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 journalArticle

3 Citations (Scopus)

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 1 2016

Fingerprint

Lysine
lysine
antibodies
Antibodies
Peptides
peptides
peptide libraries
Cell signaling
Peptide Library
Acetylation
consensus sequence
HEK293 Cells
post-translational modification
Consensus Sequence
acetylation
Post Translational Protein Processing
Mass spectrometry
Mass Spectrometry
complement
Proteins

ASJC Scopus subject areas

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

Cite this

Kim, S. Y., Sim, C. K., Zhang, Q., Tang, H., Brunmeir, R., Pan, H., ... Xu, F. (2016). An alternative strategy for pan-acetyl-lysine antibody generation. PLoS One, 11(9), [e0162528]. https://doi.org/10.1371/journal.pone.0162528

An alternative strategy for pan-acetyl-lysine antibody generation. / Kim, Sun Yee; Sim, Choon Kiat; Zhang, Qiongyi; Tang, Hui; Brunmeir, Reinhard; Pan, Hong; Karnani, Neerja; Han, Weiping; Zhang, Kangling; Xu, Feng.

In: PLoS One, Vol. 11, No. 9, e0162528, 01.09.2016.

Research output: Contribution to journalArticle

Kim, SY, Sim, CK, Zhang, Q, Tang, H, Brunmeir, R, Pan, H, Karnani, N, Han, W, Zhang, K & Xu, F 2016, 'An alternative strategy for pan-acetyl-lysine antibody generation', PLoS One, vol. 11, no. 9, e0162528. https://doi.org/10.1371/journal.pone.0162528
Kim SY, Sim CK, Zhang Q, Tang H, Brunmeir R, Pan H et al. An alternative strategy for pan-acetyl-lysine antibody generation. PLoS One. 2016 Sep 1;11(9). e0162528. https://doi.org/10.1371/journal.pone.0162528
Kim, Sun Yee ; Sim, Choon Kiat ; Zhang, Qiongyi ; Tang, Hui ; Brunmeir, Reinhard ; Pan, Hong ; Karnani, Neerja ; Han, Weiping ; Zhang, Kangling ; Xu, Feng. / An alternative strategy for pan-acetyl-lysine antibody generation. In: PLoS One. 2016 ; Vol. 11, No. 9.
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