Comparison of the immunochemical, receptor binding and biological properties of natural insulin-like growth factor I/Somatomedin C (IGF-I/Sm C) and its recombinant DNA-synthesized Thr-59 analog

Abstract

We have synthesized a gene coding for an analog of TGF-I/Sm C and cloned it into a plasmid for expression in E. coli. This analog, containing Thr-59 and 8 extra amino acids at the N-terminal, was purified to homogeneity and tested for various activities. The 125I-labeled IGF-I analog has an affinity for Sm binding protein as well as antibodies raised against human-derived IGF-I that is slightly less than the natural molecule. Both 125I-labeled IGF-I and its analog bind to a specific cell membrane receptor on cultured human fibroblasts (HF) and rat chondrocytes (RC) in a time-, temperature-, and concentration-dependent manner. The analog stimulates the incorporation of 3H-thymidine into DNA in cultured RC and HF in a dose-response manner, with half-maximal stimulation at 12.5 and 62.5 ng/ml respectively, and is equipotent to natural IGF-I. In contrast, the Thr-59 analog did not enhance 35S-sulfate incorporation into chondroitin sulfate in RC in a fashion similar to the natural hormone. We conclude that this IGF-I analog has similar, though not identical, immunochemical, receptor binding and biological properties to human-derived IGF-I/Sm C. Nevertheless, we anticipate this peptide will be useful for studying the activity of this Sm.