Protein-Resistant NTA-Functionalized Polymer Brushes for Selective and Stable Immobilization of Histidine-Tagged Proteins
Protein-Resistant NTA-Functionalized Polymer Brushes for Selective and Stable Immobilization of Histidine-Tagged Proteins
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DOI：
10.1021/am9006484
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发表时间：
2010-01-01
影响因子：
9.5
通讯作者：
Ramstedt, Madeleine
中科院分区：
文献类型：
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作者：
Gautrot, Julien E.;Huck, Wilhelm T. S.;Ramstedt, Madeleine
高选择性固定特定生物分子的抗蛋白质聚合物涂层对于生物医学应用如微阵列、生物传感、多相催化和生物工程是必不可少的。聚合物刷在这方面特别令人感兴趣，因为它们的化学结构和物理性质可以很容易地定制以满足特定的需求。本文探讨了两种蛋白质抗性聚合物刷，聚（低聚乙二醇甲基丙烯酸酯）（POEGMA）和聚（甲基丙烯酸羟乙酯）（PHEMA），与氨三乙酸（NTA）部分，可以复杂的组氨酸标记（His标记）的蛋白质选择性和可逆的功能化。利用荧光显微镜、红外光谱、X射线光电子能谱、表面等离子体共振和椭圆偏振光谱等技术，我们证明了组氨酸标记的绿色荧光蛋白可以高稳定性和高负载量地固定在NTA刷上。NTA-POEGMA达到的负载饱和度高于NTA-PHEMA，因为前者刷的溶胀增加。尽管具有更高的负载能力，NTA-POEGMA仍然保持高度的蛋白质抗性，这表明其具有“清洁”和特异性蛋白质固定的潜力。最后，我们表明，NTA-POEGMA刷的保留的蛋白质抗性可以用于通过孵育和洗涤的简单协议来产生定义明确的二元生物功能模式。这些模式可能会在细胞阵列和筛选中找到应用。
Protein-resistant polymeric coatings that allow highly selective immobilization of specific biomolecules are essential for biomedical applications such as microarrays, biosensing, heterogeneous catalysis, and bioengineering. Polymer brushes are particularly interesting for this purpose because their chemical structure and physical properties can easily be tailored to meet specific needs. This article explores the functionalization of two protein-resistant polymer brushes, poly(oligoethylene glycol methacrylate) (POEGMA) and poly(hydroxyethyl methacrylate) (PHEMA), with nitrilotriacetic acid (NTA) moieties that can complex histidine-tagged (His-tagged) proteins selectively and reversibly. Using Fluorescence microscopy, IR spectroscopy, X-ray photoelectron spectroscopy, surface plasmon resonanace, and ellipsometry, we demonstrate that His-tagged green fluorescent protein can be immobilized on NTA brushes with high stability and loading. The loading saturation reached for NTA-POEGMA is higher than that for NTA-PHEMA because of increased swelling of the former brush. Despite this higher loading capacity, NTA-POEGMA remained highly protein-resistant, which shows its potential for "clean" and specific protein immobilization. Finally, we showed that the preserved protein resistance of NTA-POEGMA brushes can be used to generate well-defined binary biofunctional patterns via a simple protocol of incubations and washes. These patterns may find applications in cell arraying and screening.