Department of Materials, ‡Department of Bioengineering, §Institute of Biomedical Engineering, and ⊥Department of Chemistry, Imperial College London , London SW7 2AZ, United Kingdom.
ACS Nano. 2016 Dec 27;10(12):11096-11104. doi: 10.1021/acsnano.6b05975. Epub 2016 Dec 7.
Cells in the body use a variety of mechanisms to ensure the specificity and efficacy of signal transduction. One way that this is achieved is through tight spatial control over the position of different proteins, signaling sequences, and biomolecules within and around cells. For instance, the extracellular matrix protein fibronectin presents RGDS and PHSRN sequences that synergistically bind the αβ integrin when separated by 3.2 nm but are unable to bind when this distance is >5.5 nm.1 Building biomaterials to controllably space different epitopes with subnanometer accuracy in a three-dimensional (3D) hydrogel is challenging. Here, we synthesized peptides that self-assemble into nanofiber hydrogels utilizing the β-sheet motif, which has a known regular spacing along the peptide backbone. By modifying specific locations along the peptide, we are able to controllably space different epitopes with subnanometer accuracy at distances from 0.7 nm to over 6 nm, which is within the size range of many protein clusters. Endothelial cells encapsulated within hydrogels displaying RGDS and PHSRN in the native 3.2 nm spacing showed a significant upregulation in the expression of the alpha 5 integrin subunit compared to those in hydrogels with a 6.2 nm spacing, demonstrating the physiological relevance of the spacing. Furthermore, after 24 h the cells in hydrogels with the 3.2 nm spacing appeared to be more spread with increased staining for the αβ integrin. This self-assembling peptide system can controllably space multiple epitopes with subnanometer accuracy, demonstrating an exciting platform to study the effects of ligand density and location on cells within a synthetic 3D environment.
细胞利用多种机制来确保信号转导的特异性和功效。一种实现方式是通过严格控制细胞内外不同蛋白质、信号序列和生物分子的位置。例如，细胞外基质蛋白纤维连接蛋白呈现 RGDS 和 PHSRN 序列，当它们之间的距离为 3.2nm 时，这些序列能够协同结合αβ整合素，但当距离大于 5.5nm 时，它们就无法结合。1 构建生物材料，以亚纳米精度在三维（3D）水凝胶中可控地间隔不同表位是具有挑战性的。在这里，我们合成了自组装成纳米纤维水凝胶的肽，利用β-折叠基序，该基序沿肽骨架具有已知的规则间隔。通过修饰肽上的特定位置，我们能够以亚纳米精度在 0.7nm 至 6nm 以上的距离可控地间隔不同的表位，这在许多蛋白质簇的尺寸范围内。在以天然 3.2nm 间距显示 RGDS 和 PHSRN 的水凝胶中包封的内皮细胞与在间距为 6.2nm 的水凝胶中的细胞相比，α5 整合素亚基的表达显著上调，证明了间距的生理相关性。此外，在 24 小时后，在间距为 3.2nm 的水凝胶中的细胞似乎更分散，αβ整合素的染色增加。这种自组装肽系统能够以亚纳米精度可控地间隔多个表位，展示了一个令人兴奋的平台，可以在合成的 3D 环境中研究配体密度和位置对细胞的影响。
