Lysine-8 and lysine-12 are biotinylated in human histone H4

GABRIELA CAMPOREALE, GAUTAM SARATH, JANOS ZEMPLENI

Washington, DC - EEUU

Conferencia; Federation of American Societies for Experimental Biology (FASEB); 2004

Folding of DNA into chromatin is mediated by binding to histones such as H4; association of DNA with histones is regulated by covalent modifications of histones. Here we tested the hypothesis that specific amino acids in histone H4 are biotinylated. Peptides derived from human histone H4 were synthesized chemically and biotinylated enzymatically using biotinidase. Peptides were electrophoresed and biotin was probed with streptavidin-peroxidase. A peptide corresponding to the first 22 residues from the N-terminus of H4 was efficiently recognized by biotinidase as substrate for biotinylation. In contrast, a peptide corresponding to the C-terminal 24 residues was not biotinylated. Substitution of lysines 8 or 12 by alanine or arginine in synthetic peptides based on the N-terminal sequence of H4 diminished covalent modification by biotin, suggesting that these two lysines are targets for biotinylation. Chemical acetylation of a given lysine decreased enzymatic biotinylation of neighboring lysines, suggesting cross-talk among known modifications of histones. Using a biotinylation site-specific antibody, histone H4 (biotinylated at lysine-12) was detected in extracts from human cells. We conclude that lysine-8 and lysine-12 in histone H4 are targets for biotinylation. Current studies in our laboratory suggest that these novel modifications of histone H4 plays a role in cellular response to DNA damage.