*4.5. Bio-Surface Display of Enzymes*

Bacteriophages, spores, yeast, and bacterial cells can be used as enzyme carriers in socalled surface protein display systems [186,187]. *E. coli* bacterium is one of the most studied cells for enzyme surface display. Surface display of the targeted enzyme (the passenger) is realized by genetically fusing it to a carrier protein, which facilitates export across the cell envelope (inner membrane and outer membrane separated by the periplasm). The protein of interest is ultimately secreted and immobilized on the cell wall. Interestingly, enzymes are thus simultaneously produced and immobilized on the bio-component. The anchoring motif can be fused to the N- or C- end of the protein, or the protein can be inserted within the sequence of the anchor protein. The anchor protein must be able to ensure transport of the heterologous protein through the secretory pathway, proper folding of the fusion protein, and strong binding at the cell wall surface. Contrary to purified immobilized proteins, expressing enzymes on cell surfaces is considered advantageous as enzymes can be used repeated for extended periods with no loss of enzyme activity. Stability of immobilized proteins by such a natural procedure against organic solvents, extreme pH, or high temperature has been reported [188,189]. Yeast and spores are also currently explored as enzyme display surfaces allowing enzyme enhanced thermal stability [190,191]. However, the efficiency of the display systems in terms of immobilized enzyme amount and specific activity needs to be improved for envisioning any biotechnological application [192].
