Self-assembling protein hydrogels with modular integrin binding domains
Self-assembling protein hydrogels with modular integrin binding domains
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DOI：
10.1021/bm050157p
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发表时间：
2006-01-01
影响因子：
6.2
通讯作者：
Harden, JL
中科院分区：
文献类型：
--
作者：
Mi, LX;Fischer, S;Harden, JL
利用嵌入RGD序列的相关蛋白制备了具有整合素结合活性的水凝胶。这些蛋白是一种改良的AC(10)Bcys三片段设计，由酸性a和碱性B亮氨酸拉链相关结构域组成，其两侧是一个新的可溶性无序线圈片段，包含9个重复的AGAGAGPEG和3个拷贝的RGD整合素结合序列。与没有嵌入RGD序列的原始AC(10)Bcys设计一样，在一定的溶液pH和温度条件下，这些蛋白质在超过约50 mg/mL的浓度下自组装成稳定的水凝胶。水凝胶的组装机制是A和B螺旋结构域的分子间结合成束，通过可溶的中心无序线圈结构域连接成交联。蛋白质的二级结构和它们形成的水凝胶的机械性能不受RGD序列的存在的不利影响。嵌入无序线圈区域的RGD序列支持人成纤维细胞在蛋白包被表面上的粘附、扩散和极化。共聚焦显微镜研究表明，这些细胞中存在黏附复合物和有组织的肌动蛋白应力纤维。相比之下，将成纤维细胞播种到原AC(10)Bcys蛋白包被的表面上，成纤维细胞保持圆形，不形成局灶性粘附，表明嵌入RGD序列的存在赋予了生物活性。这种水凝胶形成的生物活性蛋白具有潜在的细胞和组织培养应用。
Hydrogels with integrin binding activity were created from associating proteins with embedded RGD sequences. These proteins are a modified AC(10)Bcys triblock design composed of acidic A and basic B leucine zipper associating domains flanking a new soluble disordered coil block that contains nine repeats of AGAGAGPEG and three copies of the RGD integrin binding sequence. As with the original AC(10)Bcys design without the embedded RGD sequences, these proteins self-assemble into stable hydrogels at concentrations above approximately 50 mg/mL in a range of solution pH and temperature conditions. The mechanism for hydrogel assembly is the intermolecular association of A and B helical domains into bundles which act as cross-links connected by the soluble central disordered coil domains. The secondary structure of the proteins and the mechanical properties of the hydrogels they form are not adversely affected by the presence of the RGD sequences. The RGD sequences embedded in the disordered coil region support the adhesion, spreading, and polarization of human fibroblast cells on protein coated surfaces. Confocal microscopy studies demonstrated the presence of focal adhesion complexes and organized actin stress fibers in these cells. In contrast, fibroblasts seeded onto surfaces coated with the original AC(10)Bcys protein remained rounded and did not form focal adhesions, indicating that bioactivity is conferred by the presence of the embedded RGD sequences. Such hydrogel-forming bioactive proteins have potential for cell and tissue culture applications.