Enrichment of Integral Membrane Proteins for Proteomic Analysis Using Liquid Chromatography−Tandem Mass Spectrometry

2002 ◽  
Vol 1 (4) ◽  
pp. 351-360 ◽  
Author(s):  
Josip Blonder ◽  
Michael B. Goshe ◽  
Ronald J. Moore ◽  
Ljiljana Pasa-Tolic ◽  
Christophe D. Masselon ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Membrane Proteins ◽  
Tandem Mass Spectrometry ◽  
Proteomic Analysis ◽  
Integral Membrane Proteins ◽  
Tandem Mass ◽  
Chromatography Tandem Mass Spectrometry ◽  
Liquid Chromatography Tandem Mass

scholarly journals Total, Membrane, and Immunogenic Proteomes of Macrophage- and Tick Cell-Derived Ehrlichia chaffeensis Evaluated by Liquid Chromatography-Tandem Mass Spectrometry and MALDI-TOF Methods

2008 ◽  
Vol 76 (11) ◽  
pp. 4823-4832 ◽  
Author(s):  
Gwi-Moon Seo ◽  
Chuanmin Cheng ◽  
John Tomich ◽  
Roman R. Ganta
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Membrane Proteins ◽  
Tandem Mass Spectrometry ◽  
Tandem Mass ◽  
Tick Cell ◽  
Chromatography Tandem Mass Spectrometry ◽  
Liquid Chromatography Tandem Mass ◽  
Immunogenic Proteins ◽  
Total Membrane

ABSTRACT Ehrlichia chaffeensis, a tick-transmitted rickettsial, is the causative agent of human monocytic ehrlichiosis. To examine protein expression patterns, we analyzed total, membrane, and immunogenic proteomes of E. chaffeensis originating from macrophage and tick cell cultures. Total proteins resolved by one-dimensional gel electrophoresis and subjected to liquid chromatography-electrospray ionization ion trap mass spectrometry allowed identification of 134 and 116 proteins from macrophage- and tick cell-derived E. chaffeensis, respectively. Because a majority of immunogenic proteins remained in the membrane fraction, individually picked total and immunogenic membrane proteins were also surveyed by liquid chromatography-tandem mass spectrometry and matrix-assisted laser desorption ionization-time of flight methods. The analysis aided the identification of 48 additional proteins. In all, 278 genes of the E. chaffeensis genome were verified as functional genes. They included genes for DNA and protein metabolism, energy metabolism and transport, membrane proteins, hypothetical proteins, and many novel proteins of unknown function. The data reported in this study suggest that the membrane of E. chaffeensis is very complex, having many expressed proteins. This study represents the first and the most comprehensive analysis of E. chaffeensis-expressed proteins. This also is the first study confirming the expression of nearly one-fourth of all predicted genes of the E. chaffeensis genome, validating that they are functionally active genes, and demonstrating that classic shotgun proteomic approaches are feasible for tick-transmitted intraphagosomal bacteria. The identity of novel expressed proteins reported in this study, including the large selection of membrane and immunogenic proteins, will be valuable in elucidating pathogenic mechanisms and developing effective prevention and control methods.

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