Single molecule DNA origami nanoarrays with controlled protein orientation

Area of Science:

• Biophysics
• Nanotechnology
• Molecular Biology

Background:

• Precise nanoscale organization of biomolecules is crucial for developing advanced molecular devices.
• Controlling both position and orientation of single molecules remains a significant challenge.

Purpose of the Study:

• To fabricate nanoarrays of individual proteins with controlled orientation on solid substrates.
• To demonstrate a general strategy for organizing functional biomolecules with single-molecule resolution.

Main Methods:

• Utilized focused ion beam lithography to create nanopatterned surfaces with nanoapertures.
• Employed functionalized DNA origami structures for selective immobilization of biomolecules.
• Engineered proteins with new chemistry for residue-specific linkage to DNA origami.

Main Results:

• Achieved 88% immobilization of DNA origami structures in nanoapertures.
• Demonstrated high occupancy of single DNA origami units (85%) with minimal double occupancy (3%).
• Successfully assembled and organized orientation-controlled proteins on DNA origami within nanoarrays, exhibiting single-molecule traces.

Conclusions:

• Developed a robust method for fabricating protein nanoarrays with precise spatial and orientational control.
• The strategy is broadly applicable for single-molecule investigations in defined nanoarray configurations.
• Enables the development of next-generation biomolecular devices and assays requiring controlled molecular organization.