A DNA-based molecular clamp for probing protein interactions and structure under force

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Cellular mechanotransduction is crucial for biological processes and diseases.
• Studying protein mechanics under controlled tension is challenging due to a lack of suitable methods.
• Force-induced protein conformational changes are key to regulating cellular functions.

Purpose of the Study:

• To develop a novel DNA-based device for applying defined mechanical loads to proteins.
• To investigate tension-induced protein conformational changes and ligand binding.
• To enable biochemical and structural studies of proteins under mechanical stress.

Main Methods:

• Development of a DNA-based tension device utilizing DNA strand transitions.
• Attachment of protein fragments (talin rod domain) to the DNA device.
• Application of negative-stain electron microscopy for structural analysis.
• Utilization of pull-down assays to assess protein-ligand interactions.

Main Results:

• The DNA device successfully applied programmable tension to the talin rod domain.
• Electron microscopy confirmed programmable extension of the protein.
• Pull-down assays demonstrated tension-induced binding of ARPC5L and vinculin to cryptic sites within talin.
• The talin fragment exhibited tension-induced conformational changes.

Conclusions:

• The developed DNA clamp is a valuable tool for biochemical studies of protein mechanics.
• This method allows for the investigation of force-driven protein conformational changes.
• The findings open new avenues for structural analysis of proteins under mechanical load.
• This technology has implications for understanding mechanotransduction in health and disease.