Biochemical and immunological characterization of a recombinant Mu-class glutathione S-transferase from Echinococcus granulosus.

Echinococcus granulosus poses a major threat to human and animal health. Mu-class glutathione S-transferases (GSTs) are important detoxification enzymes in helminths and may participate in parasite-host interactions. In our previous proteomic analysis of extracellular vesicle (EV) preparations, E. granulosus Mu-class GST (Eg.GST-Mu) was identified in preparations derived from plasma collected from mice at 20 weeks post-infection and from hydatid cyst fluid. In the present study, recombinant Eg.GST-Mu (rEg.GST-Mu) was expressed and biochemically characterized, and the immunological responses associated with rEg.GST-Mu immunization in mice were preliminarily evaluated. The Eg.GST-Mu gene was cloned into the pET28a(+) vector and expressed in Escherichia coli BL21(DE3). The purified recombinant protein was verified using an anti-His antibody and rEg.GST-Mu-specific antiserum and retained detectable GST catalytic activity after refolding. Male C57BL/6J mice were subsequently immunized with rEg.GST-Mu, and immune responses were evaluated by measuring antigen-specific IgG and its subclasses IgG1 and IgG2c, together with flow-cytometric analysis of splenic CD4⁺ T-cell subsets. rEg.GST-Mu immunization elicited a robust antigen-specific humoral response and was associated with changes in the distribution of splenic CD4⁺ T-cell subsets and the Treg/Th17 ratio. In vitro, rEg.GST-Mu attenuated the proliferative expansion of activated CD4⁺ T cells. These findings demonstrate that rEg.GST-Mu possesses detectable GST catalytic activity and immunogenicity and provide an experimental basis for further investigation of its immunological functions. However, the role of native Eg.GST-Mu during natural E. granulosus infection, its potential association with EVs, and the mechanisms underlying its effects on host immune responses remain to be directly established.