Total, membrane, and immunogenic proteomes of macrophage- and tick cell-derived Ehrlichia chaffeensis evaluated by liquid chromatography-tandem mass spectrometry and MALDI-TOF methods.
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.
Seo
Gwi-Moon
GM
Department of Diagnostic Medicine/Pathobiology, College of Veterinary Medicine, Kansas State University, 1800 Denison Avenue, Manhattan, KS 66506, USA.
Cheng
Chuanmin
C
Tomich
John
J
Ganta
Roman R
RR
D000818:Animals; D001426:Bacterial Proteins / Q000032:analysis*; D015153:Blotting, Western; D002462:Cell Membrane / Q000737:chemistry; D002853:Chromatography, Liquid; D004285:Dogs; D018469:Ehrlichia chaffeensis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D005798:Genes, Bacterial; D008264:Macrophages / Q000382:microbiology*; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry; D017282:Tick-Borne Diseases / Q000235:genetics* / Q000378:metabolism; D013987:Ticks / Q000382:microbiology
D001426:Bacterial Proteins; D020543:Proteome