Last but not least: characterization of Ce-FAR-8 from Caenorhabditis elegans, a potential contributor in the evolution of nematode parasitism.

LUCIANA RODRIGUEZ SAWICKI; CAMILA A. REYES; JOSE F. LOMBARDO; GISELA R. FRANCHINI

Rosario

Congreso; L Reunión Anual de la Sociedad Argentina de Biofísica; 2022

Sociedad Argentina de Biofísica

Fatty Acid and Retinol Binding Proteins (FAR) are found exclusively in nematodes and are conserved in both free-living and parasitic species where they are mainly present in the excretory/secretory products. For parasitic nematodes these proteins are hypothesized to participate in the acquisition of nutrients from their host and/or to manipulate host defense mechanisms. Eight FAR genes are present in the model organism Caenorhabditis elegans (Ce). Ce-FAR proteins 1 to 7 have been recombinantly produced and characterized while Ce-FAR-8 was left unattended as being too divergent. Notably, three Ce-FAR-8 homologues were found in Caenorhabditis bovis, a particularly unusual species within the genus regarding its niche. This species is found in the inflamed ears of Zebu cattle in Eastern Africa where it is believed to cause bovine parasitic otitis. C. bovis is therefore of particular interest to researchers interested in the evolution of nematode parasitism. Our hypothesis is that Ce-FAR-8 is closely related to FAR nematode parasitic proteins. The structures of Ce-FAR-8 and the three C. bovis homologues were calculated using Alphafold2 and structural alignments between the four proteins were performed. We could observe the conservation of structure and folding patterns, even in two of C. bovis homologues proteins considerably shorter than Ce-FAR-8. The structural alignment of Ce-FAR-8 with apo-Ce-FAR-7 exhibits a high RMSD (10 A), making it difficultto compare the structural features of each protein. On the other hand, in the structural alignment of Ce-FAR-8 with holo-NaFAR1 from a parasitic nematode, we observed that the cavity of Ce-FAR-8 is bigger and a-helixes 7 and 8 from Ce-FAR-8 participate in its expansion, while a-helix 7 in NaFAR1 is responsible of the narrowing. Additionally, we obtained the first recombinant Ce-FAR-8, aiming to characterize its structure and lipid interaction. The cDNA of Ce-FAR-8 was cloned from the mARN of C. elegans, expressed as a recombinant protein and purified. Using fatty acid fluorescent analogues such as DAUDA we demonstrated that Ce-FAR-8 is indeed a lipid binding protein.