Team:Korea U Seoul/Project/sub application

Enzyme purification is costly, and purified enzymes cannot be used multiple times due to decrease in activity after few reactions. To overcome this problem, ‘whole-cell biocatalysis’, which uses the microorganism itself as a catalyst, was introduced. Related to this, ‘Cell surface display’ can provide higher rate of access to the substrate, but due to the possibility of toxicity, its result has a low rate of actual expression rate. To make more efficient and stable surface display system,“sortase-based surface display at gram-positive bacteria” has been researched in many fields lately. Sortase functions as ‘protein ligase’ that recognizes specific amino acids sequence(LPXT) of each peptides and links them together. It is also associated with cell wall-bound protein and pili synthesis. ‘Sortase-based surface display’ can induce stable enzyme surface display by reducing cell toxicity and holding enzymes tightly to the cell wall with covalent bond.

However, it cannot overcome low display yield and it should include an extra step of treating sortase in vitro. Therefore, referring to sortase-based study, we are going to make enzyme polymer (enzyme pili) on cell wall by modifying pili units used in pili synthesis in other species in Corenebacterium genus. This study provides more efficient way of enzymatic process and application of pili biosynthesis

Our protein whip platform that is sortase-based surface modified pili display is worthy of note in that many types of enzymes can be applied to our platform for enzymatic pili display. That is, with just a simple specific amino acid sequence, any enzymes can be made into pili form, resulting in mass production of enzymatic product, with constant supply of its substrate. For example, substitution of SpaA with agarose would result in agarose degrading bacteria, which can be economically valuable when commercialized. Also, with poly-His tag or cysteine, we can build up heavy metal binding bacteria.

Our study challenges the typical synthetic biology by using novel gram positive bacteria, C. glutamicum, to develop our protein whip platform, sortase based surface display. This broadens the usage of model organism in synthetic biology, which could contribute to the development of synthetic biology. We expect optimized results for enzymatic process by using C. glutamicum that is confirmed to be suitable for industrial mass production.