Abstract：
:Histone acetylation is a dynamic epigenetic modification that functions in the regulation of DNA-templated reactions, such as transcription. This lysine modification is reversibly controlled by histone (lysine) acetyltransferases and deacetylases. Here, we present methods employing isotopic labeling and mass spectrometry (MS) to comprehensively investigate histone acetylation dynamics. Turnover rates of histone acetylation are determined by measuring the kinetics of labeling from (13)C-labeled precursors of acetyl-CoA, which incorporates (13)C-carbon onto histones via the acetyltransferase reaction. Overall histone acetylation states are assessed from complete protease digestion to single amino acids, which is followed by MS analysis. Determination of site-specific acetylation stoichiometry is achieved by chemically acetylating endogenous histones with isotopic acetic anhydride, followed by trypsin digestion and LC-MS analysis. Combining metabolic labeling with stoichiometric analysis permits determination of both acetylation level and acetylation dynamics. When comparing genetic, diet, or environmental perturbations, these methods permit both a global and site-specific evaluation of how histone acetylation is dynamically regulated.
journal_name
Methods Enzymoljournal_title
Methods in enzymologyauthors
Fan J,Baeza J,Denu JMdoi
10.1016/bs.mie.2016.01.007subject
Has Abstractpub_date
2016-01-01 00:00:00pages
125-148eissn
0076-6879issn
1557-7988pii
S0076-6879(16)00020-3journal_volume
574pub_type
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