Characterization of Protein Unfolding with Solid-state Nanopores

Abstract

In this work, we review the process of protein unfolding characterized by a solid-state nanopore based device. The occupied or excluded volume of a protein molecule in a nanopore depends on the protein’s conformation or shape. A folded protein has a larger excluded volume in a nanopore thus it blocks more ionic current flow than its unfolded form and produces a greater current blockage amplitude. The time duration a protein stays in a pore also depends on the protein’s folding state. We use Bovine Serum Albumin (BSA) as a model protein to discuss this current blockage amplitude and the time duration associated with the protein unfolding process. BSA molecules were measured in folded, partially unfolded, and completely unfolded conformations in solid-state nanopores. We discuss experimental results, data analysis, and theoretical considerations of BSA protein unfolding measured with silicon nitride nanopores. We show this nanopore method is capable of characterizing a protein’s unfolding process at single molecule level. Problems and future studies in characterization of protein unfolding using a solid-state nanopore device will also be discussed.

SDS-assisted protein transport through solid-state nanopores

“…In a previous study, Li and colleagues made a survey of different denaturants that facilitate the translocation of unfolded proteins through nanopores. 28 However, no detailed study has been reported on the use of SDS for nanopore translocation of proteins. More recently, Kennedy et al used SDS to translocate proteins through a sub-nanometer pore, however, no information on the detergent-protein interaction is provided.…”

Observing Changes in the Structure and Oligomerization State of a Helical Protein Dimer Using Solid-State Nanopores

“…This is likely due to the heterogeneous charge structure of the GCN4-p1 amino acid sequence ( Figure 1D). Solid-state nanopore induced protein denaturation has been studied elsewhere, 3,25,40,41 and could explain the shift away from larger amplitude (ΔI) events at high voltages. In particular, dimers have a larger excluded volume within the pore than monomers.…”

“…We interpret these blockades as a temperaturebased unfolding of the GCN4-p1 dimer into its monomer form. 9,10,22,25,44,46 Temperature-based unfolding will expose hydrophobic leucines. Similar hydrophobic exposure has been shown to drive adsorption of protein Note the appearance of long-lived (>5 ms) half-amplitude events at room temperature.…”

Hydrogen Peroxide Sensing Based on Inner Surfaces Modification of Solid-State Nanopore

“…A biased voltage is applied across a thin membrane containing a nanopore, resulting in an ionic current from one cell to another [25]. Protein molecules including folded and unfolded structures are detected and analyzed by solid-state nanopore [26–29]. The interaction of proteins can also be detected using solid-state nanopore [30, 31].…”