Tip60 acetylates six lysines of a specific class in core histones in vitro.
BACKGROUND
Tip60, an HIV-1-Tat interactive protein, is a nuclear histone acetyltransferase (HAT) with unique histone substrate specificity. Since the acetylation of core histones at particular lysines mediates distinct effects on chromatin assembly and gene regulation, the identification of lysine site specificity of the HAT activity of Tip60 is an initial step in the analysis of its molecular function.


RESULTS
Tip60 significantly acetylates amino-terminal tail peptides of histones H2A, H3 and H4, but not H2B, consistent with substrate preference on intact histones. Preferred acetylation sites for Tip60 are the Lys-5 of histone H2A, the Lys-14 of histone H3, and the Lys-5, -8, -12, -16 of histone H4, determined by a method which combined matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) measurements and Lys-C endopeptidase digestion, or a method detecting the incorporation of radiolabelled acetate into synthetic peptides.


CONCLUSIONS
The lysine site specificity of Tip60 in histone amino-terminal tail peptides in vitro has been characterized by an assay measuring the molecular mass of endopeptidase digested peptides, or a previously described assay. These results agree well with our proposed classification of lysines in core histones. The classification may be useful for an analysis of the relationships between HATs and the substrates of other uncharacterized HATs.
Kimura
A
A
Department of Cellular Biology, University of Tokyo, Japan.
Horikoshi
M
M
D000107:Acetylation; D000123:Acetyltransferases; D000595:Amino Acid Sequence; D051548:Histone Acetyltransferases; D006657:Histones / Q000378:metabolism*; D008239:Lysine / Q000378:metabolism; D000074443:Lysine Acetyltransferase 5; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D013379:Substrate Specificity
D006657:Histones; D010446:Peptide Fragments; D011506:Proteins; D000123:Acetyltransferases; D051548:Histone Acetyltransferases; C098489:KAT5 protein, human; D000074443:Lysine Acetyltransferase 5; D008239:Lysine