Synthesis of a potentially bioactive, hydroxyapatite-nucleating molecule
Synthesis of a potentially bioactive, hydroxyapatite-nucleating molecule
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DOI：
10.1007/s00223-005-0118-4
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发表时间：
2006-01-01
影响因子：
4.2
通讯作者：
Clarkson, B
中科院分区：
文献类型：
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作者：
Chang, S;Chen, H;Clarkson, B
我们实验室之前从未成熟的根尖总 RNA 中克隆了人类磷酸化蛋白 (PP) cDNA。该cDNA包含2,364 bp，编码788个氨基酸。超过 80% 的序列排列为 (DSS)(n) (n = 1-16)、DS 和 NSS 基序。我们假设 PP 结合 Ca2+ 和使羟基磷灰石成核的能力可能取决于这些重复序列。基于人PP cDNA序列合成了两种多肽来检验该假设。一种多肽具有氨基酸序列DDPNSSDESNGNDD（合成多肽1，SP1），其来自PP的N-末端；另一种多肽 DSKSDSSKSESDSS（合成多肽 2，SP2）是 PP 重复序列基序。多肽的磷酸化是通过将其与三磷酸腺苷和酪蛋白激酶 I 和 II 反应来完成的。在稳态琼脂糖凝胶系统中测试了这些分子引起矿化的能力。结果表明，磷酸化SP2 (P-SP2) 沉淀的Ca + PO4 总量约占PP 沉淀总Ca + PO4 的60%。 P-SP1 的沉淀量约为 PP 沉淀量的 23%，并且与对照凝胶（即不添加肽）中的沉淀量相似。透射电子显微镜和X射线衍射分析表明，含P-SP2的凝胶中形成的沉淀物是羟基磷灰石。 P-SP2 使 Ca + PO4 成核并沉淀羟基磷灰石的能力是重复序列基序的结果，其中含有高比例的磷酸化丝氨酸。该分子可用于牙齿组织的修复和再生。
A human phosphophoryn (PP) cDNA was previously cloned from immature root apex total RNA in our laboratory. This cDNA comprises 2,364 bp, encoding 788 amino acids. More than 80% of the sequences are arranged as (DSS)(n) (n = 1-16), DS, and NSS motifs. We hypothesize that the capability of PP to bind Ca2+ and nucleate hydroxyapatite may depend on these repeated sequences. Two polypeptides were synthesized based on the human PP cDNA sequence to test the hypothesis. One polypeptide has the amino acid sequence DDPNSSDESNGNDD (synthetic polypeptide 1, SP1), which is from the N-terminal end of PP; the other polypeptide, DSKSDSSKSESDSS (synthetic polypeptide 2, SP2), is the PP repeated sequence motif. Phosphorylation of the polypeptides was accomplished by reacting them with adenosine triphosphate and casein kinases I and II. The ability of these molecules to cause mineralization was tested in a steady-state agarose gel system. The results show that phosphorylated SP2 (P-SP2) precipitated approximately 60% of the total Ca + PO4 precipitated by PP. P-SP1 precipitated about 23% of that precipitated by PP and was similar to the amount precipitated in the control gel, that is, without added peptides. Transmission electron microscopy and X-ray diffraction analysis showed that the precipitate formed in the P-SP2-containing gel was hydroxyapatite. The capability of P-SP2 to nucleate Ca + PO4 and precipitate hydroxyapatite is a result of the repeated sequence motif, which contains a high percentage of phosphorylated serine. This molecule could be used in the repair and regeneration of dental tissue.