Characterization of radiation-induced thymine-tyrosine crosslinks by electrospray ionization mass spectrometry.
Exposure to ionizing radiation leads to formation of covalent crosslinks between DNA and proteins. The nature, extent and site of the modifications are not well understood due to the difficulty in assessing free radical-induced damage in biopolymers. Electrospray ionization mass spectrometry (ESI-MS) permits direct analyses of intact oligopeptides, permitting characterization of the radiation-induced DNA-protein covalently crosslinked constituents. Our first application of this methodology to free radical-induced damage was in a model system where angiotensin, a small 10-amino acid peptide, is irradiated at various doses in the presence of excess thymine. The relative yield of crosslinks, which ranged from 0.1 to 15%, was linearly related to radiation dose for doses from 0.1 to 100 Gy. Detection of thymine-tyrosine moieties in this model system was possible at doses as low as 0.1 Gy with a signal-to-noise ratio of 4 to 1. ESI-MS revealed that the site of crosslink was located exclusively on the tyrosine residue as expected.
Weir Lipton
M S
MS
Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA 99352, USA.
Fuciarelli
A F
AF
Springer
D L
DL
Edmonds
C G
CG
D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000737:chemistry* / Q000528:radiation effects; D003432:Cross-Linking Reagents; D004247:DNA / Q000737:chemistry* / Q000528:radiation effects; D013058:Mass Spectrometry / Q000379:methods; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry; D011506:Proteins / Q000737:chemistry* / Q000528:radiation effects; D013941:Thymine / Q000737:chemistry / Q000528:radiation effects*; D014443:Tyrosine / Q000737:chemistry / Q000528:radiation effects*
D003432:Cross-Linking Reagents; D009842:Oligopeptides; D011506:Proteins; D000804:Angiotensin II; D014443:Tyrosine; D004247:DNA; D013941:Thymine