Structural and biophysical characterization of a novel nematode lipid binding protein of Dioctophyme renale

SISTI, MARTÍN S.; A. NAHILI GIORELLO; JULIO J. CARAMELO; LUCRECIA M. CURTO; JOSÉ M. DELFINO; FRANCHINI, GISELA R.

Congreso; XLIX Reunión Anual de la Sociedad Argentina de Biofísica; 2021

Sociedad Argentina de Biofísica

The giant kidney worm, Dioctophyme renale, is a debilitating and potentially lethalparasite that typically inhabits and destroys, the host?s right kidney, and may also befound in ectopic sites. It is circumglobally distributed, mainly in wild carnivores that liveclose to rivers and is increasingly regarded as a threat to other domestic animals andhumans. Previous results from our laboratory identified two major proteins present in thepseudocelomic fluid (PF), one of 17 kDa (P17), that has a prosthetic heme group and ispossibly related to oxygen transport, and another of 44 kDa (P44), that binds lipids and issecreted. Both proteins would play a crucial role in the biochemistry of the parasitebecause lacking the metabolic pathways for the syntheses of the heme group, fatty acids,and cholesterol, this organism must obtain these compounds from its host. The aim of thiswork is to carry out a structural and biophysical characterization of P44, emphasizing itspossible biological functions. Therefore, the protein was purified from PF by SEC and thendelipidated by HPLC. Purity was checked by SDS-PAGE followed by Western blotting.Glycosylated residues were evidenced using a Schiff based dyeing and by an enzymaticdeglycosylation assay. P44 presents both far and near UV CD spectra consistent with awell folded protein. Additionally, denaturation curves were studied by fluorescence,showing high stability towards denaturing agents such urea and guanidinium chloride. Toassess function, fluorescence displacement experiments were carried out. Affinity for oleicacid (Kdapp 6.9 µM) and cholesterol (Kdapp 27.6 µM) was measured by their ability todisplace DAUDA or ANS probes, respectively. In summary, P44 emerges as a novelnematode lipid binding protein likely involved in the modulation of the host´s immuneresponse.