Engineering, Cloning and Expression of DNA Sequence Coding of OMP31 Epitope of <i>Brucella melitensis</i> linked to IL-2 in <i>Escherichia coli</i>

Abstract

Brucellosis is a zoonetic disease that is caused by Brucella melitensis, which affects human health and animal husbandry industry. Omp31 is the most exposed outer membrane proteins (OMPs) and is a dominant antigen in smooth strains of B. melitensis, thus it is considered a desirable protein in subunit vaccine against brucellosis disease. Besides, IL-2 acts as a growth and differentiation stimulator of T-cells in cellular immune responses, therefore, it can serve as an adjuvant in subunit vaccines against intracellular pathogens. It seems that chimera scaffold, consisting of some copies of OMP31 experimental epitope region and IL-2, can provide a good immunization. In this regard, these tow fragments are linked together through using rigid linker and splicing overlap extension (SOE) PCR technique. Successful PCR products were TA cloned and then sub-cloned into pET-22b (+) vector as an expression vector. Recombinant protein was expressed in Escherichia coli BL21 (DE3) as an expressing lineage bacterium and then confirmed by SDS-PAGE and Western-blotting analysis. Secondary and tertiary structure prediction of recombinant protein and its immunity characterization were evaluated using in silico process. Further studies on the immunogenicity properties of this protein are expected to be carried out in the future to confirm the results in vivo.