An alternative strategy for pan-acetyl-lysine antibody generation

Sun Yee Kim, Choon Kiat Sim, Qiongyi Zhang, Hui Tang, Reinhard Brunmeir, Hong Pan, Neerja Karnani, Weiping Han, Kangling Zhang, Feng Xu

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Lysine acetylation is an important post-translational modification in cell signaling. In acetylome studies, a high-quality pan-acetyl-lysine antibody is key to successful enrichment of acetylated peptides for subsequent mass spectrometry analysis. Here we show an alternative method to generate polyclonal pan-acetyl-lysine antibodies using a synthesized random library of acetylated peptides as the antigen. Our antibodies are tested to be specific for acetyl-lysine peptides/proteins via ELISA and dot blot. When pooled, five of our antibodies show broad reactivity to acetyl-lysine peptides, complementing a commercial antibody in terms of peptide coverage. The consensus sequence of peptides bound by our antibody cocktail differs slightly from that of the commercial antibody. Lastly, our antibodies are tested in a proof-of-concept to analyze the acetylome of HEK293 cells. In total we identified 1557 acetylated peptides from 416 proteins. We thus demonstrated that our antibodies are well-qualified for acetylome studies and can complement existing commercial antibodies.

Original languageEnglish (US)
Article numbere0162528
JournalPloS one
Volume11
Issue number9
DOIs
StatePublished - Sep 2016

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

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