A relationship between the starting secondary structure of recombinant porcine growth hormone solubilised from inclusion bodies and the yield of native (monomeric) protein after in vitro refolding

• Published in: FEBS letters Vol. 305; no. 3; pp. 177 - 180
• Main Authors: Puri, N.K., Cardamone, M.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 06-07-1992; Elsevier
• Subjects: Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; Cationic surfactant; Cetrimonium; Cetrimonium Compounds - chemistry; Escherichia coli - chemistry; Fundamental and applied biological sciences. Psychology; General aspects, investigation methods; Growth hormone; Growth Hormone - chemistry; Growth Hormone - isolation & purification; Guanidine; Guanidines - chemistry; In vitro refolding; Inclusion body; Protein Denaturation; Protein Folding; Protein Structure, Secondary; Proteins; Recombinant Proteins - chemistry; Recombinant Proteins - isolation & purification; Secondary structure; Solubility; Swine; Urea - chemistry; Cationic surfactant; Inclusion body; Secondary structure; Growth hormone; In vitro refolding; Chemical compound; Refolding; STH; Solubilization; In vitro; Pig; Protein hormone; Vertebrata; Denaturation renaturation; Mammalia; Artiodactyla; Recombinant protein; Ungulata
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(92)80661-Y

Recombinant porcine growth hormone (rPGH) was solubilised from inclusion bodies (IB's) using either 6 M guanidinium hydrochloride (GnHCl), 7.5 M urca or by a novel method using a cationic surfactant, cetyltrimethylammonium chloride (CTAC). Circular dichroism (CD) analysis of the secondary (2°) structure of the urea- and GnHCl-solubilised rPGH showed the absence of α-helical content with the majority of the molecule existing in a ‘random coil’ structure. In contrast, the CTAC-solubilised rPGH displayed significant starting 2° structure (10–15% α helix; 30–40% β structure). The three rPGH preparations were refolded in vitro against weak urea, GnHCl or aqueous buffers, resulting in an average refolding efficiency of 50% native (monomeric) rPGH for CTAC solubilised IB's and only 20% for urea or GnHCl solubilised IB's. We conclude that the method of solubilisation of IB's and the resultant difference in the starting 2° structure of rPGH, particularly α-helical content, is a major in vitro factor that apparently predetermines the aggregation/refolding behaviour rPGH irrespective of refolding environment.