RESUMEN
Objective: The apical membrane antigen-1 [AMA-1] is considered as a promising candidate for development of a malaria vaccine against Plasmodium parasites. The correct conformation of this protein appears to be necessary for the stimulation of parasite-inhibitory responses, and these responses, in turn, seem to be antibody-mediated. Therefore, in the present investigation, we expressed the Plasmodium vivax AMA-1 [PvAMA-1] ectodomain in Escherichia coli [E. coli], purified it using standard procedures and characterized it to determine its biological activities for it to be used as a potential target for developing a protective and safe vivax malaria vaccine
Materials and Methods: In this experimental investigation, the ectodomain of PvAMA-1 antigen [GenBank accession no. JX624741] was expressed in the E. coli M15- pQE30 expression system and purified with immobilized-metal affinity chromatography. The correct conformation of the recombinant protein was evaluated by Western blotting and indirect immunofluorescence antibody [IFA] test. In addition, the immunogenic properties of PvAMA-1 were evaluated in BALB/c mice with the purified protein emulsified in Freund's adjuvant
Results: In the present study, the PvAMA-1 ectodomain was expressed at a high-level [65 mg/L] using a bacterial system. Reduced and non-reduced sodium dodecyl sulfate-polyacrylamide gel electrophoresis [SDS-PAGE] as well as Western blot analysis confirmed the appropriate conformation and folding of PvAMA-1. The evaluation of immunogenic properties of PvAMA-1 showed that both T helper-1 and 2 cells [Th1 and Th2] responses were present in mice after three immunizations and persisted up to one year after the first immunization. Moreover, the antibodies raised against the recombinant PvAMA-1 in injected mice could recognize the native protein localized on P. vivax parasites
Conclusion: We demonstrate that our recombinant protein had proper conformation and folding. Also, there were common epitopes in the recombinant forms corresponding to native proteins. These results; therefore, indicate that the expressed PvAMA-1 has the potential to be used as a vivax malaria vaccine
RESUMEN
Objective: Apical membrane antigen-1 [AMA-1] is one of the most promising bloodstage candidate antigens for production of a malaria vaccine against the Plasmodium parasite. Genetic diversity in protective antigens, which is a common phenomenon in a complex pathogen such as the Plasmodium parasite, is responsible for problems with the development of an effective malaria vaccine. This phenomenon will increase the parasite's ability to evade immune responses. Therefore, malaria vaccine development requires the evaluation of immune responses to different allelic forms of the vaccine candidate antigens
Methods: In this investigation, the two variant forms of PvAMA-1 [PvAMA-1A and B] were expressed in an Escherichia coli M15-pQE30 system using genomic DNA from Iranian individuals with patent Plasmodium vivax infection. The IgG responses of two antigens were evaluated in BALB/c mice with the purified protein emulsified in Freund's adjuvant. In addition, the correct conformation of the recombinant proteins was evaluated by the indirect immunofluorescence antibody test [IFA]
Results: The evaluation of immunogenic responses of two variant forms of PvAMA-1 showed the presence of IgG responses in mice after three immunizations. Cross-reactions were observed. Monitoring of IgG responses showed the persistence up to one year after the last immunization. The antibodies raised against recombinant PvAMA-1s in injected mice recognized the native protein [PvAMA-1] localized on Plasmodium vivax merozoites
Conclusion: The present outcomes confirmed the presence of common epitopes in recombinant forms of the protein that corresponded to native proteins. These emulsified proteins in Freund's adjuvant were immunogenic in BALB/c mice and IgG responses persisted for up to one year. The IgG responses to two PvAMA-1 variants did not differ significantly. The presence of cross-reactive antibodies has implied that one of these two forms of protein could be used in a universal blood-stage vaccine based on the PvAMA-1 antigen