A step closer to membrane protein multiplexed nanoarrays using biotin-doped polypyrrole.
A step closer to membrane protein multiplexed nanoarrays using biotin-doped polypyrrole.
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DOI：
10.1021/nn406252h
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发表时间：
2014-02-25
期刊：
影响因子：
17.1
通讯作者：
Martinez KL
中科院分区：
文献类型：
--
作者：
Della Pia EA;Holm JV;Lloret N;Le Bon C;Popot JL;Zoonens M;Nygård J;Martinez KL
无论是基础生物学研究还是诊断和药物发现应用，蛋白质微阵列和纳米阵列都是有吸引力的技术，因为它们的低样品消耗，高通量和多路复用能力。然而，到目前为止开发的阵列平台仍然不能处理膜蛋白，需要选择性地固定这些疏水和脆弱分子的特定方法来了解它们的功能和结构复杂性。在这里，我们整合了两种技术，电聚合和amphipols，以证明与膜蛋白的微和纳米表面的电寻址功能化。在生物素存在下，通过电生成聚合物膜来选择性地修饰金表面，然后可以选择性地固定抗生物素蛋白缀合物。该方法被成功地应用于制备蛋白质多重阵列的顺序电聚合和生物分子功能化的步骤。结合到电极上的蛋白质的表面密度可以通过调节电聚合过程中沉积的生物素的量来容易地调节。Amphipol是专门设计的两亲聚合物，提供了一种简单的方法来稳定和添加功能的膜蛋白。利用生物素对链霉亲和素的强亲和力，我们通过生物素标记的两性分子将不同的膜蛋白锚在不同的电极上。抗体识别事件表明，蛋白质是稳定固定的，并且带有生物素单元的聚吡咯膜的电沉积与蛋白质结合活性是相容的。由于聚吡咯膜显示出良好的导电性能，这里描述的平台特别适合于制备电子转导的生物纳米传感器。
Whether for fundamental biological research or for diagnostic and drug discovery applications, protein micro- and nanoarrays are attractive technologies because of their low sample consumption, high-throughput, and multiplexing capabilities. However, the arraying platforms developed so far are still not able to handle membrane proteins, and specific methods to selectively immobilize these hydrophobic and fragile molecules are needed to understand their function and structural complexity. Here we integrate two technologies, electropolymerization and amphipols, to demonstrate the electrically addressable functionalization of micro- and nanosurfaces with membrane proteins. Gold surfaces are selectively modified by electrogeneration of a polymeric film in the presence of biotin, where avidin conjugates can then be selectively immobilized. The method is successfully applied to the preparation of protein-multiplexed arrays by sequential electropolymerization and biomolecular functionalization steps. The surface density of the proteins bound to the electrodes can be easily tuned by adjusting the amount of biotin deposited during electropolymerization. Amphipols are specially designed amphipathic polymers that provide a straightforward method to stabilize and add functionalities to membrane proteins. Exploiting the strong affinity of biotin for streptavidin, we anchor distinct membrane proteins onto different electrodes via a biotin-tagged amphipol. Antibody-recognition events demonstrate that the proteins are stably immobilized and that the electrodeposition of polypyrrole films bearing biotin units is compatible with the protein-binding activity. Since polypyrrole films show good conductivity properties, the platform described here is particularly well suited to prepare electronically transduced bionanosensors.
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