BIOCHEMICAL CHARACTERIZATION OF Yersinia OUTER PROTEIN P (YopP)-GALECTIN-1 INTERACTIONS

RABINOVICH GA; GOMEZ BARROSO JA; DI GENARO MS; ELIÇAVE J; BRENDA JOFRE; MARIÑO K; DAVICINO RC

Merlo

Congreso; XXXV Reunión Científica Anual de la Sociedad de Biología de Cuyo; 2017

Sociedad de Biología de Cuyo

Yersinia enterocolitica (Ye) is a Gram-negative enteropathogenic bacterium. Yersinia outer proteins (Yops) are effector proteins of Ye that are injected into host cells. YopP causes suppression of pro-inflammatory cytokines and induces apoptosis in both macrophages and dendritic cells. Galectin-1 (Gal-1) is a ?proto-type? β-galactoside-binding lectin widely distributed in host tissues with important immunomodulatory roles. We previously demonstrated that the Ye-induced apoptosis of macrophages depends on both YopP and Gal-1, and that Gal-1 binds only to YopP preventing its auto-degradation. The aim of this study was to characterize biochemically the YopP-Gal-1 binding. Glycosylation of YopP was confirmed by Schiff staining and lectin blots with both Gal-1 and other biotinylated plant lectins. Different glycosylation patterns were observed for the Yops. Only YopP presented permissive glycoepitopes for Gal-1 binding. Moreover, we isolated the Gal-1-YopP complex using sodium alginate beads. The presence of Gal-1 and YopP in the complex was confirmed by Western blot using anti-Gal-1 or anti-YopP antibodies. The pure complex was subjected to reducing or non-reducing conditions, and then analyzed on partially denaturingpolyacrylamide gels to evaluate possible different conformations. Additionally, polyacrylamide gels using transverse urea gradient, evidenced strong interactions only inhibited by 2-4 M urea. This interaction was not affected by the oxidation of vicinal diols. We conclude that the specific binding of Gal-1 to YopP under in vitro conditions is protein-glycan type.