Resolving Sulfation Posttranslational Modifications on a Peptide Hormone using Nanopores.

Area of Science:

• Biochemistry
• Molecular Biology
• Nanotechnology

Background:

• Post-translational modifications (PTMs) on peptide hormones are vital for receptor recognition and biological function.
• Tyrosine sulfation is critical for plant peptide hormone activity, influencing growth and development.
• Current methods for detecting and localizing sulfotyrosine face significant technical hurdles.

Purpose of the Study:

• To demonstrate the capability of nanopore technology for sensitive, single-molecule detection of PTMs on peptide hormones.
• To accurately identify and distinguish sulfation from phosphorylation on tyrosine residues of plant peptide hormones.
• To assess the accuracy of nanopore measurements in determining the presence and position of PTMs on multiple tyrosine residues.

Main Methods:

• Translocation of post-translational modification variants of the plant pentapeptide hormone phytosulfokine (PSK) through a nanopore.
• Sensitive, single-molecule detection of PTMs by analyzing changes in electrical current during translocation.
• Distinguishing between sulfation and phosphorylation modifications on specific tyrosine residues.

Main Results:

• Accurate identification of both sulfation and phosphorylation on the two tyrosine residues of PSK.
• Sulfation was clearly detected and distinguished from phosphorylation on the same residue with >90% accuracy.
• The presence or absence of PTMs on adjacent tyrosine residues was determined with >96% accuracy.

Conclusions:

• Nanopore measurements offer extraordinary sensitivity for detecting protein PTMs at the single-molecule level.
• This technology provides a powerful tool for identifying position-specific sulfation on peptide hormones.
• The findings suggest broad applicability for nanopore analysis in identifying various protein PTMs.