Eimeria tenella: identification of secretory and surface proteins from expressed sequence tags.
To identify new vaccine candidates, Eimeria tenella expressed sequence tags (ESTs) from public databases were analysed for secretory molecules with an especially developed automated in silico strategy termed DNAsignalP. A total of 12,187 ESTs were clustered into 2881 contigs followed by a blastx search, which resulted in a significant number of E. tenella contigs with homologies to entries in public databases. Amino acid sequences of appropriate homologous proteins were analysed for the occurrence of an N-terminal signal sequence using the algorithm signalP. The resulting list of 84 entries comprised 51 contigs whose deduced proteins showed homologies to proteins of apicomplexan parasites. Based on function or localisation, we selected candidate proteins classified as (i) secreted proteins of Apicomplexa parasites, (ii) secreted enzymes, and (iii) transport and signalling proteins. To verify our strategy experimentally, we used a functional complementation system in yeast. For five selected candidate proteins we found that these were indeed secreted. Our approach thus represents an efficient method to identify secretory and surface proteins out of EST databases.
Klotz
Christian
C
Department of Molecular Parasitology, Humboldt University, Berlin, Germany.
Marhöfer
Richard J
RJ
Selzer
Paul M
PM
Lucius
Richard
R
Pogonka
Thomas
T
D000595:Amino Acid Sequence; D000818:Animals; D016782:Apicomplexa / Q000737:chemistry / Q000235:genetics; D016000:Cluster Analysis; D016384:Consensus Sequence; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D016786:Eimeria tenella / Q000737:chemistry* / Q000201:enzymology / Q000235:genetics; D020224:Expressed Sequence Tags; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D021382:Protein Sorting Signals / Q000235:genetics; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D016053:RNA, Protozoan / Q000235:genetics; D017385:Sequence Homology; D014162:Transfection; D043324:beta-Fructofuranosidase / Q000235:genetics / Q000378:metabolism
D018076:DNA, Complementary; D008565:Membrane Proteins; D021382:Protein Sorting Signals; D015800:Protozoan Proteins; D016053:RNA, Protozoan; D043324:beta-Fructofuranosidase