Solid-phase refolding of inclusion body protein in a packed and expanded bed adsorption chromatography

Abstract

Lipoprotein kringle “LK68” is a polypeptide of a modified angiostatin consisting of three kringle structures that might be clinically useful as potential cancer therapeutics. It can be produced by overexpressing it as an inclusion body in recombinant Escherichia coli. In this study, solid-phase refolding processes using a packed bed (PBA) and expanded bed adsorption (EBA) column were carried out to compare their refolding yields with that of the conventional, solution-phase refolding process. For the solution-phase and the PBA-mediated processes employing Q-Sepharose, washed inclusion body was used as the starting material, whereas both washed inclusion body and E. coli homogenate were used for the EBA-mediated process employing STREAMLINE DEAE. On a per unit mass of wet cell basis, the EBA- and PBA-mediated process showed about 4.3- and 1.7-fold higher yields, respectively, than the solution-phase refolding method. The solid-phase refolded LK68 demonstrated the same lysine-binding bioactivity and the retention time in the RP- and SEC-HPLC as those of the native protein.