Analysis of heterologous expression and purification of Plasmodium falciparum Generative Cell Specific1 as a transmission blocking vaccine

Background: Malaria is the most infectious and deadly disease in human that is caused by an obligatory intracellular parasite belong to Plasmodium genus. The current tools to control the malaria are not sufficient and to achieve the elimination and eradication of malaria, new tools such as effective malaria vaccine is necessary. Transmission blocking vaccines (TBVs) are the most effective vaccines for this purpose. Generative cell specific1 (GCS1) is an ideal TBV candidate antigen that is expressed on the surface of male gamete and antibodies raised to this antigen could prevent of gamete fertilization and zygote formation.Objectives: expression and purification of Plasmodium falciparum Generative Cell Specific1 as a transmission blocking vaccineMaterial and Methods: In this study, gcs1 gene was codon optimized according to E. coli usage codons, synthesized, and cloned in pET23a plasmid. The recombinant plasmid was transformed to different expression hosts (E. coli BL21 DE3, E. coli BL21 DE3 pLYSs, E. coli Rosetta DE3) to get the best expression. The expressed PfGCS1 was purified by metal affinity chromatography and analyzed by western blotting using anti-His antibodies.Results: The recombinant PfGCS1-pET23a was confirmed by restriction enzyme analysis, and sequence analysis of recombinant plasmid showed no mutation, insertion or deletion in the cloned gene. The recombinant PfGCS1 was successfully expressed in E. coli Rosetta (DE3) with a His-tag fusion in C-terminal. Analysis of the purified rPfGCS1 showed a 35kDa protein without any non-specific bands. This purified protein was recognized by anti-His antibodies.Conclusion: In the next phase of study, polyclonal antibodies raised to this antigen could be evaluated to inhibit the sexual stage development of P. falciparum in Anopheles.

Keywords:

Malaria , Transmission blocking vaccine , Generative Cell specific 1