A non-isotopic in vitro assay for histone acetylation

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• 作者：David Kuninger ; James Lundblad ; Anthony Semirale ; Peter Rotwein
• 关键词：Non-isotopic ; Acetyl-transferase assay ; p300 ; PCAF ; Histone H3 ; Histone H4
• 刊名：Journal of Biotechnology
• 出版年：2007
• 出版时间：15 September 2007
• 年：2007
• 卷：131
• 期：3
• 页码：253-260
• 全文大小：751 K

文摘

We describe a simple, robust, and relatively inexpensive non-radioactive in vitro assay for measuring histone acetyl-transferase activity. The assay takes advantage of easy to purify recombinant E. coli-derived fusion proteins containing the NH₂-terminal tails of histones H3 and H4 linked to epitope-tagged maltose-binding protein (MBP), and immunoblotting with antibodies specific to acetylated H3 and H4. Here we show the specificity and dynamic range of this assay for the histone acetyl-transferases, p300 and PCAF. This assay may be adapted readily for other substrates by simply generating new fusion proteins and for other acetyl-transferases by modifying reaction conditions.