Abstract

Alveolar echinococcosis (AE), caused by the metacestode larval of Echinococcus multilocularis , is one of the most lethal helminthic diseases in humans. Current treatment options, such as albendazole, are limited in their efficacy, highlighting the need for a deeper understanding of the parasite-host interaction to identify new therapeutic targets. One promising area of research involves helminth-derived cystatins, which are known to modulate host immune responses to facilitate parasite survival. A cystatin homologue from E. multilocularis ( Em Cystatin-B) was identified and analyzed. Em Cystatin-B was cloned, expressed and purified. Its expression patterns were evaluated by western blot, qPCR and Immunohistochemistry The Em Cystatin-B structure was solved by X-ray crystallography. Em Cystatin-B was expressed in the mature protoscoleces as well as in the cytosol and nucleus of the metacestode vesicles. Moverover, Em Cystatin-B adopts a conserved typical cystatin fold, but also exhibits unique structural features. Notably, a novel feature characterized by two intermolecular disulfide bridges between Cys4 in a Em Cystatin-B molecular and Cys76 in adjacent molecule was discovered. Further investigation demonstrated this distinctive feature appears to be involved in the oligomerization of Em Cystatin-B, facilitating a monomer-dimer-tetramer assembly pathway. The crystal structure of Em Cystatin-B reveals a novel feature in classical stefins, and provides species-specific insights into the sequence, structure, and functional characteristics of Em Cystatin-B.