Differential recovery of membrane proteins after extraction by aqueous methanol and trifluoroethanol

Huoming Zhang, Qingsong Lin, Sukumar Ponnusamy, Narasimhan Kothandaraman, Kwang Lim Teck, Changqing Zhao, Sook Kit Hon, Biswas Arijit, Mary Rauff, Choy Leong Hew, Maxey Ching Ming Chung, Shashikant B. Joshi, Mahesh Choolani*

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    51 Scopus citations

    Abstract

    Cell membrane proteome analysis is limited by inherent membrane hydrophobicity. Conventional membrane protein extraction techniques use detergents, chaotropes and organic acids that require sample clean-up or pH adjustment, and are associated with significant sample loss. We extracted membrane proteins from red blood cells (RBCs) using methanol (MeOH), trifluoroethanol (TFE) and urea, and identified membrane proteins using 2-D LC coupled with MALDI-TOF/TOF-MS. We show that organic solvents MeOH- and TFE-based methods have membrane protein analysis efficiencies comparable to urea, and are complementary for the recovery of both hydrophilic and hydrophobic peptides. The mean grand average of hydropathicity (GRAVY) value of identified peptides from the TFE-based method (-0.107) was significantly higher than that of the MeOH-based method (-0.465) (p<0.001). Sequential and adjunctive use of the organic solvents MeOH and TFE increases the number of proteins identified, and the confidence of their identification. We show that this strategy is effective for shotgun membrane proteome analysis.

    Original languageEnglish (US)
    Pages (from-to)1654-1663
    Number of pages10
    JournalProteomics
    Volume7
    Issue number10
    DOIs
    StatePublished - May 1 2007

    Keywords

    • 2-D LC-MALDI-TOF/TOF-MS
    • Human erythrocyte membrane protein
    • Methanol
    • Trifluoroethanol

    ASJC Scopus subject areas

    • Biochemistry
    • Molecular Biology

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