First evidence of a glutaredoxin-like protein in Entamoeba histolytica.

BIROCCO F; GUERRERO SA; IGLESIAS AA; ARIAS DG

Salta

Congreso; SAIB LV Reunión Anual; 2019

SAIB

Entamoeba histolytica, an intestinalparasitic protozoan, is the causative agent of amoebiasis. The parasite usuallylives and multiplies within the human gut, an environment of reduced oxygenpressure. During tissue invasion, E. histolytica is exposed toelevated amounts of exogenous reactive oxygen species (ROS), which are highlytoxic for the parasite. The metabolic pathway for ROS detoxification in thisorganism is a matter of controversy. Because neither glutathione nor itsassociated enzymes were found to occur, it has been proposed the cysteine as amain intracellular thiol and one of the compounds responsible for maintainingthe intracellular redox balance. In this work, we present the functional characterizationof a glutaredoxin-like protein from E. histolytica (EhGrx1). Biochemical assaysshowed that EhGrx1 was able to catalyse the in vitroreduction of GSH-derivate low molecular mass disulfides and cystine. The protein obtainedby recombinant expression in Escherichiacoli presented an apo-monomeric structure; however, a holo-protein form was obtained from supplemented culture media withferric citrate and cysteine. The ability to ligate iron-sulfur centers (ISCs)was evaluated by UV-Vis spectroscopy and gel filtration chromatography, showingevidence that EhGrx1 could bind ISCs. The Grx activity was not detectedin holo-EhGrx1, suggesting that its catalytic cysteine residue would belinked to ISC. We also evaluated by western blot the relative abundance of EhGrx1in E. histolytica cells exposed to exogenous oxidative species and metronidazole(the preferred drug for amoebiasis treatment). The results showed that theprotein level increases respect to no-treated cells. Similar behaviour wasobserved in the subcellular localization analysis, carried out for differentoxidative conditions byconfocalimmunofluorescence microscopy. Altogether, the results suggest that EhGrx1could be involved in oxidative and nitrosative stress protection in theparasite. To the best of our knowledge this is the first characterization of thistype of protein in E. histolytica.Grantedby ANPCyT (PICT2016-1778and PICT2017-2268).