Improved Solubilization of Recombinant Human Growth Hormone Inclusion Body Produced in Escherichia coli

Recombinant human growth hormone (r-hGH) over-expressed in Escherichia coli forms inactive and insoluble aggregates as inclusion bodies in the cytoplasm. The efficient solubilization of inclusion bodies is critical for cost-effective production. Contrary to a previous report, in our production system, the solubilization method by alkaline treatment including 2M urea was ineffective. Hence various buffers containing different concentrations of urea or guanidine hydrochloride (GnHCl) at neutral and alkaline pH were attempted. Efficient solubilization (about 90%) was observed in 100 mM Tris buffer, pH 8.0, with more than 4M GnHCl, and at pH 12.5 with more than 2 M GnHCl, but not with about 8M of urea. The r-hGH solubilized at pH 12.5 containing 2M GnHCl was refolded by simple dilution and purified by DEAE. Sepharose anion-exchange chromatography. The biological activity of the resulting r-hGH was comparable with commercially available r-hGH in in vitro cell proliferation assay using the hGH-dependent cell line.