|Protein refolding techniques are essential for the |
manufacturing process of valuable recombinant proteins. Numerous
research have been done to increase the refolding yield, while we
are using ionic liquids as additives which are known to increase
the enzyme activity and stability and increase the
protein refolding yield.
|Since recombinant DNA technology was developed, it opens new era for the|
production of heterogeneous proteins in microbial hosts. The abundance of protein
expression systems renders the efficient bacterial production of most proteins
possible. But the expression of recombinant proteins in microorganism frequently
leads to the formation of insoluble aggregates, inclusion bodies (IBs). (Figure 1)
In order to get the water-soluble correctly folded active proteins, additional
protein refolding process are needed.
|Figure 1. Production of recombinant proteins and inclusion body|
|There are several works that we are doing now:|
|Size-Exclusiong Chromatography Refolding|
- Small molecules, such as dithiotheritol (DTT) and urea, enter the pores of the resin and are separated from the
unfolded protein molecules.
- As the chaotrope concentration surroudding the unfolded species is reduced, the protein begin to fold yielding
compact and partially folded protein that can enter the resin pores.
|Figure 2. Schematic of protein refolding within the SEC Media|
|Protein Refolding Using ILs|
About one third of all enzymes need a specific metal ion for the enzyme to be active. If a metal ion is the
cofactor which enhance the enzyme activity, the increased refolding yield through the stabilization of enzyme
active sites will be obtained by adding this metal ion during the protein refolding process.
|Figure 3. Laccase from Trametes versicolor
||Figure 4. Copper-based IL|
|Figure 5. Laccase refolding with copper-based IL|