Resolving Isomeric Posttranslational Modifications Using a Biological Nanopore as a Sensor of Molecular Shape

Area of Science:

• Biochemistry
• Nanotechnology
• Proteomics

Background:

• Posttranslational modifications (PTMs) are critical in diseases like cancer.
• Current PTM detection methods (mass spectrometry, immunoassays) lack selectivity and specificity.
• Accurate PTM identification requires precise localization of modifications on peptides.

Purpose of the Study:

• To develop a novel method for differentiating peptides based on PTM position.
• To utilize protein nanopore technology for label-free PTM analysis.
• To overcome limitations of existing PTM detection techniques.

Main Methods:

• Employed a protein nanopore to analyze peptides derived from human histone H4.
• Differentiated peptides of identical mass with varying acetylation and methylation on lysine residues.
• Utilized molecular dynamics simulations to understand the pore's electric field sensitivity to molecular shape.

Main Results:

• Successfully differentiated peptides based on PTM location using a protein nanopore.
• Demonstrated label-free detection of PTMs and their precise positions.
• Identified a nonuniform electric field within the pore as key to shape-sensing sensitivity.

Conclusions:

• Protein nanopore technology enables label-free, high-throughput characterization of protein isoforms.
• This shape-sensing principle offers a versatile approach for PTM analysis.
• The method provides a promising alternative for diagnosing diseases linked to PTMs.