Single-molecule protein identification by sub-nanopore sensors.

Area of Science:

• Biophysics
• Analytical Chemistry
• Proteomics

Background:

• Top-down mass spectrometry offers intact protein identification but lacks sensitivity due to low ion counts.
• Nanopore technology excels at single-molecule detection but is primarily used for DNA sequencing.
• A need exists for highly sensitive methods for single-molecule protein identification.

Purpose of the Study:

• To investigate the potential of sub-nanopore technology for single-molecule protein identification (SMPI).
• To develop and validate an algorithm for analyzing electrical current blockade signals (nanospectra) generated by protein translocation.
• To assess the feasibility of using this method for protein identification in complex biological samples.

Main Methods:

• Utilized sub-nanopore devices to translocate denatured, linearly charged proteins.
• Developed an algorithm to identify characteristic electrical current blockade signals (nanospectra) during protein translocation.
• Analyzed identification p-values to determine the accuracy and sensitivity of the method.

Main Results:

• Demonstrated that sub-nanopore technology can generate distinct electrical signals for individual proteins.
• The developed algorithm successfully identified proteins based on their nanospectra.
• Current technology shows sufficient capability for matching nanospectra against small protein databases, such as those from bacterial proteomes.

Conclusions:

• Sub-nanopore technology holds significant promise for highly sensitive single-molecule protein identification.
• The developed nanospectrum analysis algorithm enables accurate protein identification from translocation events.
• This approach is a viable method for protein identification in applications like bacterial proteome analysis.