An abundant, trans-spliced mRNA from Toxocara canis infective larvae encodes a 26-kDa protein with homology to phosphatidylethanolamine-binding proteins.
A full-length mRNA encoding a secreted 26-kDa antigen of infective larvae of the ascarid nematode parasite Toxocara canis has been identified. This was characterized as a 1,082-base pair clone highly abundant (0.8-1.9%) in cDNA prepared from infective stage larvae but absent from cDNA from adult male worms. Sequence analysis revealed an open reading frame corresponding to a hydrophilic 263-amino acid residue polypeptide with a 20-residue N-terminal signal peptide, indicating that it is secreted. The 5' end of the cDNA was isolated by polymerase chain reaction using a primer containing the nematode-spliced leader sequence, SL1, showing that the mRNA is trans-spliced. The molecular mass of the putative protein with the signal peptide removed is 26.01 kDa, and antibody to the recombinant protein expressed in bacterial vectors reacts with a similarly sized protein in T. canis excretory/secretory (TES) products. An identical sequence was obtained from a genomic clone isolated by expression screening with mouse antibody to TES. The 72 amino acid residues adjacent to the signal peptide form two homologous 36-residue motifs containing 6 cysteine residues; this motif is found also in the T. canis-secreted glycoprotein TES-120 and in genes of Caenorhabditis elegans. Sequence data base searches revealed significant similarity to 7 other sequences in a newly recognized gene family of phosphatidylethanolamine-binding proteins that includes yeast, Drosophila, rat, bovine, simian, and human genes and a representative from the filarial nematode Onchocerca volvulus. Assays with the T. canis recombinant 26-kDa protein expressed as a fusion with maltose-binding protein have confirmed phosphatidylethanolamine-binding specificity for this novel product.
Gems
D
D
Wellcome Research Centre for Parasitic Infections, Department of Biology, Imperial College of Science, Technology and Medicine, London, United Kingdom.
Ferguson
C J
CJ
Robertson
B D
BD
Nieves
R
R
Page
A P
AP
Blaxter
M L
ML
Maizels
R M
RM
D000595:Amino Acid Sequence; D000818:Animals; D000947:Antigens, Helminth / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D015698:Genomic Library; D015801:Helminth Proteins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D007814:Larva; D008297:Male; D008969:Molecular Sequence Data; D010714:Phosphatidylethanolamines / Q000378:metabolism; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics*; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D017166:Toxocara canis / Q000235:genetics* / Q000254:growth & development
D000947:Antigens, Helminth; D002352:Carrier Proteins; D018076:DNA, Complementary; D015801:Helminth Proteins; D010714:Phosphatidylethanolamines; D012333:RNA, Messenger; D011994:Recombinant Proteins; C045159:TES-26 protein, Toxocara canis