Detailed functional characterization of glycosylated and nonglycosylated variants of malaria vaccine candidatePfAMA1 produced inNicotiana benthamianaand analysis of growth inhibitory responses in rabbits

摘要: Summary One of the most promising malaria vaccine candidate antigens is Plasmodium falciparum apical membrane antigen 1 (PfAMA1). Several studies have shown that this blood-stage can induce strong parasite growth inhibitory antibody responses. PfAMA1 contains up to six recognition sites for N-linked glycosylation, a post-translational modification absent in P. falciparum. To prevent any potential negative impact N-glycosylation, been knocked out variants expressed eukaryotic hosts. However, glycosylation may increase efficacy by improving immunogenicity and/or focusing response towards relevant epitopes glycan masking. We describe production glycosylated and nonglycosylated Nicotiana benthamiana its detailed characterization terms yield, integrity protective efficacy. Both accumulated high levels (>510 μg/g fresh leaf weight) after transient expression, high-mannose-type N-glycans were confirmed variant. No significant differences between N. benthamiana Pichia pastoris detected conformation-sensitive ligand-binding studies. Specific titres >2 × 106 induced rabbits, reactivity with P. falciparum schizonts was observed immunofluorescence assays, as well 100% inhibition both variants, IC50 values ~35 μg/mL. Competition assays indicated number shielded from immune N-glycans, warranting further determine how be used directed targeting These results highlight plant expression systems platform candidates.