Mizuno M, Zhou H Y, Yamada H, Kawamura M, Hirano H, Kuboki Y
Department of Biochemistry, School of Dentistry, Hokkaido University, Sapporo, Japan.
Calcif Tissue Int. 1992 Aug;51(2):156-61. doi: 10.1007/BF00298505.
Time-dependent changes of the [3H]-proline-labeled noncollagenous proteins synthesized by the osteogenic cell-line MC3T3-E1 were analyzed over a range starting from cell confluency to 13 days after confluency during which time cells formed a bone-like structure. It was found that the 40 kDa protein on SDS-polyacrylamide gel (SDS-PAGE) remarkably increased in the cell-matrix layer at about 9 days after cell confluence, just before calcification. This protein was highly purified and was found to contain high amounts of glutamic acid, glycine, and serine. An internal amino acid sequence of this protein was revealed to be K-X-M-A-P-E-E-X-P, which showed homology with the sequence of the EF-hand domain in osteonectin/SPARC (secreted protein, acidic, and rich in cysteine). This protein co-migrated with collagen in gel filtration, and ion-exchange chromatography. Furthermore, it showed high affinity to type I collagen.
分析了成骨细胞系MC3T3-E1合成的[3H]-脯氨酸标记的非胶原蛋白在从细胞汇合到汇合后13天（在此期间细胞形成类骨结构）的时间依赖性变化。发现在SDS-聚丙烯酰胺凝胶（SDS-PAGE）上40 kDa的蛋白质在细胞汇合后约9天，即钙化前，在细胞-基质层中显著增加。该蛋白质被高度纯化，发现含有大量的谷氨酸、甘氨酸和丝氨酸。该蛋白质的内部氨基酸序列显示为K-X-M-A-P-E-E-X-P，与骨粘连蛋白/SPARC（分泌蛋白，酸性，富含半胱氨酸）中EF-手结构域的序列具有同源性。该蛋白质在凝胶过滤和离子交换色谱中与胶原蛋白共迁移。此外，它对I型胶原显示出高亲和力。
