ANALYSIS OF RADIATION-INDUCED NUCLEOBASE-PEPTIDE CROSS-LINKS BY ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY

Upon exposure to ionizing radiation, DNA undergoes a variety of modifi cations including the production of a covalent bond between the nucleo base thymine and the amino acid tyrosine, These crosslinked lesions, p roduced in cells exposed to ionizing radiation, if unrepaired are thou ght to result in cell death, We have used electrospray ionization mass spectrometry (ESI-MS) to study a model system consisting of the pepti de angiotensin, a 10 amino acid peptide containing only one tyrosine r esidue, irradiated in the presence of the nucleobase thymine, The pres ence of the covalently crosslinked species has been determined by ESI- MS, by the appearance of additional species in the irradiated samples which correspond to the adduction of thymine as well as a hydrated spe cies containing thymine and water (5-hydroxy-6-hydrothymine). The form ation of 5-hydroxy-6-hydrothymine adduct is reversible and the relativ e abundance of the thymine and 5-hydroxy-6-hydrothymine adducts is dep endent on the pH of the spray solution, High resolution experiments us ing Fourier transform ion cyclotron resonance mass spectrometry confir ms the presence of the thymine and hydrated thymine adducts, The high resolution nature of these experiments also allows the detection of a 5,6-dihydrothymine adduct. (C) 1997 by John Wiley & Sons, Ltd.