| Budget Amount *help |
¥5,300,000 (Direct Cost: ¥5,300,000)
Fiscal Year 1987: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1986: ¥4,300,000 (Direct Cost: ¥4,300,000)
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| Research Abstract |
The purpose of this study is to verify the working hypothesis that the individual matrix protein in bone and dentin functions independently as well as cooperatively undergoing quantitative and qualitative changes in order to regulate calcification. First, we have established a systematic methods for the isolation of major noncollagenous proteins including phosphophoryn, osteonectin, osteocalcin, alpha-2HS pr otein, 62K protein and 24K protein from bovene bone and dentin. Systematic methods comprised the extraction with 1.0 M NaAl and 0.5 M EDTA/1 M NC1, pH 7.4, the various types of conventional chromatography and a high performance liquid chromatography using Asahipak GS-520. By these methods, phosphophoryn was found to increase its content with the development of dentin, which suggested an unique function of this highly phosphorylated protein. Secondly, we have found that the distilled water solution of EDTA/NaC1-extracted proteins was precipitated by the addition of trace amount of calcium ions ( less than 2 mM in final concentration ).
Moreover, it was found that the precipitation occurred even in the 150 mM/5mM Tris-HC1 (pH7.4) solution of the bone proteins when the calcium ions were added to the final concentration of 1mM. High performance liquid chromatography revealed that the proteins involved in the precipitation were at least osteonectin and 62K proteins, and that these two proteins associated to form the higher molecular components at the calcium concentation of less than 250 uM. These association of bone proteins preceded the apparent calciuminduced precipitation. Thus it was shown that these two proteins interact with calcium ions, associate themselves and finaly prcipitate with a small change of calcium concenration. The results suggested that these two proteins may play a role of immobilization of the initial clusters of calcium and phosphate ions and regulation of the calcification if bone tissue.
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