Nonequilibrium Capture Rates Induce Protein Accumulation and Enhanced Adsorption to Solid-State Nanopores

Single molecule capturing of analytes using an electrically biased nanopore is the fundamental mechanism in which nearly all nanopore experiments are conducted. With pore dimensions being on the order of a single molecule, the spatial zone of sensing only contains approximately a zeptoliter of volume. As a result, nanopores offer high precision sensing within the pore but provide little to no information about the analytes outside the pore. In this study, we use capture frequency and rate balance theory to predict and study the accumulation of proteins at the entrance to the pore. Protein accumulation is found to have positive attributes such as capture rate enhancement over time but can additionally lead to negative effects such as long-term blockages typically attributed to protein adsorption on the surface of the pore. Working with the folded and unfolded states of the protein domain PDZ2 from SAP97, we show that applying short (e.g., 3–25 s in duration) positive voltage pulses, rather than a constant voltage, can prevent long-term current blockades (i.e., adsorption events). By showing that the concentration of proteins around the pore can be controlled in real time using modified voltage protocols, new experiments can be explored which study the role of concentration on single molecular kinetics including protein aggregation, folding, and protein binding.