Measuring a frequency spectrum for single-molecule interactions with a confined nanopore

Abstract

Nanopore analysis is a powerful technique for single molecule analysis by virtue of its electrochemically confined effects. As a single molecule translocates through the nanopore, the featured ionic current pattern on the time scale contains single molecule characteristics including volume, charge, and conformational properties. Although the characteristics of a single molecule in a nanopore have been written to the featured ionic current, extracting the dynamic information from a complex current trace is still a big challenge. Here, we present an applicable nanopore analysis method employing the Hilbert-Huang Transform (HHT) to study the vibrational features and interactions of a single molecule during the dynamic translocation process through the confined space of a nanopore. The HHT method is specially developed for analyzing nonlinear and non-stationary data that is highly compatible with nanopore data with a high frequency resolution. To provide proof-of-concept, we applied HHT to measure the frequency response for the wild-type (WT) aerolysin and mutant K238E aerolysin nanopores with and without the presence of poly(dA)4, respectively. The energy-frequency-time distribution spectra demonstrate that the biological nanopore contributes greatly to the characteristics of the high frequency component (>2 kHz) in the current recording. Our results suggest that poly(dA)4 undergoes relatively more consistent and confined interactions with K238E than WT, leading to a prolonging of the duration time. Therefore, the characteristics in frequency analysis could be regarded as an "single-molecule ionic spectrum" inside the nanopore, which encodes the detailed behaviours of single-molecule weak interactions.

Detecting topological variations of DNA at single-molecule level

“…We conclude the analysis by pointing out that a recent trend in nanopore signal processing is to use the Hilbert–Huang Transform (HHT) 35 . HHT is a useful empirical tool for analyzing nanopore traces as it is designed to work for signals that are nonstationary and nonlinear (see Supplementary Discussion 2).…”

Time series analysis and long short-term memory (LSTM) network prediction of BPV current density

“…An alternative to STL decomposition that would provide a different view of the constituent patterns making up the current density profile is Empirical Mode Decomposition (EMD) used when applying the Hilbert-Huang Transform (HHT) to nonlinear and non-stationary data. EMD combined with HHT is a timefrequency analysis method that has been used to study single molecule interaction within nanopores, among various other applications 34,35 . EMD is an iterative process that decomposes a signal into individual mono-components called intrinsic mode functions (IMFs), each containing content of a narrow range of frequencies that make up the signal.…”

“…EMD is an iterative process that decomposes a signal into individual mono-components called intrinsic mode functions (IMFs), each containing content of a narrow range of frequencies that make up the signal. The HHT is then applied to 34,35 . Nonetheless, the STL decomposition allowed the two current density profiles to be decomposed in to their constitutive patterns, providing insights that aid the understanding of the performance of the LSTM network discussed in the next section.…”

From current trace to the understanding of confined media

“…[41,43] Recently, the same team has developed an analysis method based on the Hilbert-Huang Transform to study single analytes through the confined space of an Aerolysin channel. [52] Another type of complex blockades analysis is the discrimination between two situations: a single or two macromolecules enter the pore together ( fig. 2(c)).…”