Direct expression of a synthetic somatomedin C gene in Escherichia coli by use of a two-cistron system.
Direct expression of a growth-promoting peptide hormone, somatomedin C/insulin-like growth factor I, which is quite difficult due to the instability of somatomedin C itself in Escherichia coli, has been achieved by the use of a two-cistron system. Assuming that basic somatomedin C might be stabilized by forming a complex with an acidic polypeptide, we constructed synthetic genes consisting of two cistrons; an acidic 93-amino-acid polypeptide was coded in the first cistron followed by a synthetic somatomedin C gene in the second cistron. The chain termination codon for the first polypeptide overlapped the initiation codon for the second polypeptide in the intercistronic region, as occurs in the polycistronic E. coli tryptophan operon, whose products are associated in multi-subunit enzyme complexes. In the expression of the resulting genetic system, recombinant somatomedin C associated with the acidic polypeptide was accumulated to high levels in the cells. After treatment of the product with acetic acid to dissociate the two components, the recombinant somatomedin C was isolated in a yield of 4.0 mg from a liter of culture broth at A600 = 1.6. It was determined to be Met-somatomedin C by chymotryptic mapping as well as amino-terminal analysis.
Saito
Y
Y
Central Research Laboratories, Fujisawa Pharmaceutical Co., Ltd., Osaka.
Ishii
Y
Y
Niwa
M
M
Ueda
I
I
D000595:Amino Acid Sequence; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005813:Genes, Synthetic; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000235:genetics*; D010957:Plasmids; D013002:Somatomedins / Q000235:genetics*
D013002:Somatomedins; D007334:Insulin-Like Growth Factor I