Diversity in the structures and ligand binding sites of nematode fatty acid and retinol binding proteins revealed by Na-FAR-1 from Necator americanus

Fatty acid and retinol binding proteins (FARs) comprise a family of unusual α-helix rich lipid binding proteins found exclusively in nematodes. They are secreted into host tissues by parasites of plants, animals and humans. The structure of a FAR protein from the free-living nematode Caenorhabditis elegans is available, but this protein (Ce-FAR-7) is from a subfamily of FARs that does not appear to be important at the host-parasite interface. We have therefore examined Na-FAR-1 from the blood-feeding intestinal parasite of humans, Necator americanus . The three dimensional structure of Na-FAR-1 in its ligand-free and ligand-bound forms, determined by nuclear magnetic resonance spectroscopy (NMR) and X-ray crystallography, respectively, reveals an a-helical fold similar to Ce-FAR-7, but Na-FAR-1 possesses a larger and more complex internal ligand binding cavity and an additional C-terminal a-helix. Titration of apo -Na-FAR-1 with oleic acid, analysed by NMR chemical shift perturbation, reveals that at least four distinct protein:ligand complexes can be formed. Na-FAR-1, and possibly other FARs, may have a wider repertoire for hydrophobic ligand binding, as confirmed here by our finding that a range of neutral and polar lipids co-purify with the bacterial recombinant protein. Finally, we show by immunohistochemistry that Na-FAR-1 is present in adult worms with a tissue distribution indicative of possible roles in nutrient acquisition by the parasite and in reproduction in the male