Abstract
Membrane proteins solubilized in a starting buffer containing high concentration of SDS are directly entrapped and immobilized into gel matrix when the membrane protein solution is absorbed by the vacuum-dried polyacrylamide gel. After the detergent and other salts are removed by washing, the proteins are subjected to in-gel digestion, and the tryptic peptides are extracted and analyzed by CapLC-MS/MS. The newly developed method not only avoids protein loss and the adverse protein modifications during gel-embedment but also improves the subsequent in-gel digestion and the recovery of tryptic peptides, particularly hydrophobic peptides. Thus, this method facilitates the identification of membrane proteins, especially integral membrane proteins.
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Acknowledgments
This work was supported by grants from the Special Program for Key Basic Research of the Ministry of Science (2004CCA00300), National High Technology Research and Development Program or “863 Program” of China (2006AA02Z141), and National Basic Research Program or “973 Program” of China (2010CB529800, 2007CB914203).
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Wang, X., Liang, S. (2019). Gel Absorption-Based Sample Preparation Method for Shotgun Analysis of Membrane Proteome. In: Kurien, B., Scofield, R. (eds) Electrophoretic Separation of Proteins. Methods in Molecular Biology, vol 1855. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8793-1_41
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DOI: https://doi.org/10.1007/978-1-4939-8793-1_41
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