciências Naturais e Humanas, Universidade Federal do ABC, Santo André-SP, Brazil.
Bolin
Celeste
C
Cardozo-Pelaez
Fernando
F
D001205:Ascorbic Acid / Q000737:chemistry; D003300:Copper / Q000737:chemistry*; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D006160:Guanosine Triphosphate / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D017665:Hydroxyl Radical / Q000737:chemistry*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010101:Oxygen Consumption; D017382:Reactive Oxygen Species / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D017382:Reactive Oxygen Species; D017665:Hydroxyl Radical; D003300:Copper; D006160:Guanosine Triphosphate; D004247:DNA; D001205:Ascorbic Acid