Applications of Cell Surface Engineering in Bacteria

Cell surface engineering in bacteria is a principal technology for cell surface protein engineering by modifying the surface of a bacterial spore. Most bacteria have abundant cell surface-anchoring proteins that can be easily modified. These anchoring proteins are possible ways to display large numbers of recombinant protein molecules on the cell surface of many bacteria. The first study of a bacterial cell surface display system was reported in 1968 with a 32-amino acid residue outer membrane protein A (OmpA) of Escherichia coli K12 strain. So far, a number of bacterial cell surface display systems have been investigated to achieve attachment of heterologous proteins to the surface of different host cells including Gram-negative and Gram-positive bacteria.

Principle of Cell Surface Engineering in Bacteria

• coli is the most widely used host because there are well-established gene modification tools. Systems displaying passengers at a high density on the outer membrane (OM) via major carrier proteins have been established in E. coli. The carrier protein, which has an anchor domain, mediates the traverse of passengers through the inner membrane into the periplasm, directs them to the OM, and finally secretes them out of cells. The molecular size limitation of passenger proteins, fusion patterns, and localization patterns at the cell surface depends on the properties of the carrier protein. Therefore, the carrier protein involved is a main actor in the surface display system and directly affects the secretion efficiency and folding state of target passengers.

Applications of Surface Engineered Bacteria

• Medical application: vaccine delivery

Scientists have successfully engineered E. coli as a vaccine model against Foot-and-mouth disease. They fused several different Foot and Mouth Disease Virus (FMDV) epitopes containing the immunogenic regions of VP1 to the E. coli ompA. The immunogenicity of these recombinant bacteria was tested by immunizing the mice. The results showed extra stimulation in the immune system of the mice with the daily feed of these engineered bacteria.

Moreover, different antigens are expressed on the Bacillus subtilis spore surface by using different spore coat proteins as a carrier. In one study, the urease subunit (UreA) of animal pathogen Helicobater acinonychis was expressed on the B. subtilis spore surface using three different spore coat as carriers. This subunit is recognized as a major antigen of H. Pylori and induces protection against infection, and this system can be used for vaccine delivery.

Fig.1 Spore layers and surface display of β-galactosidase on B. subtilis spore. (Tafakori, 2012)

• Environmental application: whole-cell biocatalysts

Whole-cell biocatalysts displaying a single enzyme can catalyze only simple, one-step reactions, while the surface display of multiple enzymes enables the development of novel cells that can catalyze a set of multiple reactions at one time. Bioremediation is an application for multiple enzyme display. Organophosphorus pesticides (OPs) are highly toxic pollutants in soil and water and are common targets for bioremediation. Therefore, intensive efforts have been made in several studies to develop biocatalysts that can efficiently degrade OPs. Scientists have developed a whole-cell catalyst that co-displays two enzymes for OP degradation and its monitoring; OP hydrolase (OPH) and a methyl parathion hydrolase (MPH)-GFP fusion were co-displayed via ice nucleation proteins (INPs) and Lpp-OmpA, respectively. Degradation of OPs by OPH/MPH generates protons to influence the fluorescence of GFP through an environmental pH change, and thus can be monitored by detecting the fluorescence change.

Table 1. Development of whole-cell biocatalysts by cell surface display on bacteria. (Nakatani, 2017)

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References

1. Nakatani, H.; Hori, K. Cell surface protein engineering for high-performance whole-cell catalysts. Frontiers of Chemical Science and Engineering. 2017, 11(1):46-57.
2. Tafakori, V.; et al. Microbial cell surface display; its medical and environmental applications. Iranian Journal of Biotechnology. 2012, 10(4):231-9.