Novel Bacterial Surface Display System Based on the Protein MipA.

Bacterial surface display systems have been developed for various applications in biotechnology and industry. Particularly, the discovery and design of anchoring motifs is highly important for the successful display of a target protein or peptide on the surface of bacteria. In this study, an efficient display system on was developed using novel anchoring motifs designed from the mipA gene. Using the C-terminal fusion system of an industrial enzyme, lipase, six possible fusion sites, V, V, K, V, V, and K, which were truncated from the C-terminal end of the gene (MV, MV, MV, MV, MV, and MV) were examined. The whole-cell lipase activities showed that MV was the best among the six anchoring motifs. Furthermore, the lipase activity obtained using MV as the anchoring motif was approximately 20-fold higher than that of the previous anchoring motifs FadL and OprF but slightly higher than that of YiaTR232. Western blotting and confocal microscopy further confirmed the localization of the fusion lipase displayed on the surface using the truncated MV. Additionally the MV motif could be used for successfully displaying another industrial enzyme, α-amylase from . These results showed that the fusion proteins using the MV motif had notably high enzyme activities and did not exert any adverse effects on either cell growth or outer membrane integrity. Thus, this study shows that MipA can be used as a novel anchoring motif for more efficient bacterial surface display in the biotechnological and industrial fields.