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    Proteome-wide enrichment of proteins modified by lysine methylation

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    Main article (584.7Kb)
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    https://www.nature.com/articles/nprot.2013.164
    Permanent Link
    https://doi.org/10.1038/nprot.2013.164
    http://hdl.handle.net/11603/26714
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    • UMBC Biological Sciences Department
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    Author/Creator
    Carlson, Scott M
    Moore, Kaitlyn E
    Green, Erin
    Martín, Glòria Mas
    Gozani, Or
    Author/Creator ORCID
    https://orcid.org/0000-0003-3923-6726
    Date
    2013-12-05
    Type of Work
    27 pages
    Text
    journal articles
    postprints
    Citation of Original Publication
    Carlson, Scott M et al. “Proteome-wide enrichment of proteins modified by lysine methylation.” Nature protocols vol. 9,1 (2014): 37-50. doi:10.1038/nprot.2013.164
    Rights
    This version of the article has been accepted for publication, after peer review (when applicable) and is subject to Springer Nature’s AM terms of use, but is not the Version of Record and does not reflect post-acceptance improvements, or any corrections. The Version of Record is available online at: http://dx.doi.org/10.1038/nprot.2013.164
    Abstract
    We present a protocol for using the triple malignant brain tumor domains of L3MBTL1 (3×MBT), which bind to mono- and di-methylated lysine with minimal sequence specificity, in order to enrich for such methylated lysine from cell lysates. Cells in culture are grown with amino acids containing light or heavy stable isotopic labels. Methylated proteins are enriched by incubating cell lysates with 3×MBT, or with the binding-null D355N mutant as a negative control. Quantitative liquid chromatography and tandem mass spectrometry (LC-MS/MS) are then used to identify proteins that are specifically enriched by 3×MBT pull-down. The addition of a third isotopic label allows the comparison of protein lysine methylation between different biological conditions. Unlike most approaches, our strategy does not require a prior hypothesis of candidate methylated proteins, and it recognizes a wider range of methylated proteins than any available method using antibodies. Cells are prepared by growing in isotopic labeling medium for about 7 d; the process of enriching methylated proteins takes 3 d and analysis by LC-MS/MS takes another 1–2 d.


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    Albin O. Kuhn Library & Gallery
    University of Maryland, Baltimore County
    1000 Hilltop Circle
    Baltimore, MD 21250
    www.umbc.edu/scholarworks

    Contact information:
    Email: scholarworks-group@umbc.edu
    Phone: 410-455-3021


    If you wish to submit a copyright complaint or withdrawal request, please email mdsoar-help@umd.edu.