Open AccessGene synthesis, expression in Escherichia coli and purification of immunoreactive human insulin‐like growth factors I and II
Author(s) -
HUMMEL Michael,
HERBST Hermann,
STEIN Harald
Publication year - 1989
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1989.tb14681.x
Subject(s) - escherichia coli , fusion protein , biochemistry , amino acid , chromatography , biology , formic acid , gene , gene product , high performance liquid chromatography , chemistry , microbiology and biotechnology , gene expression , recombinant dna
We have expressed synthetic genes encoding human insulin‐like growth factors I and II in large quantities in Escherichia coli as fusion proteins with the 300 N‐terminal amino acids of the E. coli trpE gene product. The resulting hybrid proteins were purified from the insoluble fraction of crude bacterial lysates using a rapid HPLC separation procedure employing a C 8 reversed‐phase column and a gradient of 2‐propanol in formic acid. With the procedure we obtained in volatile solvents highly purified fusion proteins that were used for further biochemical and immunological procedures. Here we describe biochemical characteristics of the bacterially expressed fusion proteins and demonstrate that these proteins share substantial antigenic properties with the native polypeptides allowing the establishment of highly specific monoclonal antibodies.