Expression of Omp16 and L7/L12 <em>Brucella</em> <em>abortus</em> protective antigens as secretory fusion proteins in mammalian cells

Abstract

<em>Brucella abortus</em> is a facultative intracellular bacterial pathogen infecting animals and humans. This preliminary study was designed to express two immunogenic genes of <em>Brucella abortus</em> as recombinant fusion proteins in mammalian cells for being studied as a vaccine candidate in mammalian hosts, especially mice and cattle, in our next study. The complete open reading frame sequences of two immune dominant genes of <em>Brucella abortus</em> namely, Omp16 and L7/L12 were fused with an intermediate spacer along with N-terminal fusion with secretory signal sequence from immunoglobulin M. The complete fusion gene sequence was codon optimized for expression in mammalian cells. For expression analysis, the codon optimized synthetic gene was cloned in pDsRed-Express-N1 mammalian expression vector, with C-terminal fusion with red fluorescent protein sequence. On transfection in MDBK and HEK-293 cells, appearance of red fluorescence in transfected cells indicated expression of Omp16- L7/L12 fusion proteins along with RFP. The Omp16-L7/L12 fusion construct without RFP sequence was also expressed in mammalian cells. The expressed Omp16-L7/L12 fusion proteins were confirmed through both indirect fluorescence antibody test and western blot. This preliminary study suggested that the codon optimized Omp16-L7/L12 fusion construct is ready to be studied in hosts like mice and cattle for its vaccine efficacy.