Surface display of recombinant protein on the cell surface of Bacillus subtilis by the CotB anchor protein

We developed a novel surface display system based on the CotB anchoring motif in order to express foreign protein on the surface of vegetative Bacillus subtilis cells. CotB is a protein in the B. subtilis spore coat. In this system, three repeats of the immunodominant ovalbumin T-cell epitope (OVA(323–339)) were linked with the cholera toxin B subunit (CTB) to construct a fusion protein, CTB-OVA epi, which was then fused to the C-terminal of the CotB protein so that CTB-OVA epi was expressed in vegetatively-growing B. subtilis. The expression and localization of the CTB-OVA epi protein was confirmed by western blotting, immunofluorescence microscopy, and flow cytometry. The results indicated that a CotB-based surface display system was successfully used to express the CTB-OVA epi protein on the surface of vegetative B. subtilis cells.