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    Comparative proteomic analysis of the contractile-protein-depleted fraction from normal versus dystrophic skeletal muscle


    Carberry, Steven and Zweyer, Margit and Swandulla, Dieter and Ohlendieck, Kay (2014) Comparative proteomic analysis of the contractile-protein-depleted fraction from normal versus dystrophic skeletal muscle. Analytical Biochemistry, 446. pp. 108-115. ISSN 0003-2697

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    Abstract

    In basic and applied myology, gel-based proteomics is routinely used for studying global changes in the protein constellation of contractile fibers during myogenesis, physiological adaptations, neuromuscular degeneration, and the natural aging process. Since the main proteins of the actomyosin apparatus and its auxiliary sarcomeric components often negate weak signals from minor muscle proteins during proteomic investigations, we have here evaluated whether a simple prefractionation step can be employed to eliminate certain aspects of this analytical obstacle. To remove a large portion of highly abundant contractile proteins from skeletal muscle homogenates without the usage of major manipulative steps, differential centrifugation was used to decisively reduce the sample complexity of crude muscle tissue extracts. The resulting protein fraction was separated by two-dimensional gel electrophoresis, and 2Dlandmark proteins were identified by mass spectrometry. To evaluate the suitability of the contractileprotein- depleted fraction for comparative proteomics, normal versus dystrophic muscle preparations were examined. The mass spectrometric analysis of differentially expressed proteins, as determined by fluorescence difference in-gel electrophoresis, identified 10 protein species in dystrophic mdx hindlimb muscles. Interesting new biomarker candidates included Hsp70, transferrin, and ferritin, whereby their altered concentration levels in dystrophin-deficient muscle were confirmed by immunoblotting.

    Item Type: Article
    Keywords: Dystrophinopathy; Ferritin; Hsp70; Muscle proteomics; Transferrin;
    Academic Unit: Faculty of Science and Engineering > Biology
    Item ID: 7487
    Identification Number: https://doi.org/10.1016/j.ab.2013.08.004
    Depositing User: Prof. Kay Ohlendieck
    Date Deposited: 07 Oct 2016 10:44
    Journal or Publication Title: Analytical Biochemistry
    Publisher: Elsevier
    Refereed: Yes
    Funders: Muscular Dystrophy Ireland, Duchenne Ireland, Health Research Board (HRB)
    URI:

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