Positioning multiple proteins at the nanoscale with electron beam cross-linked functional polymers.
Positioning multiple proteins at the nanoscale with electron beam cross-linked functional polymers.
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DOI：
10.1021/ja804767j
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发表时间：
2009-01-21
影响因子：
15
通讯作者：
Maynard, Heather D.
中科院分区：
文献类型：
--
作者：
Christman, Karen L.;Schopf, Eric;Broyer, Rebecca M.;Li, Ronald C.;Chen, Yong;Maynard, Heather D.
构建与自然界中发现的复杂性相匹配的多组分蛋白质结构对于下一代医疗材料至关重要。在这份报告中，一个通用的方法来精确地安排多组分蛋白质纳米粒子在二维单层或三维多层格式，使用电子束光刻。八臂聚（乙二醇）在链端用生物素、马来酰亚胺、氨氧基或次氮基三乙酸进行修饰。通过1H NMR光谱分析显示反应是有效的，并且端基转化率为91- 100%。然后使用电子束将聚合物交联到Si表面上以形成微米尺寸的官能团图案。然后分别将具有生物素结合位点、游离半胱氨酸、N-末端α-氧代酰胺和组氨酸标签的蛋白质与底物在水溶液中孵育，不添加任何其他试剂。通过荧光显微镜实验，确定蛋白质与暴露的官能团位点特异性地反应以形成蛋白质微图案。然后制造多组分纳米级蛋白质图案。具有正交反应性的不同PEG在同一芯片上顺序图案化。两种不同的蛋白质从生物分子的混合物中同时组装形成多组分二维图案。原子力显微镜显示，形成了纳米尺寸的聚合物图案，荧光显微镜显示，获得了不同蛋白质的并排图案。此外，多层PEG制造产生一个官能团在另一个官能团之上的微米和纳米尺寸的图案。然后实现了不同蛋白质的精确三维排列。
Constructing multicomponent protein structures that match the complexity of those found in Nature is essential for the next generation of medical materials. In this report, a versatile method to precisely arrange multicomponent protein nanopatterns in two-dimensional single-layer or three-dimensional multilayer formats using electron beam lithography is described. Eight arm poly(ethylene glycol)s were modified at the chain ends with either biotin, maleimide, aminooxy, or nitrilotriacetic acid. Analysis by 1H NMR spectroscopy revealed that the reactions were efficient and that end group conversions were 91-100%. The polymers were then cross-linked onto Si surfaces using electron beams to form micron sized patterns of the functional groups. Proteins with biotin binding sites, a free cysteine, an N-terminal α-oxoamide, and a histidine tag, respectively, were then incubated with the substrate in aqueous solutions without the addition of any other reagents. By fluorescence microscopy experiments it was determined that proteins reacted site-specifically with the exposed functional groups to form protein micropatterns. Multicomponent nanoscale protein patterns were then fabricated. Different PEGs with orthogonal reactivity were sequentially patterned on the same chip. Simultaneous assembly of two different proteins from a mixture of the biomolecules formed the multicomponent two dimensional patterns. Atomic force microscopy demonstrated that nanometer sized patterns of polymer were formed and fluorescence microscopy demonstrated that side-by-side patterns of the different proteins were obtained. Moreover, multilayer PEG fabrication produced micron and nanometer sized patterns of one functional group on top of the other. Precise three-dimensional arrangements of different proteins were then realized.
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影响因子：
2.9
作者：
Arnold, M;Cavalcanti-Adam, EA;Spatz, JP
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作者：
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通讯作者：
Maynard, Heather D.
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