The complexity of the secreted NPA and FAR lipid-binding protein families of Haemonchus contortus revealed by an iterative proteomics-bioinformatics approach.
Two different classes of small nematode specific lipid-binding proteins, the nematode polyprotein allergens/antigens (NPAs) and the fatty acid- and retinol-binding (FAR) proteins, are secreted by helminth parasites. Until now, there was no evidence of the expression or secretion of these two families of proteins in Haemonchus contortus. In this study, we applied proteomic and bioinformatic tools in an iterative manner to reveal the expression and complexity of these proteins in the excretory/secretory products (ESP) of adult H. contortus at the protein and gene levels. Initial examination of the mass spectra of ESP fractions against standard databases returned nine peptides mapping to Ostertagia ostertagi NPA and FAR sequences. Searches of the H. contortus EST and genomic contig databases with the O. ostertagi and Caenorhabditis elegans homologues retrieved diverse sequences encoding H. contortus NPA and FAR proteins. H. contortus sequences were then integrated into a customized database and a new search of the mass spectra achieved a 10-fold improvement in coverage of the predicted H. contortus NPAs. The final analyses of the mass spectra achieved 49-60% coverage of H. contortus NPAs and 7-47% coverage of H. contortus FARs. Moreover, the diversity in structures of the encoding genes was revealed by assembling the genomic sequence data with predicted protein sequences confirmed by the peptide evidence. We predict there are at least one Hc-NPA gene and six Hc-FAR genes in H. contortus, and life stage gene expression of Hc-FAR-1 to -6 revealed unique transcription patterns for each of these genes.
Kuang
Lisha
L
CSIRO Livestock Industries, Queensland Biosciences Precinct, 306 Carmody Road, St. Lucia, 4067 QLD, Australia.
Colgrave
Michelle L
ML
Bagnall
Neil H
NH
Knox
Malcolm R
MR
Qian
Min
M
Wijffels
Gene
G
D000595:Amino Acid Sequence; D000818:Animals; D000947:Antigens, Helminth / Q000032:analysis*; D017173:Caenorhabditis elegans / Q000235:genetics; D029742:Caenorhabditis elegans Proteins / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics; D019295:Computational Biology / Q000379:methods; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics; D006189:Haemonchus / Q000737:chemistry*; D015801:Helminth Proteins / Q000032:analysis*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010028:Ostertagia / Q000235:genetics; D010802:Phylogeny; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D000947:Antigens, Helminth; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D018509:DNA, Helminth; C473216:FAR protein, C elegans; D015801:Helminth Proteins; D020543:Proteome