We have investigated how a pair of oppositely charged macromolecules can be driven by an electric field to form a polyelectrolyte complex inside a nanopore. To observe and isolate an individual complex pair, a model protein nanopore, embedded in artificial phospholipid membrane, allowing compartmentalization (cis/trans) is employed. A polyanion in the cis and a polycation in the trans compartments are subjected to electrophoretic capture by the pore. We find that the measured ionic current across the pore has a distinguishable signature of complex formation, which is different from the signature of the passage of individual molecules through the pore. The ionic current signature allows us to detect the interaction between the two oppositely charged macromolecules and thus, enables us to measure the lifetime of the complex inside the nanopore. After showing that we can isolate a complex pair in the nanopore, we studied the effects of molecular identity on the nature of interaction in different complex pairs. In contrast to the irreversible conductance state of the alpha hemolysin (alpha HL) channel in the complexation of poly-styrene sulfonate (PSS) and poly L lysine (PLL), a reversible conductance state is observed during complexation between single stranded DNA (ssDNA) and PLL. This suggests that there is a weak interaction between ssDNA and PLL, when compared to the interaction in a PSS PLL complex. Analysis of the PSS-PLL complexation events and its lifetime inside the nanopore supports a four step mechanism: (i) The polyanion is captured by the pore, (ii) the polyanion starts threading through the pore. (iii) The polycation is captured, a complex pair is formed in the pore, and the polyanion slides along the polycation. (iv) The complex pair can be pulled through the pore into the trans compartment or it can dissociate. Additionally, we have developed a simple theoretical model, which describes the lifetime of the complex inside the pore. The observed reversible two-state conductance across alpha HL channel during ssDNA PLL complexation, is described as the binding/unbinding of PLL during the translocation of ssDNA. This enables us to evaluate the apparent rate constants for association/dissociation and equilibrium dissociation constants for the interaction of PLL with ssDNA. This work throws light on the behavior of polyelectrolyte complexes in an electric field and enhances our understanding of the electrical aspects of inter-macromolecular interactions, which plays an extremely important role in the organization of macromolecules in the crowded and confined cellular environment.

中文翻译：

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纳米孔内电压驱动的聚电解质络合

我们研究了一对带相反电荷的大分子如何被电场驱动以在纳米孔内形成聚电解质复合物。为了观察和分离单个复杂对，采用了嵌入人工磷脂膜的模型蛋白质纳米孔，允许分隔（顺式/反式）。顺式隔室中的聚阴离子和反式隔室中的聚阳离子受到孔的电泳捕获。我们发现测量的穿过孔的离子电流具有可区分的复合物形成特征，这与单个分子通过孔的特征不同。离子电流特征使我们能够检测两个带相反电荷的大分子之间的相互作用，从而使我们能够测量纳米孔内复合物的寿命。在证明我们可以在纳米孔中分离复杂对之后，我们研究了分子同一性对不同复杂对中相互作用性质的影响。与聚苯乙烯磺酸盐 (PSS) 和聚 L 赖氨酸 (PLL) 复合中 α 溶血素 (alpha HL) 通道的不可逆电导状态相反，在单链 DNA (ssDNA) 复合过程中观察到可逆电导状态和锁相环。这表明，与 PSS PLL 复合物中的相互作用相比，ssDNA 和 PLL 之间的相互作用较弱。对 PSS-PLL 络合事件及其在纳米孔内的寿命的分析支持四步机制：(i) 聚阴离子被孔捕获，(ii) 聚阴离子开始穿过孔。(iii) 聚阳离子被捕获，在孔中形成复合对，并且聚阴离子沿着聚阳离子滑动。(iv) 复合物对可以通过孔进入反式隔室，也可以解离。此外，我们还开发了一个简单的理论模型，它描述了孔隙内复合物的寿命。在 ssDNA PLL 复合过程中观察到的跨 alpha HL 通道的可逆两态电导被描述为 ssDNA 易位过程中 PLL 的结合/解除结合。这使我们能够评估 PLL 与 ssDNA 相互作用的结合/解离的表观速率常数和平衡解离常数。这项工作揭示了聚电解质复合物在电场中的行为，并增强了我们对大分子间相互作用的电学方面的理解，