Journal
JOURNAL OF PROTEOME RESEARCH
Volume 11, Issue 4, Pages 2441-2451Publisher
AMER CHEMICAL SOC
DOI: 10.1021/pr201169q
Keywords
brain; proteomics; membrane proteins (MPs); transmembrane proteins (TMPs); mass spectrometry (MS); sample preparation; extraction
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Funding
- Swedish Governmental Agency for Innovation Systems
- Swedish Research Council [621-2008-3562]
- Uppsala University
- Swedish Institute
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This study compares 16 different extraction methods for the comprehensive extraction of mouse brain proteome in combination with shotgun-based mass spectrometry (MS). Membrane proteins (MPs) are responsible for a large part of the regulatory functions of the cell and are therefore of great interest to extract and analyze. Sixteen protein extraction protocols were evaluated in regards to protein yield and number of identified proteins with emphasis on MPs. The extracted proteins were delipidated, on-filter digested, and analyzed by reversed phase nanoliquid chromatography (RP-nanoLC) in combination with electrospray ionization (ESI) tandem mass spectrometry (MS/MS) using a 7 T hybrid LTQ: FT mass spectrometer. Detergent-based lysis buffers showed higher efficiencies and yields in the extraction of proteins from the brain tissue compared to solubilization with organic solvents or organic acids. The detergent octyl-beta-D-glucopyranoside gave the highest number of identified proteins (541) as well as numbers and percentages of identified MPs (29%). Detergent-based protocols are the best sample preparation tools for central nervous system (CNS) tissue and can readily be applied to screen for candidate biomarkers of neurological diseases.
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