Mapping intact protein isoforms in discovery mode using top-down proteomics

John C. Tran, Leonid Zamdborg, Dorothy R. Ahlf, Ji Eun Lee, Adam D. Catherman, Kenneth R. Durbin, Jeremiah D. Tipton, Adaikkalam Vellaichamy, John F. Kellie, Mingxi Li, Cong Wu, Steve M. M. Sweet, Bryan P. Early, Nertila Siuti, Richard D. LeDuc, Philip D. Compton, Paul M. Thomas and Neil L. Kelleher ()
Additional contact information
John C. Tran: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Leonid Zamdborg: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Dorothy R. Ahlf: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Ji Eun Lee: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Adam D. Catherman: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Kenneth R. Durbin: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Jeremiah D. Tipton: Northwestern University
Adaikkalam Vellaichamy: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
John F. Kellie: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Mingxi Li: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Cong Wu: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Steve M. M. Sweet: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Bryan P. Early: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Nertila Siuti: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Richard D. LeDuc: University of Wisconsin-Madison
Philip D. Compton: Northwestern University
Paul M. Thomas: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign
Neil L. Kelleher: and the Institute for Genomic Biology, University of Illinois at Urbana-Champaign

Nature, 2011, vol. 480, issue 7376, 254-258

Abstract: Intact proteins yield to proteomics Conventional 'bottom-up' proteomics, in which mass spectrometry is used to analyse peptide mixtures made by tryptic digestion of target proteins, is a powerful way of characterizing complex proteomes. However, the technique has limitations when considering different protein isoforms and combinations of post-translational modifications. The 'top-down' approach is generally thought to be impractical because of the limitations of mass spectrometry and difficulties with automation. A new top-down system presented here avoids these problems by using a four-dimensional separation system that achieves greater proteome coverage than conventional methods. A proof-of-principle experiment shows that the method is capable of identifying previously undetected isoforms and isoform-specific post-translational modifications caused by cellular senescence.

Date: 2011
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DOI: 10.1038/nature10575

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