Voltage-driven polyelectrolyte complexation inside a nanopore

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.