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Proteome Profiling of Muscle Cells and Muscle Tissue Using Stable Isotope Labeling by Amino Acids

  • Emily Canessa
  • Mansi V. Goswami
  • Alison M. Samsel
  • Michael Ogundele
  • Shefa M. Tawalbeh
  • Tchilabalo D. Alayi
  • Yetrib HathoutEmail author
Chapter
Part of the Methods in Physiology book series (METHPHYS)

Abstract

Comprehensive and accurate proteome profiling of skeletal muscle remains challenging owing to the large proteome dynamic range in this tissue and the increased sensitivity needed to detect low-abundant proteins. Sarcomeric and glycolytic enzymes are by far the most abundant proteins in muscle, masking detection and quantification of low-abundant proteins such as dystrophin, dystrophin-associated protein complex, cell signaling proteins, and transcription factors. About 5400 unique proteins have been identified so far in skeletal muscle using extensive pre-fractionation methods and mass spectrometry [1]. While this is good for cataloging the muscle proteome, pre-fractionation methods are often not compatible with quantification or comparative proteomics because of inherent technical variability from experiment to experiment leading to false and inaccurate quantification. To overcome these challenges, several stable isotope labeling strategies have been developed and tested in the past in different cell culture models and tissues. Typically, the two samples to be compared are tagged with heavy and light stable isotope-labeled moieties, respectively, either at the peptide level, after digestion of proteins with a protease (e.g., trypsin or endoproteinase Lys-C), or even at the cellular level before protein extraction and processing. This book chapter, we will focus on stable isotope labeling by amino acid in cell culture (SILAC), which is by far the most accurate method for quantitative proteomics in cell culture systems.

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Copyright information

© The American Physiological Society 2019

Authors and Affiliations

  • Emily Canessa
    • 1
  • Mansi V. Goswami
    • 1
  • Alison M. Samsel
    • 1
  • Michael Ogundele
    • 1
  • Shefa M. Tawalbeh
    • 1
  • Tchilabalo D. Alayi
    • 1
  • Yetrib Hathout
    • 1
    Email author
  1. 1.School of Pharmacy and Pharmaceutical SciencesBinghamton University – SUNYJohnson CityUSA

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