Phase transfer surfactant-aided trypsin digestion for membrane proteome analysis.

We have developed a new protocol for digesting hydrophobic proteins using trypsin with the aid of phase-transfer surfactants (PTS), such as sodium deoxycholate (SDC). SDC increases the solubility of hydrophobic proteins, enhances the activity of trypsin, and improves the accessibility to trypsin of proteins denatured during the extraction process. After digestion, SDC was successfully removed from the acidified solution containing tryptic peptides by adding a water-immiscible organic solvent, into which SDC was predominantly transferred, while the digested peptides remained in the aqueous phase. Compared with a protocol using an acid-labile surfactant, this PTS protocol increased the number of identified proteins and the recovery of hydrophobic peptides in the analysis of 400 ng of a membrane-enriched fraction of Escherichia coli. Application of the PTS protocol to 9.0 microg of a membrane-enriched pellet from human cervical cancer HeLa cells resulted in identification of a total of 1450 proteins, of which 764 (53%) were membrane proteins, by two-dimensional strong cation exchange (SCX)-C18 LC-MSMS with 5 SCX fractions. The distribution of the number of transmembrane domains in proteins identified in this study was in agreement with that in the IPI human database, suggesting that the PTS protocol can provide unbiased digestion of the membrane proteome.

[1]  P. Oeller,et al.  Global proteome discovery using an online three-dimensional LC-MS/MS. , 2005, Journal of proteome research.

[2]  J. Godovac-Zimmermann,et al.  Improvement of an in‐gel tryptic digestion method for matrix‐assisted laser desorption/ionization‐time of flight mass spectrometry peptide mapping by use of volatile solubilizing agents , 2001, Proteomics.

[3]  M. Goshe,et al.  Membrane proteomic analysis of Arabidopsis thaliana using alternative solubilization techniques. , 2007, Journal of proteome research.

[4]  Ronald J. Moore,et al.  Enrichment of integral membrane proteins for proteomic analysis using liquid chromatography-tandem mass spectrometry. , 2002, Journal of proteome research.

[5]  J. Yates,et al.  Proteomics of organelles and large cellular structures , 2005, Nature Reviews Molecular Cell Biology.

[6]  Oliver Mirus,et al.  Hiding behind Hydrophobicity , 2004, Journal of Biological Chemistry.

[7]  R. Mhatre,et al.  Strategies for locating disulfide bonds in a monoclonal antibody via mass spectrometry. , 1999, Rapid communications in mass spectrometry : RCM.

[8]  Ole Nørregaard Jensen,et al.  In situ liquid-liquid extraction as a sample preparation method for matrix-assisted laser desorption/ionization MS analysis of polypeptide mixtures. , 2003, Analytical chemistry.

[9]  Juri Rappsilber,et al.  Microcolumns with self-assembled particle frits for proteomics. , 2002, Journal of chromatography. A.

[10]  P. Andrews,et al.  Surfactant effects on protein structure examined by electrospray ionization mass spectrometry , 1994, Protein science : a publication of the Protein Society.

[11]  Thomas P Conrads,et al.  A detergent‐ and cyanogen bromide‐free method for integral membrane proteomics: Application to Halobacterium purple membranes and the human epidermal membrane proteome , 2004, Proteomics.

[12]  J. Langridge,et al.  Identification of proteins from two‐dimensional polyacrylamide gels using a novel acid‐labile surfactant , 2002, Proteomics.

[13]  Martin Gilar,et al.  Enzyme-friendly, mass spectrometry-compatible surfactant for in-solution enzymatic digestion of proteins. , 2003, Analytical chemistry.

[14]  M. Busman,et al.  Mass spectrometric analysis of integral membrane proteins: Application to complete mapping of bacteriorhodopsins and rhodopsin , 1998, Protein science : a publication of the Protein Society.

[15]  Wei Zhang,et al.  Proteomic analysis of integral plasma membrane proteins. , 2004, Analytical chemistry.

[16]  Yasushi Ishihama,et al.  Proteomic LC-MS systems using nanoscale liquid chromatography with tandem mass spectrometry. , 2005, Journal of chromatography. A.

[17]  A. Burlingame,et al.  Proteomic analysis of plasma membrane vesicles isolated from the rat renal cortex , 2005, Proteomics.

[18]  J. Yates,et al.  The application of mass spectrometry to membrane proteomics , 2003, Nature Biotechnology.

