Defining the protein repertoire of microneme secretory organelles in the apicomplexan parasite Eimeria tenella.
The apicomplexan pathogen Eimeria causes coccidiosis, an intestinal disease of chickens, which has a major welfare and economic impact on the poultry industry. There is an urgent need to identify molecules that are rational targets for drug design and novel vaccines against coccidiosis. Apicomplexan secretory organelles, including micronemes and rhoptries, are essential for invasion of the host intestinal epithelium and establishment of parasitism. However, relatively little is known about the precise molecular function of these organelles, partly because few organelle proteins have been characterized. In this study, proteomics tools have been harnessed to define the protein repertoire of micronemes. Purified microneme proteins from Eimeria tenella sporozoites were excised from two-dimensional (2-D) gels and analyzed using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) and chemically assisted fragmentation (CAF)-MALDI with de novo sequencing. Peptide mass profiles were searched against the NCBI non-redundant (nr) database and against Eimeria-specific databases using the Mascot search algorithm, resulting in the identification of 37 of 96 spots excised from the 2-D gels. In addition, we have found CAF-MALDI to be a useful adjunct for identifying proteins, without the need for tandem MS. This global approach to protein characterization will be vital to gain greater understanding of the processes involved in apicomplexan host cell invasion.
Bromley
Elizabeth
E
Institute for Animal Health, Compton, Berkshire RG20 7NN, UK. liz.bromley@bbsrc.ac.uk
Leeds
Nicola
N
Clark
Julie
J
McGregor
Emma
E
Ward
Malcolm
M
Dunn
Michael J
MJ
Tomley
Fiona
F
D000818:Animals; D016786:Eimeria tenella / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D008854:Microscopy, Electron; D015388:Organelles / Q000378:metabolism* / Q000648:ultrastructure; D020543:Proteome; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D020543:Proteome