Characterization of lysine-guanine cross-links upon one-electron oxidation of a guanine-containing oligonucleotide in the presence of a trilysine peptide.
Formation of DNA-protein cross-links involving the initial formation of a guanine radical cation was investigated. For this purpose, riboflavin-mediated photosensitization of a TGT oligonucleotide in aerated aqueous solution in the presence of the KKK tripeptide was performed. We have shown that the nucleophilic addition of the epsilon-amino group of the central lysine residue of KKK to the C8 atom of either the guanine radical cation or its deprotonated form gives rise to the efficient formation of a Nepsilon-(guanin-8-yl)-lysine cross-link. Interestingly, the time course of formation of the above-mentioned cross-link was found to be not linear with the time of irradiation, and its formation rapidly reached a plateau. This is explained by secondary decomposition of the initially generated cross-link which could be further oxidized more efficiently than starting TGT oligonucleotide. One-electron oxidation of the initially generated cross-link was found to produce mainly two diastereomeric cross-links exhibiting a spiroimino-trilysine-dihydantoin structure as inferred from enzymatic digestion, CD, UV, NMR and mass spectrometry measurements. In addition, other minor cross-links, for which formation was favored at acidic pH, were assigned as lysine-guanine adducts in which the modified guanine base exhibits a guanidino-trilysine-iminohydantoin structure. A proposed mechanism for the formation of the different detected oligonucleotide-peptide cross-links is given. The high yield of formation of the detected cross-links strongly suggests that a DNA-protein cross-link involving a lysine residue linked to the C8 position of guanine could be generated in cellular systems if a lysine is located in the close vicinity of a guanine radical cation.
Perrier
Sandrine
S
Laboratoire Lésions des Acides Nucléiques, DRFMC/SCIB UMR-E3 CEA-UJF, CEA-Grenoble, 17 rue des Martyrs, F-38054 Grenoble Cedex 9, France.
Hau
Jörg
J
Gasparutto
Didier
D
Cadet
Jean
J
Favier
Alain
A
Ravanat
Jean-Luc
JL
D002851:Chromatography, High Pressure Liquid; D004583:Electrons; D006147:Guanine / Q000737:chemistry*; D008239:Lysine / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular; D009841:Oligonucleotides / Q000737:chemistry*; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization
D009841:Oligonucleotides; D010455:Peptides; D006147:Guanine; D008239:Lysine