E. coli outer membrane protein T (OmpT) nanopore for peptide sensing

Area of Science:

• Biophysics
• Nanotechnology
• Biochemistry

Background:

• Sensitive peptide detection is crucial for diagnosing diseases linked to peptide biomarkers.
• Nanopore sensing offers a cost-effective, high-throughput, and scalable method for peptide analysis.
• Omptins proteins, capable of forming β-barrel pores, show promise for nanopore biosensor development.

Purpose of the Study:

• To investigate the channel properties of E. coli OmpT.
• To develop OmpT as a functional nanopore biosensor for peptide detection.
• To assess OmpT's specificity in peptide detection against nucleic acids.

Main Methods:

• Construction and characterization of the OmpT biological nanopore channel.
• Measurement of channel conductance in a 500 mM NaCl buffer.
• Analysis of OmpT gating phenomena under specific voltage conditions.
• Development and testing of the OmpT nanopore as a peptide biosensor.

Main Results:

• The constructed OmpT nanopore exhibited a conductance of 1.49 nS.
• A distinct three-step gating phenomenon was observed for OmpT at negative voltages exceeding 100 mV.
• The OmpT biosensor successfully detected peptides, showing no interference from single-stranded DNA (ssDNA) and deoxynucleotide triphosphates (dNTPs).

Conclusions:

• The engineered OmpT protein functions as a viable biological nanopore.
• The OmpT nanopore biosensor demonstrates potential for specific and sensitive peptide detection.
• This research introduces a novel strategy for peptide detection utilizing protease-specific binding within a nanopore system.