Single Molecule Study of Hydrogen Bond Interactions Between Single Oligonucleotide and Aerolysin Sensing Interface

Area of Science:

• Biophysics
• Nanotechnology
• Molecular Biology

Background:

• Aerolysin nanopore exhibits sensitive detection of single oligonucleotides via electrochemical signals.
• Stronger aerolysin nanopore-oligonucleotide interactions lead to longer signal duration, enhancing sensitivity.

Purpose of the Study:

• Investigate hydrogen bond interactions to optimize aerolysin nanopore sensing performance.
• Understand the role of specific amino acid residues in nanopore-analyte interactions.

Main Methods:

• Site-directed mutagenesis was used to alter single amino acid residues near K238.
• Mutations K238C and K238Y were created and tested for their effect on analyte translocation.
• Electrochemical signals and translocation duration times were measured for wild-type (WT) and mutant aerolysin nanopores.

Main Results:

• The K238C mutation resulted in a six-fold increase in (dA)4 translocation duration compared to WT aerolysin.
• The K238Y mutation shortened translocation time, indicating faster analyte movement and reduced sensitivity.
• Hydrogen bonds enhance analyte-nanopore interactions within the confined space, rather than dominating translocation dynamics.

Conclusions:

• Specific mutations, like K238C, can significantly enhance aerolysin nanopore sensitivity for oligonucleotide detection.
• Hydrogen bonding plays a crucial role in modulating analyte-nanopore interactions for improved sensing.
• These findings provide insights for rational design of nanopore sensing mechanisms for biomolecules.