title: Calcium and collagen binding properties of osteopontin, bone sialoprotein, and bone acidic glycoprotein-75 from bone.
abstract: Calcium binding properties of bone acidic glycoprotein-75, osteopontin, and bone sialoprotein were determined in 10 mM imidazole buffer (pH 6.8), containing either 60 mM KCl or 150 mM NaCl. Proteins assayed were first bound to nitrocellulose to mimic substrate-bound forms in vivo; retention of phosphoproteins was determined through use of radioiodinated tracers. Binding studies were carried out both as a function of calcium concentration and the amount of phosphoprotein. In the presence of 60 mM KCl, bone acidic glycoprotein-75 exhibited the largest calcium binding capacity (139 atoms/molecule at saturation), with bone sialoprotein intermediary (83 atoms/molecule) and osteopontin lowest (50 atoms/molecule). Sites detected for each phosphoprotein exhibited overall binding constants in the 0.5-1.0 mM extracellular range. In 150 mM NaCl and 1-2 mM total calcium, phosphoproteins bound between 72 and 19 mol of calcium/mol with the same relative order. Binding was proportional to amount of phosphoprotein in either salt condition. The presence of 5 mM calcium had a different effect on concentration-dependent binding to type I collagen for each phosphoprotein. Bone acidic glycoprotein-75 alone was found to undergo an unusual calcium-enhanced polymerization reaction, confirmed by light scattering measurements, wherein collagen binding was greatest with polymeric forms. These findings demonstrate that acidic phosphoproteins from bone bind calcium atoms with a range of capacities. Calcium appears to induce conformational changes in bone acidic glycoprotein-75 which influences its self-association and binding to different substrata.
authors.affiliation: Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri, Kansas City 64110-2499.
authors.forename: Y
authors.initials: Y
authors.lastname: Chen
authors.forename: B S
authors.initials: BS
authors.lastname: Bal
authors.forename: J P
authors.initials: JP
authors.lastname: Gorski
mesh_terms: D000818:Animals; D001345:Autoradiography; D001842:Bone and Bones / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002132:Calcium Radioisotopes; D003094:Collagen / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008027:Light; D008297:Male; D053495:Osteopontin; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D012542:Scattering, Radiation; D012795:Sialoglycoproteins / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D002132:Calcium Radioisotopes; D006023:Glycoproteins; D010750:Phosphoproteins; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; C506897:Spp1 protein, rat; C512787:bone acidic glycoprotein-75, rat; D053495:Osteopontin; D003094:Collagen; D002118:Calcium