Nanopore Detection Using Supercharged Polypeptide Molecular Carriers

Area of Science:

• Biophysics
• Molecular Biology
• Analytical Chemistry

Background:

• Single-molecule analysis of proteins is crucial for understanding biological processes and diseases, especially for low-abundance targets.
• Nanopore sensing offers label-free detection of single proteins, with applications in biomarker screening and drug discovery.
• Current nanopore sensing faces challenges in controlling protein translocation and correlating structure/function with readouts.

Purpose of the Study:

• To develop a method for enhanced control and detection of proteins using nanopore sensing.
• To investigate the use of genetically fused supercharged unstructured polypeptides (SUPs) as molecular carriers for proteins.
• To overcome spatiotemporal limitations in current single-molecule protein analysis.

Main Methods:

• Genetically fusing supercharged unstructured polypeptides (SUPs) with target proteins.
• Utilizing cationic SUPs to modulate protein translocation through nanopores via electrostatic interactions.
• Analyzing nanopore current signals to differentiate proteins based on size and shape.

Main Results:

• SUPs significantly slow down protein translocation through nanopores.
• Characteristic subpeaks in nanopore current allow differentiation of individual proteins.
• Demonstrated a viable method for controlling molecular transport at the single-molecule level.

Conclusions:

• Genetically engineered SUPs serve as effective molecular carriers for enhanced nanopore detection of proteins.
• This approach facilitates single-molecule protein differentiation and interaction studies.
• SUPs offer a promising strategy to overcome limitations in nanopore sensing for biological applications.