Adduci, I.; Sajovitz-Grohmann, F.; Wortha, L. N.; Dutkiewicz, Z.; Weidinger, H.; Rohrer, S. B.; Joachim, A.; Wittek, T.; Werling, D.; Wilson, I. B. H.; Lichtmannsperger, K.; Yan, S.

The H11 antigens, located on the intestinal microvilli of Haemonchus contortus, comprise a group of homologous aminopeptidases essential for the parasite\'s digestion of blood meals. Native H11 proteins are promising vaccine antigens, capable of eliciting robust protective immunity against H. contortus in sheep and goats. However, recombinant forms of H11, produced either in conventional expression systems or in transgenic Caenorhabditis elegans, failed to replicate the protective efficacy of the native form, most likely due to two critical factors: improper glycosylation and protein misfolding. To address these limitations, we developed a novel strategy to produce recombinant Haemonchus antigens in glycoengineered insect cells. By introducing three C. elegans genes that alter the native N-glycosylation pathways of Hi5 insect cells we successfully expressed soluble H11 and GA1 antigens featuring nematode-specific glycan epitopes, including tri-fucosylated structures and the Gal{beta}1,4Fuc motif. The glycoengineered H11 proteins retained aminopeptidase activity and stimulated cytokine secretion from ovine peripheral blood mononuclear cells in vitro. These findings establish a platform for producing bioactive vaccine antigens against the parasitic nematode H. contortus.