Hunter G K, Hauschka P V, Poole A R, Rosenberg L C, Goldberg H A
Faculty of Dentistry, University of Western Ontario, London, Canada.
Biochem J. 1996 Jul 1;317 ( Pt 1)(Pt 1):59-64. doi: 10.1042/bj3170059.
Many proteins found in mineralized tissues have been proposed to function as regulators of the mineralization process, either as nucleators or inhibitors of hydroxyapatite (HA) formation. We have studied the HA-nucleating and HA-inhibiting properties of proteins from bone [osteocalcin (OC), osteopontin (OPN), osteonectin (ON) and bone sialoprotein (BSP)], dentine [phosphophoryn (DPP)] and calcified cartilage [chondrocalcin (CC)] over a wide range of concentrations. Nucleation of HA was studied with a steady-state agarose gel system at sub-threshold [Ca] x [PO4] product. BSP and DPP exhibited nucleation activity at minimum concentrations of 0.3 microgram/ml (9 nM) and 10 micrograms/ml (67 nM) respectively. OC, OPN, ON and CC all lacked nucleation activity at concentrations up to 100 micrograms/ml. Inhibition of HA formation de novo was studied with calcium phosphate solutions buffered by autotitration. OPN was found to be a potent inhibitor of HA formation [IC50 = 0.32 microgram/ml (0.01 microM)] whereas OC was of lower potency [IC50 = 6.1 micrograms/ml (1.1 microM)]; BSP, ON and CC all lacked inhibitory activity at concentrations up to 10 micrograms/ml. The effect of OPN on HA formation de novo is mainly to inhibit crystal growth, whereas OC delays nucleation. These findings are consistent with the view that BSP and DPP may play roles in the initiation of mineralization in bone and dentine respectively. OPN seems to be the mineralized tissue protein most likely to function in the inhibition of HA formation, possibly by preventing phase separation in tissue fluids of high supersaturation.
矿化组织中发现的许多蛋白质被认为可作为矿化过程的调节剂，充当羟基磷灰石（HA）形成的成核剂或抑制剂。我们研究了来自骨骼[骨钙素（OC）、骨桥蛋白（OPN）、骨粘连蛋白（ON）和骨唾液蛋白（BSP）]、牙本质[磷蛋白（DPP）]和钙化软骨[软骨钙素（CC）]的蛋白质在广泛浓度范围内的HA成核和HA抑制特性。在亚阈值[Ca]×[PO4]乘积条件下，使用稳态琼脂糖凝胶系统研究HA的成核作用。BSP和DPP分别在最低浓度为0.3微克/毫升（9纳摩尔）和10微克/毫升（67纳摩尔）时表现出成核活性。OC、OPN、ON和CC在浓度高达100微克/毫升时均缺乏成核活性。通过自动滴定缓冲的磷酸钙溶液研究了从头开始形成HA的抑制作用。发现OPN是HA形成的有效抑制剂[IC50 = 0.32微克/毫升（0.01微摩尔）]，而OC的效力较低[IC50 = 6.1微克/毫升（1.1微摩尔）]；BSP、ON和CC在浓度高达10微克/毫升时均缺乏抑制活性。OPN对从头开始形成HA的作用主要是抑制晶体生长，而OC则延迟成核。这些发现与以下观点一致，即BSP和DPP可能分别在骨骼和牙本质矿化的起始中发挥作用。OPN似乎是最有可能在抑制HA形成中起作用的矿化组织蛋白，可能是通过防止高过饱和度组织液中的相分离来实现的。
