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Probing RNA Conformations Using a Polymer-Electrolyte Solid-State Nanopore.

Chalmers Chau1,2,3, Fabio Marcuccio2,3, Dimitrios Soulias2,3

  • 1School of Molecular and Cellular Biology and Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds LS2 9JT, U.K.

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|October 24, 2022
PubMed
Summary
This summary is machine-generated.

This study introduces a novel polymer-electrolyte nanopore system enabling biomolecule analysis under physiological conditions. The optimized system successfully distinguished conformations of Chikungunya virus RNA fragments.

Keywords:
DNAPEGRNAnanopipettenanoporepolymer−electrolytesingle molecule

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Area of Science:

  • Biophysics
  • Nanotechnology
  • Molecular Biology

Background:

  • Solid-state nanopore systems analyze biomolecules at single-molecule resolution.
  • High salt concentrations are typically required, limiting analysis under physiological conditions.
  • Biomolecular conformation is sensitive to electrolyte composition.

Purpose of the Study:

  • To develop a polymer-electrolyte solid-state nanopore system for biomolecular analysis under physiological conditions.
  • To investigate the effect of polymer-electrolyte baths on analyte translocation dynamics.
  • To analyze the conformational states of Chikungunya virus (CHIKV) RNA fragments.

Main Methods:

  • Implementation of a solid-state nanopore system using alkali metal halide salts in 50% poly(ethylene) glycol (PEG).
  • Optimization of salt composition, identifying Cesium Bromide (CsBr) as optimal for signal enhancement.
  • Probing CHIKV RNA genome fragments (∼300 to ∼2000 nt) using the developed nanopore system.

Main Results:

  • The polymer-electrolyte bath significantly influences analyte translocation dynamics.
  • CsBr in PEG provided the largest signal enhancement for nanopore analysis.
  • The system successfully differentiated conformations of co-transcriptionally folded and natively refolded CHIKV RNA fragments.
  • Fingerprinting of CHIKV RNA fragments from ∼300 to ∼2000 nt was achieved.

Conclusions:

  • The polymer-electrolyte solid-state nanopore system enables biomolecular conformational analysis under physiologically relevant conditions.
  • This approach overcomes limitations of traditional high-salt nanopore systems.
  • The system holds promise for future structural and conformational studies of individual biomolecules.