Hydrogels are excellent scaffolds to accommodate sensitive enzymes in a protective environment. However, the lack of suitable immobilization techniques on substrates and the lack of selectivity to anchor a biocatalyst are major drawbacks preventing the use of hydrogels in bioanalytical devices. Here, nanofilm coatings on surfaces were made of a recombinant spider silk protein (rssp) to induce rssp self-assembly and thus the formation of fibril-based nanohydrogels. To functionalize spider silk nanohydrogels for bioselective binding of proteins, two different antithrombin aptamers were chemically conjugated with the rssp, thereby integrating the target-binding function into the nanohydrogel network. Human thrombin was selected as a sensitive model target, in which the structural integrity determines its activity. The chosen aptamers, which bind various exosites of thrombin, enabled selective and cooperative embedding of the protein into the nanohydrogels. The change of the aptamer secondary structure using complementary DNA sequences led to the release of active thrombin and confirmed the addressable functionalization of spider silk nanohydrogels.

水凝胶是在保护性环境中容纳敏感酶的优秀支架。然而，缺乏合适的基质固定技术以及缺乏锚定生物催化剂的选择性是阻碍水凝胶在生物分析装置中使用的主要缺点。在这里，表面的纳米薄膜涂层由重组蜘蛛丝蛋白（rssp）制成，以诱导rssp自组装，从而形成基于原纤维的纳米水凝胶。为了使蜘蛛丝纳米水凝胶功能化以生物选择性地结合蛋白质，将两种不同的抗凝血酶适体与rssp化学缀合，从而将靶标结合功能整合到纳米水凝胶网络中。选择人凝血酶作为敏感模型靶标，其中结构完整性决定其活性。所选择的适体结合了凝血酶的各种外部位点，使得蛋白质能够选择性且协同地嵌入纳米水凝胶中。使用互补DNA序列改变适体二级结构导致活性凝血酶的释放，并证实了蜘蛛丝纳米水凝胶的可寻址功能化。