The extraction and characterization of soluble anionic phosphoproteins from bone

Embryonic bovine jaw and femur bone, and 14-week-old chicken metatarsal bone were demineralized in 0.2 M EDTA. Analysis of the extracts revealed proteins rich in the acidic amino acids, aspartic and glutamic acids, and containing from 0.75% organic phosphorus in the femur extracts to 0.54% organic phosphorus in the chicken bone extracts. In addition, hexose, hexosamine, uronic acid and sialic acid were also present. Further purification of the peptides solubilized in EDTA was achieved by Sephadex G-100 molecular sieve filtration and Whatman DE-52 ion exchange chromatography. From these procedures a component was isolated that appeared to be homogeneous by acrylamide disc gel electrophoresis and ion exchange chromatography. Its amino acid composition revealed a high content of anionic amino acids and the presence of hydroxyproline in both bovine jaw bone and femur bone, but not in chicken bone. Organic phosphorus and serine were present in almost equimolar amounts. A possible role for these anionic phosphorus-containing proteins in facilitating the initiation of mineralization is discussed.