Bradley Ledden

Electrical characterization of protein molecules by a solid-state nanopore

The authors measured ionic current blockages caused by protein translocation through voltage-biased silicon nitride nanopores in ionic solution. By calculating the mean amplitude, time duration, and the integral of current blockages, they estimated the relative charge and size of protein molecules at a single molecule level. The authors measured the change in protein charge of bovine serum albumin (BSA) protein induced by pH variation. They also confirmed that BSA molecules indeed traverse nanopores using an improved chemiluminescent analysis. They demonstrated that a larger protein fibrinogen could be distinguished from BSA by a solid-state nanopore measurement.

Sensing Single Protein Molecules with Solid-State Nanopores

This chapter is focused on the development of experiments and theory of using solid-state nanopores for sensing single protein molecules in their native and unfolded states. Proteins serve diverse roles such as transport carriers, catalysts, molecular motors, cellular structural support, and others that make life possible. Because of these widely differing roles, proteins have an enormously diverse set of shapes, sizes, and charge structures as compared to polynucleic acids. Solid-state nanopores are particularly suitable for characterizing single protein molecules because they can be fabricated with adjustable dimensions and are stable under conditions that denature proteins. This chapter describes the nanopore experimental setup, signal recording, data analysis, and basic principles related to the experiments and the theory connecting the electrical signal with the properties of proteins. Examples of experimental results illustrate the ability of solid-state nanopores to differentiate proteins in their folded and unfolded states. Native-state protein nanopore translocation follows biased one-dimensional diffusion of charged particles that is sensitive to size and electrical charge. Due to the heterogeneous charge sequence of polypeptides, unfolded proteins obey a coupled electrophoretic and thermally activated process that is sequence specific. The chapter concludes with a discussion of future directions and open challenges for single protein characterization using solid-state nanopores.