Isolation and amino-terminal sequence of a novel cellular growth inhibitor (inhibitory diffusible factor 45) secreted by 3T3 fibroblasts

A growth inhibitory protein named inhibitory diffusible factor 45 (IDF45) has been purified to homogeneity from medium conditioned by dense cultures of mouse 3T3 cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of fast protein liquid chromatography-purified bioactive material showed a single band corresponding to a protein of 45 kDa. The molecule is a monomer. The pI of the molecule, as determined by isoelectric focusing, is about 6.5. The amino-terminal sequence of the protein was established as Ser-Ala-Gly-Ala-Val-Gly-Ala-Gly-Pro-Val-Val-Arg. The protein inhibits DNA synthesis in chicken embryo fibroblasts in a dose-dependent manner (ED50: 40 ng/ml, approximately 1 nM). The chemical properties of IDF45, i.e. molecular weight and amino-terminal sequence, clearly distinguish it from other known growth inhibitory proteins.