Ac-FAR-1, a 20 kDa fatty acid- and retinol-binding protein secreted by adult Ancylostoma caninum hookworms: gene transcription pattern, ligand binding properties and structural characterisation.
Antibody against adult Ancylostoma caninum excretory-secretory (ES) products was used to immunoscreen a cDNA expression library leading to the isolation of cDNAs encoding putative hookworm fatty-acid and retinol-binding proteins. Ac-far-1 and Ac-far-2 cDNAs encode open reading frames corresponding to approximately 20kDa proteins with 91 percent amino acid identity. Ac-FAR-1 and Ac-FAR-2 exhibit clear similarities to other FARs of parasitic nematodes, most closely to two of the FAR proteins of Caenorhabditis elegans (Ce-FAR-1 and Ce-FAR-2). By reverse transcriptase polymerase chain reaction (RT-PCR) assay, Ac-far-1 mRNA was detected in both adult and third-stage larvae of A. caninum. However, the respective proteins were detectable by immunoblot only in adult hookworm ES products and adult extracts. Using fluorescence-based binding assays, bacterial recombinant Ac-FAR-1 was found to bind fatty acids and retinol (Vitamin A) with dissociation constants in the micromolar region. Circular dichroism spectra indicated that Ac-FAR-1 possesses a high level of alpha-helix, similar to Ov-FAR-1 from Onchocerca volvulus. This is the first demonstration of a functional FAR secreted by adult hookworms and provides further evidence that FAR proteins secreted by parasitic nematodes are crucial to parasitism.
Basavaraju
Sridhar V
SV
Department of Microbiology and Tropical Medicine, The George Washington University and Sabin Vaccine Institute, DC 20037, USA.
Basavaraju
Sridhar
S
Zhan
Bin
B
Kennedy
Malcolm W
MW
Liu
Yueyuan
Y
Hawdon
John
J
Hotez
Peter J
PJ
D000595:Amino Acid Sequence; D000722:Ancylostoma / Q000235:genetics / Q000378:metabolism*; D000724:Ancylostomiasis / Q000469:parasitology; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004285:Dogs; D005227:Fatty Acids / Q000378:metabolism*; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008024:Ligands; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000032:analysis; D012177:Retinol-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014158:Transcription, Genetic
D002352:Carrier Proteins; D005227:Fatty Acids; D015801:Helminth Proteins; D008024:Ligands; D011994:Recombinant Proteins; D012177:Retinol-Binding Proteins