Evaluation of immune response to recombinant Bacillus anthracis LFD1PA4 chimeric protein

Background: Anthrax is a particularly dangerous infectious disease that affects humans and livestock. Efficacious vaccines that can rapidly induce a longterm immune response are required to prevent anthrax infection in humans. Domains 4 and 1 of the protective antigen (PA) and lethal factor (LF), respectively, have very high antigenic properties. Aims: In this experimental study, the pET28alfD1pa4 expression vector was designed, constructed and transferred into E. coli BL21 (DE3) plysS. Methods: For this purpose, pa4 gene was amplified by polymerase chain reaction (PCR) and cloned in a pGEM Teasy vector. The pGEMpa4 and pGEMlfD1 were digested by XbaI and HindIII enzymes. The ligation reaction was performed by ligase T4 enzyme and the gene cassette, lfD1pa4, was subcloned in pET28a and transferred to E. coli BL21 (DE3) PlysS. Expression and purification of chimeric proteins were confirmed by sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDSPAGE) and Western blotting techniques. The chimera LFD1PA4 and mixed LFD1+PA4 proteins were injected four times into mice and antibody production was relativity evaluated by enzymelinked immunosorbent assay (ELISA) test. Results: The results showed that both chimeric and mixed proteins are immunogenic, but LFD1PA4 has a higher potential to stimulate mice immune system. Conclusion: LFD1PA4 chimeric protein induced a higher immune response than LFD1+PA4 mixed protein and elicited antibody responses to LF and edema factor (EF), therefore, it holds promise to be a more effective trivalent vaccine candidate to use in anthrax prevention.