Application of poly-arginine fused minichaperone to renaturation of cyclodextrin glycosyltransferase expressed in recombinant Escherichia coli

Minichaperone, a monomeric polypeptide binding domain fragment of GroEL, has chaperone activities in vivo and in vitro. Fed-batch cultures to optimize production of poly-arginine fused minichaperone (MiniELR10) in recombinant Escherichia coli were done to produce dry cell weight of 75.2 g/L, MiniELR10 expression level of 30.7%, and MiniELR10 concentration of 11.1 g/L in yeast extract supplemented conditions. MiniELR10 was simply purified through a single step of cation exchange chromatography with high purity. In a prewashing step and an adsorption process by controlling NaCl concentration, removal of contaminant intracellular E. coli proteins and selective binding of MiniELR10 on a cation exchanger were achieved without loss of the target protein. The purified MiniELR10 was efficiently immobilized on the cation exchanger and applied to the refolding of Bacillus macerans cyclodextrin glycosyltransferase (CGTase), which was expressed as inclusion body in recombinant E. coli. The refolding yield of CGTase was enhanced by 71% by the immobilized MiniELR10 compared with a conventional refolding method. Thus, minichaperone immobilized by poly-arginine fusion could assist refolding of heterologous proteins expressed as inclusion body.