Tsuji Toru, Onuma Kazuo, Yamamoto Akira, Iijima Mayumi, Shiba Kiyotaka
Division of Protein Engineering, Cancer Institute, Japanese Foundation for Cancer Research, Koto, Tokyo 135-8550, Japan.
Proc Natl Acad Sci U S A. 2008 Nov 4;105(44):16866-70. doi: 10.1073/pnas.0804277105. Epub 2008 Oct 28.
An animal's hard tissue is mainly composed of crystalline calcium phosphate. In vitro, small changes in the reaction conditions affect the species of calcium phosphate formed, whereas, in vivo, distinct types of crystalline calcium phosphate are formed in a well-controlled spatiotemporal-dependent manner. A variety of proteins are involved in hard-tissue formation; however, the mechanisms by which they regulate crystal growth are not yet fully understood. Clarification of these mechanisms will not only lead to the development of new therapeutic regimens but will also provide guidance for the application of biomineralization in bionanotechnology. Here, we focused on the peptide motifs present in dentin matrix protein 1 (DMP1), which was previously shown to enhance hydroxylapatite (HAP) formation when immobilized on a glass substrate. We synthesized a set of artificial proteins composed of combinatorial arrangements of these motifs and successfully obtained clones that accelerated formation of HAP without immobilization. Time-resolved static light-scattering analyses revealed that, in the presence of the protein, amorphous calcium phosphate (ACP) particles increased their fractal dimension and molecular mass without increasing their gyration radii during a short period before precipitation. The protein thus facilitated reorganization of the internal structure of amorphous particles into ordered crystalline states, i.e., the direct transformation of ACP to HAP, thereby acting as a nucleus for precipitation of crystalline calcium phosphate. Without the protein, the fractal dimension, molecular mass, and gyration radii of ACP particles increased concurrently, indicating heterogeneous growth transformation.
动物的硬组织主要由结晶磷酸钙组成。在体外，反应条件的微小变化会影响所形成的磷酸钙种类，而在体内，不同类型的结晶磷酸钙则以一种严格受控的时空依赖方式形成。多种蛋白质参与硬组织的形成；然而，它们调节晶体生长的机制尚未完全了解。阐明这些机制不仅将导致新治疗方案的开发，还将为生物矿化在生物纳米技术中的应用提供指导。在此，我们聚焦于牙本质基质蛋白1（DMP1）中存在的肽基序，该蛋白先前被证明固定在玻璃基质上时可增强羟基磷灰石（HAP）的形成。我们合成了一组由这些基序的组合排列组成的人工蛋白质，并成功获得了无需固定就能加速HAP形成的克隆。时间分辨静态光散射分析表明，在蛋白质存在的情况下，无定形磷酸钙（ACP）颗粒在沉淀前的短时间内增加了其分形维数和分子量，而其回转半径并未增加。因此，该蛋白质促进了无定形颗粒内部结构重组成有序的结晶状态，即ACP直接转化为HAP，从而充当结晶磷酸钙沉淀的晶核。没有该蛋白质时，ACP颗粒的分形维数,分子量和回转半径同时增加，表明发生了非均相生长转变。
