Dependence of DNA-protein crosslinking on guanine radical protonation state

CHED 810

Jonah-Lynne Padigus, Alina Estonactoc, Amanda Madison, Julianna Scala, April Wayman, and Eric D. A. Stemp. Department of Physical Sciences and Mathematics, Mount St. Mary's College, 12001 Chalon Road, Los Angeles, CA 90049
Oxidative damage to DNA can cause disease and one form of oxidative damage is the DNA-protein crosslink. Guanine (G) is easily oxidized, and proton transfer from the guanine radical cation to cytosine is favorable. We are studying how the G radical protonation state influences the DNA-protein crosslinking reaction with histone. In DNA 20-mer duplexes, methyl or bromo was substituted at the C5 position to alter the cytosine basicity. Crosslinking was then induced by the flash-quench technique and gel shift and the chloroform extraction assays revealed that the crosslinking was most efficient with the 5'-bromocytosine DNA. This was expected because the cation radical is more reactive and the more weakly basic 5-bromocytosine should allow the guanine cation radical to persist longer. Preliminary results indicate that this trend holds at pH 4.5 as well as pH 7 but that the crosslink appears more labile at acidic pH.