[19]  A. Krogh,et al.  Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. , 2001, Journal of molecular biology.

[20]  D. Russell,et al.  Proteolysis in mixed organic-aqueous solvent systems: applications for peptide mass mapping using mass spectrometry. , 2001, Analytical chemistry.

[21]  M. Mann,et al.  Organellar proteomics: turning inventories into insights , 2006, EMBO reports.

[22]  R. Doolittle,et al.  A simple method for displaying the hydropathic character of a protein. , 1982, Journal of molecular biology.

[23]  M. Mann,et al.  Stop and go extraction tips for matrix-assisted laser desorption/ionization, nanoelectrospray, and LC/MS sample pretreatment in proteomics. , 2003, Analytical chemistry.

[24]  G. Bensi,et al.  Characterization and identification of vaccine candidate proteins through analysis of the group A Streptococcus surface proteome , 2006, Nature Biotechnology.

[25]  M. Mann,et al.  Modular stop and go extraction tips with stacked disks for parallel and multidimensional Peptide fractionation in proteomics. , 2006, Journal of proteome research.

[26]  Nan Zhang,et al.  Effects of common surfactants on protein digestion and matrix-assisted laser desorption/ionization mass spectrometric analysis of the digested peptides using two-layer sample preparation. , 2004, Rapid communications in mass spectrometry : RCM.

[27]  F. McLafferty.  Tandem mass spectrometry. , 1981, Science.

[28]  Wei Zhang,et al.  Complete disulfide bond assignment of a recombinant immunoglobulin G4 monoclonal antibody. , 2002, Analytical biochemistry.

[29]  Zhen Liu,et al.  Evaluation of the application of sodium deoxycholate to proteomic analysis of rat hippocampal plasma membrane. , 2006, Journal of proteome research.

[30]  R. Caprioli,et al.  Mass spectrometry of intracellular and membrane proteins using cleavable detergents. , 2003, Analytical chemistry.

[31]  D. Richards,et al.  Development and applications of in-gel CNBr/tryptic digestion combined with mass spectrometry for the analysis of membrane proteins. , 2003, Journal of proteome research.

[32]  J. Yates,et al.  Large-scale analysis of the yeast proteome by multidimensional protein identification technology , 2001, Nature Biotechnology.

[33]  Y. Ishihama,et al.  Efficient in-gel digestion procedure using 5-cyclohexyl-1-pentyl-beta-D-maltoside as an additive for gel-based membrane proteomics. , 2004, Rapid communications in mass spectrometry : RCM.

[34]  A. Roda,et al.  Bile acid structure-activity relationship: evaluation of bile acid lipophilicity using 1-octanol/water partition coefficient and reverse phase HPLC. , 1990, Journal of lipid research.

[35]  Natalie G Ahn,et al.  Analysis of membrane proteins from human chronic myelogenous leukemia cells: comparison of extraction methods for multidimensional LC-MS/MS. , 2006, Journal of proteome research.

[36]  O. Jensen,et al.  Characterization of N-palmitoylated human growth hormone by in situ liquid-liquid extraction and MALDI tandem mass spectrometry. , 2007, Journal of mass spectrometry : JMS.

[37]  Y. Ishihama,et al.  Multiplexed two-dimensional liquid chromatography for MALDI and nanoelectrospray ionization mass spectrometry in proteomics. , 2006, Journal of proteome research.

[38]  John R Yates,et al.  Optimization of mass spectrometry-compatible surfactants for shotgun proteomics. , 2007, Journal of proteome research.

[39]  N. Kelleher,et al.  Processing complex mixtures of intact proteins for direct analysis by mass spectrometry. , 2002, Analytical chemistry.

[40]  M. Mann,et al.  Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips , 2007, Nature Protocols.

[41]  Richard D. Smith,et al.  Affinity labeling of highly hydrophobic integral membrane proteins for proteome-wide analysis. , 2003, Journal of proteome research.

[42]  Neil Hall,et al.  Analysis of the Plasmodium falciparum proteome by high-accuracy mass spectrometry , 2002, Nature.

[43]  D. Wishart,et al.  Comparison of SDS‐ and methanol‐assisted protein solubilization and digestion methods for Escherichia coli membrane proteome analysis by 2‐D LC‐MS/MS , 2007, Proteomics.