PlpE Epitopes of Fusion Protein as Novel Subunit Vaccine Candidates.

BACKGROUND

is the causative agent of many diseases. Antimicrobial treatment disadvantages highlight the need to find other possible ways such as prophylaxis to manage infections. Current vaccines against this agent include inactivated bacteria, live-attenuated bacteria, and nonpathogenic bacteria, which have disadvantages such as lack of immunogenicity, reactogenicity, or reversion to virulence wild bacteria. Using bioinformatical approaches, potentially immunogenic and protective epitopes identified and merged to design the best epitope fusion form in case of immunogenicity as a vaccine candidate.

MATERIALS AND METHODS

In this study, the fusion protein () and full genes () were cloned in pET28a in BL21 (DE3) firstly and later in pBAD/gIII A and expressed in Top10 coli. Overlap polymerase chain reaction (PCR) using different primers for 5' and 3' end of each segment produced fusion segment 1 + 2 and (1 + 2) +3 fragments and was used for cloning.

RESULTS

Cloning of both and into the pET28a vector and their transform into the BL21 (DE3) host was successful, as the presence of the cassettes was proved by digestion and colony PCR, however, their expression faced some challenges independent of expression inducer (isopropyl β-d-1-thiogalactopyranoside) concentration.

CONCLUSION

Changing the vector to pBAD/gIII A and consequently changing the host to Top10 have resulted in sufficient expression, which shows that Top10 may be a good substitute for such cases. Furthermore, it is concluded that adding 8M urea results in sufficient purification, which hypothesizes that denature purification is better for such cases than native one. Purified proteins headed for further analysis as vaccine candidates.