Epidemiologic evidence shows a strong correlation between chronic inflammation and the subsequent likelihood of damaged cells becoming cancerous. However, the mechanism whereby inflammation initiates or contributes to carcinogenesis is unknown. Emerging data in the field of epigenetics suggest that histone modifications play an integral role to modify gene expression paving the way for yet another gene regulatory system that may become perturbed, leading to cancer. Hypochlorous acid, formed during the “respiratory burst” of inflammation-mediated processes, has been shown to oxidize and chlorinate host lipids, proteins, RNA, and DNA while fulfilling its main antimicrobial purpose. We hypothesize that HOCl-mediated damage of histones may affect the gene regulatory system and show that we have a method and a marker to measure the amounts of inflammation-mediated damage to histone proteins. Chlorotyrosine (ClY) has been shown to be a stable product of HOCl-mediated damage to proteins. Measuring this marker with a sensitive gas chromatography-negative chemical ionization-mass spectroscopy (GC-NCI-MS) allows us to quantitatively measure the amounts of ClY formed when calf thymus histones H2a, H2b, H3, and H4 are reacted with HOCl. Digesting these reactants with proteases and analyzing with MALDI/TOF and LC/ESI mass spectroscopy allow us to determine where these chlorination adducts are formed in the protein sequence. We found that ClY is formed in higher amounts than expected from the reaction rates of tyrosine groups to HOCl. We, like others, have also found that tyrosines found in an YXXK or KXXY are chlorinated more than other tyrosines, suggesting that there may be some sequence specificity to this reaction. By elucidating the mechanism of HOCl-mediated histone damage, we hope to better understand how chronic inflammation may perturb the epigenetic regulation of gene expression and promote the transformation of normal cells to cancer.
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