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Differential recovery of membrane proteins after extraction by aqueous methanol and trifluoroethanol
Author(s) -
Zhang Huoming,
Lin Qingsong,
Ponnusamy Sukumar,
Kothandaraman Narasimhan,
Lim Teck Kwang,
Zhao Changqing,
Kit Hon Sook,
Arijit Biswas,
Rauff Mary,
Hew ChoyLeong,
Chung Maxey Ching Ming,
Joshi Shashikant B.,
Choolani Mahesh
Publication year - 2007
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200600579
Subject(s) - chemistry , chaotropic agent , membrane , chromatography , proteome , urea , membrane protein , methanol , shotgun proteomics , aqueous solution , extraction (chemistry) , biochemistry , proteomics , organic chemistry , gene
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.