Full Width at Half Maximum of Nanopore Current Blockage Controlled by a Single-Biomolecule Interface.

A biological nanopore is one of the predominant single-molecule approaches as a result of its controllable single-biomolecule interface, which could reflect the "intrinsic" information on an individual molecule in a label-free way. Because the current blockage is normally treated as the most important parameter for nanopore identification of every single molecule, the fluctuation of current blockage for certain types of molecules, defined as full width at half maximum (fwhm) of current blockage, actually owns a dominant influence on nanopore resolution. Therefore, controlling the fwhm of current blockage of molecules is critical for the sensing capability of the nanopore. Here, taking an aerolysin nanopore as a model, by precisely controlling the functional group in this single-biomolecule interface, we could narrow the fwhm of nanopore current blockage for DNA identification and prolong the duration inside the nanopore. Moreover, a substantial correlation between fwhm of current blockage and duration is established, showing a non-monotonic variation. Besides, the mechanism is also clarified with studying the detailed current blockage events. This proposed correlation is further demonstrated to be applied uniformly across different mutant aerolysins for a certain DNA. This study proposes a new strategy for regulating molecular sensing from the duration of the analyte, which could guide the resolution of heterogeneity analysis using nanopores.

3D Blockage Mapping for Identifying Familial Point Mutations in Single Amyloid-ß Peptides with a Nanopore.

Accurate discrimination of amyloid-ß (Aß) peptides containing familial point mutations would advance the knowledge of their roles in early-onset Alzheimer's disease. Herein, we simultaneously identified the mutant A21G, E22G, E22Q, and the wild-type (WT) Aß18-26 peptides with aerolysin nanopore using a 3D blockage mapping strategy. The standard deviation of current blockade fluctuations (σb ) was proposed as a new supplement to current blockage (Ib /I0 ) and duration time (tD ) to profile the blockage characteristics of single molecules. Although the WT and A21G Aß18-26 are indistinguishable in a traditional Ib /I0 -tD 2D description, ∼87 % of the blockade events can be accurately classified with half reduction of false identification using a combination of Ib /I0 , tD, and σb . This work offers an easy and reliable strategy to promote nanopore sensitivity of peptide mutants, leading to a more precise analysis of pathogenic mutations for developing effective diagnosis and treatment.

Profiling the chemistry- and confinement-controlled sensing capability of an octameric aerolysin-like protein.

Octameric Aep1 was employed, for the first time to the best of our knowledge, as a nanopore to expand applications. After investigating the optimized conditions of Aep1 for single-channel recording, the sensing features were characterized. Cyclic and linear molecules of varying sizes and charges were employed to probe the radius and chemical environment of the pore, providing deep insights for expected future endeavors at predicting the structure of octameric Aep1. γ-CD showed unique suitability as an 8-subunit adapter in octameric Aep1, enabling the discrimination of ß-nicotinamide mononucleotide.