Established methods for characterizing proteins typically require physical or chemical modification steps or cannot be used to examine individual molecules in solution. Ionic current measurements through electrolyte-filled nanopores can characterize single native proteins in an aqueous environment, but currently offer only limited capabilities. Here we show that the zeptolitre sensing volume of bilayer-coated solid-state nanopores can be used to determine the approximate shape, volume, charge, rotational diffusion coefficient and dipole moment of individual proteins. To do this, we developed a theory for the quantitative understanding of modulations in ionic current that arise from the rotational dynamics of single proteins as they move through the electric field inside the nanopore. The approach allows us to measure the five parameters simultaneously, and we show that they can be used to identify, characterize and quantify proteins and protein complexes with potential implications for structural biology, proteomics, biomarker detection and routine protein analysis.
In this work we use a combination of 3D-TEM tomography, energy filtered TEM, single molecule DNA translocation experiments, and numerical modeling to show a more precise relationship between nanopore shape and ionic conductance and show that changes in geometry while in solution can account for most deviations between predicted and measured conductance. We compare the structural stability of Ion Beam Sculpted (IBS), IBS-annealed, and TEM drilled nanopores. We demonstrate that annealing can significantly improve the stability of IBS made pores. Furthermore, the methods developed in this work can be used to predict pore conductance and current drop amplitudes of DNA translocation events for a wide variety of pore geometries. We discuss that chemical dissolution is one mechanism of the geometry change for SiNx nanopores and show that small modification in fabrication procedure can significantly increase the stability of IBS nanopores.
Ion and water transport through biological membranes by osmosis is essential in many biological processes and the complex nature of biological pores and channels involved in these processes is not fully understood. We are developing simplified robust models of ion channels based on smooth, narrow and hydrophobic pores of carbon nanotubes. Incorporation of these pores into liposomes gives us a convenient system to study osmotically-induced transport using dynamic light scattering. We will present our measurements of the transport of charged and uncharged chemical species through the carbon nanotube pores and also discuss the role of the charges at the nanotube pore ends in regulating the ion transport selectivity.
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