Production and characterization of recombinant chicken insulin-like growth factor-I from Escherichia coli

J Mol Endocrinol. 1992 Aug;9(1):83-92. doi: 10.1677/jme.0.0090083.

Abstract

Recombinant chicken insulin-like growth factor-I (cIGF-I) has been produced in Escherichia coli after first modifying a plasmid that coded for a human IGF-I (hIGF-I) fusion protein, in order to introduce codons for the eight amino acid substitutions. The cIGF-I fusion protein, deposited in bacterial inclusion bodies, was dissolved under reducing conditions, desalted, subjected to anion-exchange chromatography to remove proteinases, refolded and partially purified by reverse-phase high-performance liquid chromatography. The fusion protein was cleaved with hydroxylamine after which cIGF-I was purified to homogeneity by three additional chromatographic steps. Recombinant cIGF-I was equipotent with hIGF-I in cell culture bioassays of protein synthesis and breakdown using rat L6 myoblasts and chick embryo fibroblasts. Binding of radiolabelled cIGF-I and hIGF-I was also equivalent in the two cell lines, as was their binding in ligand blots of chicken, sheep and human plasma. The cross-reactivity of cIGF-I in a polyclonal hIGF-I radioimmunoassay was 60% of that observed with hIGF-I. The availability of recombinant cIGF-I will facilitate investigations into the role of IGF-I in chicken growth and development.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Chick Embryo
  • Chromatography, High Pressure Liquid
  • Chromatography, Ion Exchange
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics*
  • Insulin-Like Growth Factor I / biosynthesis*
  • Insulin-Like Growth Factor I / genetics
  • Insulin-Like Growth Factor I / metabolism
  • Molecular Sequence Data
  • Plasmids
  • Radioimmunoassay
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism

Substances

  • Recombinant Proteins
  • Insulin-Like Growth Factor I