Solubilization of growth hormone and other recombinant proteins from Escherichia coli inclusion bodies by using a cationic surfactant

Abstract

Recombinant pig growth hormone (rPGH) was solubilized from inclusion bodies by using the cationic surfactant cetyltrimethylammonium chloride (CTAC). The solubilizing action of CTAC appeared to be dependent on the presence of a positively charged head group, as a non-charged variant was inactive. Relatively low concentrations of CTAC were required for rapid solubilization, and protein-bound CTAC was easily removed by ion-exchange chromatography. Compared with solubilization and recovery of rPGH from inclusion bodies with 7.5 M-urea and 6 M-guanidinium chloride, the relative efficiency of solubilization was lower with CTAC. However, superior refolding efficiency resulted in final yields of purified rPGH being in the order of CTAC greater than urea greater than or equal to guanidinium chloride. Detailed comparison of the different rPGH preparations as well as pituitary-derived growth hormone by h.p.l.c., native PAGE, c.d. spectral analysis and radioreceptor-binding assay showed that the CTAC-derived rPGH was essentially indistinguishable from the urea and guanidinium chloride preparations. The CTAC-derived rPGH was of greater biopotency than pituitary-derived growth hormone. The advantages of CTAC over urea and guanidinium chloride for increasing recovery of monomeric rPGH by minimizing aggregation during refolding in vitro were also found with recombinant sheep interleukin-I beta and a sheep insulin-like growth factor II fusion protein. In addition, the bioactivity of the CTAC-derived recombinant interleukin-1 beta was approximately ten-fold greater than that of an equivalent amount obtained from urea and guanidinium chloride preparations. It is concluded that CTAC represents, in general, an excellent additional approach or a superior alternative to urea and in particular guanidinium chloride for solubilization and recovery of bioactive recombinant proteins from inclusion bodies.