A flexible nanoarray approach for the assembly and probing of molecular complexes

Area of Science:

• Biophysics
• Materials Science
• Molecular Biology

Background:

• Single-molecule force spectroscopy, including atomic force microscopy (AFM), relies on effective molecular immobilization.
• Studying molecular interactions requires precise control over the positioning and interaction of molecules.

Purpose of the Study:

• To introduce a novel flexible nanoarray (FNA) technique for probing molecular interactions.
• To measure the interaction between internally immobilized amyloid β peptides using AFM.
• To assess the capability of the FNA technique for reversible and multiple intermolecular interaction analyses.

Main Methods:

• Synthesized a flexible nanoarray (FNA) tether using DNA synthesis chemistry with incorporated peptide anchoring points.
• Utilized atomic force microscopy (AFM) force spectroscopy to pull the FNA tether and measure peptide interactions.
• Immobilized the FNA tether to an AFM substrate and tip using covalent (maleimide-thiol) and non-covalent (biotin-streptavidin) linkages.

Main Results:

• Successfully measured amyloid β peptide dimer formation and rupture using the FNA-AFM approach.
• Identified unique rupture fingerprints (position and force) for the peptide dimers on force curves.
• Demonstrated reversible association and dissociation of peptide monomers within the same molecular complex.
• Computational analysis confirmed that FNA tether flexibility allows natural dimer formation.

Conclusions:

• The flexible nanoarray (FNA) technique provides a novel and effective method for studying molecular interactions.
• FNA enables precise immobilization and measurement of interactions between internally tethered molecules like amyloid β peptides.
• The FNA technique supports reversible experiments, allowing analysis of multiple intermolecular interactions within a single complex.