Charge, Diffusion, and Current Fluctuations of Single-Stranded DNA Trapped in an MspA Nanopore

Area of Science:

• Biophysics
• Nanotechnology
• Molecular Biology

Background:

• Single-stranded DNA (ssDNA) translocation through nanopores is crucial for sequencing and diagnostics.
• Understanding ssDNA-nanopore interactions requires characterizing molecular properties like charge and diffusion.

Purpose of the Study:

• To investigate the influence of nucleotide identity on the effective charges and diffusion constants of ssDNA oligonucleotides.
• To explore correlations between these ssDNA properties and nanopore current signals.
• To identify novel sources of current fluctuations during ssDNA-nanopore experiments.

Main Methods:

• Utilizing MspA nanopore system to trap ssDNA oligonucleotides.
• Employing escape time experiments with NeutrAvidin-tethered DNA to measure effective charges and diffusion constants.
• Analyzing current blockages and fluctuations to correlate with ssDNA characteristics.

Main Results:

• Nucleotide identity substantially affects ssDNA effective charges and diffusion constants.
• Oligonucleotide effective charges correlate with current blockages.
• Diffusion constants correlate with DNA-induced current blockage fluctuations.
• An unexpected discrete current level structure in fluctuations was observed, potentially due to NeutrAvidin-MspA interactions.

Conclusions:

• ssDNA sequence composition is a key determinant of its behavior in nanopores.
• Nanopore current measurements provide valuable information about ssDNA charge and diffusion.
• NeutrAvidin-MspA interactions represent a significant factor influencing current fluctuations.