Bone sialoprotein (BSP) and osteopontin, the major phosphorylated proteins of mammalian bone, have been proposed to function in initiation mineralization. To test this hypothesis, effects BSP osteopontin on hydroxyapatite crystal formation were determined by using a steady-state agarose gel system. At low calcium phosphate concentrations, no accumulation occurred control gels or containing osteopontin. Gels at 1-5 micrograms/ml, however, exhibited visible precipitation band significantly...

Many proteins found in mineralized tissues have been proposed to function as regulators of the mineralization process, either nucleators or inhibitors hydroxyapatite (HA) formation. We studied HA-nucleating and HA-inhibiting properties from bone [osteocalcin (OC), osteopontin (OPN), osteonectin (ON) sialoprotein (BSP)], dentine [phosphophoryn (DPP)] calcified cartilage [chondrocalcin (CC)] over a wide range concentrations. Nucleation HA was with steady-state agarose gel system at...

Osteopontin is a phosphorylated sialoprotein containing conserved sequence of contiguous aspartic acid residues. This protein expressed at high levels in mineralized tissues and has previously been shown to inhibit the vitro formation hydroxyapatite (HA). In present study, modification model compound studies have used identify structural features osteopontin that are responsible for its crystal-modulating properties. Using metastable calcium phosphate solutions buffered by autotitration,...

Bone sialoprotein (BSP) is a bone-specific glycoprotein containing phosphoserine and sulphotyrosine residues regions of contiguous glutamic acid residues. Recent studies in this laboratory have shown that BSP capable nucleating the bone mineral hydroxyapatite steady-state agarose gel system. We show here chemical modification carboxylate groups abolishes nucleation activity BSP, but enzymic dephosphorylation has no effect. Formation also induced by poly(L-glutamic acid) poly(D-glutamic...

Enamelin is critical for proper dental enamel formation, and defects in the human enamelin gene cause autosomal dominant amelogenesis imperfecta. We used targeting to generate a knock-in mouse carrying null allele of (Enam) that has lacZ reporter replacing Enam translation initiation site sequences through exon 7. Correct transgene was confirmed by Southern blotting PCR analyses. No protein could be detected Western Enam-null mice. Histochemical...

Osteopontin (OPN) is an acidic phosphoglycoprotein that believed to function in the prevention of soft tissue calcification. In vitro studies have shown OPN can inhibit formation hydroxyapatite (HA) and other biologically relevant crystal phases, this inhibitory activity requires phosphorylation protein; however, it not known which phosphorylated residues are involved. We synthesized peptides corresponding four phosphoserine-containing sequences rat OPN: OPN7–17, containing phosphoserines 10...

AbstractThe phosphorylated acidic glycoproteins bone sialoprotein (BSP) and osteopontin (OPN) bind to hydroxyapatite (HA) crystals may be involved in the regulation of mineralization. The HA-binding properties these proteins have been attributed glutamic acid-rich sequences BSP aspartic OPN. present study examines roles polycarboxylate binding OPN HA.Porcine BSP, synthetic polypeptides poly-L-glutamic acid [Poly(Glu)] poly-L-aspartic [Poly(Asp)] were labeled with fluorescein isothiocyanate...

Mineral-associated proteins have been proposed to regulate many aspects of biomineralization, including the location, type, orientation, shape, and texture crystals. To understand how achieve this exquisite level control, we are studying interaction between phosphoprotein osteopontin (OPN) biomineral calcium oxalate monohydrate (COM). In present study, synthesized peptides corresponding amino acids 220−235 rat bone OPN (pSHEpSTEQSDAIDpSAEK), one several highly phosphorylated, aspartic-,...

Bone sialoprotein (BSP) is an anionic phosphorylated glycoprotein that expressed almost exclusively in mineralized tissues and has been shown to be a potent nucleator of hydroxyapatite formation. The binding BSP collagen thought important for the initiation bone mineralization adhesion cells matrix. Using solid phase assay, we have investigated interaction between collagen. Initial studies showed raising ionic strength, decreasing pH below 7, or introducing divalent cations diminishes but...

Mice lacking the gene encoding matrix gla protein (MGP) exhibit massive mineral deposition in blood vessels and die soon after birth. We hypothesize that MGP prevents arterial calcification by adsorbing to growing hydroxyapatite (HA) crystals. To test this, we have used a combined experimental–computational approach. synthesized peptides covering entire sequence of human MGP, which contains three sites serine phosphorylation five γ-carboxylation, studied their effects on HA crystal growth...

Study of the calcification cartilage in endochondral ossification has yielded two apparently contradictory views role proteoglycan this process. The ability to act as a calcium-concentrating agent (Kalksalzfänger) is consistent with view that proteoglycans are promoters calcification. However, study their effect on hydroxyapatite formation vitro suggests inhibitors A resolution paradox now proposed. Proteoglycans inhibit under conditions limited calcium availability (in part) by binding...

In vitro studies have shown that the phosphoprotein osteopontin (OPN) inhibits nucleation and growth of hydroxyapatite (HA) other biominerals. vivo, OPN is believed to prevent calcification soft tissues. However, nature interaction between HA not understood. computational part present study, we used molecular dynamics simulations predict adsorption 19 peptides, each 16 amino acids long collectively covering entire sequence OPN, {100} face HA. This analysis showed there an inverse...

Biomineralization is a complex process in the development of mineralized tissues such as bone and pathological calcifications atherosclerotic plaques, kidney stones gout. Osteopontin (OPN), an anionic phosphoprotein, expressed mineralizing has previously been demonstrated to be potent inhibitor hydroxyapatite formation. The OPN-deficient (Opn-/-) mouse displays hypermineralized phenotype starting at 12 weeks postnatally. By isolating culturing Opn-/- wild-type (WT) osteoblasts, we sought...

Bone sialoprotein (BSP) was shown to be a potent nucleator of hydroxy apatite (HA) in steady-state agarose gel system (Hunter and Goldberg, 1993, PNAS 90: 8562). Nucleation HA also demonstrated with the homopolymer poly-glutamic acid but not poly-aspartic or osteopontin. Since BSP contains contiguous sequences glutamic acid, it is reasonable suggest that HA-nucleating activity resides within these regions. Purified porcine treated trypsin digests fractionated by filtration. In addition small...

Osteopontin (OPN) is a highly modified protein that found in many tissues and has been associated with variety of physiological pathological processes. Bone OPN potent inhibitor hydroxyapatite crystal formation stimulates bone resorption by osteoclasts; these activities, as well others, are dependent upon phosphorylation the protein. We have used mass spectrometry (MS) to perform comprehensive analysis post-translational modification purified from rat bone. Matrix-assisted laser desorption...