ISOLATION AND BIOLOGICAL PROPERTIES OF OSTEOPONTIN FROM BOVINE-MILK

A procedure for the isolation of osteopontin (OPN) from bovine milk us ing ion-exchange and hydrophobic chromatography is described. A DEAE-S ephacel column followed by dual phenyl-Sepharose columns yielded simil ar to 8 mg of purified protein per liter of milk. SDS-PAGE analysis re vealed that the protein migrated at M(r) 60,000. NH2-terminal sequence analysis of the first seven amino acids revealed the protein to be id entical to that previously reported for bovine OPN. Also, our preparat ion demonstrated expected biological properties of OPN including adhes ion of both endothelial and vascular smooth muscle cells to OPN in a d ose- and Arg-Gly-Asp dependent manner. Furthermore, OPN coupled to Sep harose was capable of binding the alpha(v) beta(3) integrin from a det ergent extract of endothelial cells. Thus, our procedure yielded biolo gically active OPN from an abundant and natural source. (C) 1997 Acade mic Press.