Isolation of mitogenically active C-terminal truncated pentapeptide of osteogenic growth peptide from human plasma and culture medium of murine osteoblastic cells

The osteogenic growth peptide (OGP) is a 14-amino acid stromal cell mitogen that stimulates in vivo osteogenesis and hematopoiesis. In the blood circu lation and cell culture conditioned medium immunoreactive OGP (irOGP), iden tified using antibodies raised against the OGP C-terminal region, presents free and bound forms. The bound form consists entirely of the full length p eptide. The present study was designed to investigate the identity of free irOGP under nondenaturing conditions. Fresh human serum and culture medium conditioned with murine osteoblastic MC3T3 E1 cells were fractionated using ultrafiltration (3000 molecular weight cut-off). Hydrophobic chromatograph y of the ultrafiltrate, immunoscreening of chromatographic fractions with a ntibodies directed against the OGP C-terminal region and amino acid sequenc ing of immunoreactive peaks demonstrated the presence of two mitogens, the full length OGP and a C-terminal truncated form, OGP(10-14). The OGP(10-14) derived the from both serum and conditioned medium, as well as the synthet ic pentapeptide [sOGP(10-14)], shared the in vitro OGP proliferative activi ty. However, in a competitive binding assay, devised to assess the OGP-OGP binding protein (OGPBP) complex formation, sOGP(10-14) failed to compete ou t radiolabeled OGP from the complex. It is concluded that OGP(10-14) is a n aturally occuring human and murine mitogen. In addition, the data suggests that the OGP(10-14) is generated from OGP by proteolytic cleavage upon diss ociation of the OGP-OGPBP complexes.