Hydrodynamic effects in fast AFM single-molecule force measurements.

Area of Science:

• Biophysics
• Materials Science
• Nanotechnology

Background:

• Atomic force microscopy (AFM) is crucial for measuring single-molecule forces, like protein unfolding and receptor-ligand bond rupture.
• Understanding energy landscapes requires probing dynamic strengths at various force loading rates, typically achieved by varying AFM cantilever pulling speeds.
• High pulling speeds (above 10 microm/s) are desirable for comprehensive kinetic analysis but are limited by significant hydrodynamic drag forces on the AFM cantilever.

Purpose of the Study:

• To develop and present a method for accurately evaluating AFM force measurements at extended pulling speeds.
• To overcome the limitations imposed by hydrodynamic drag forces in high-speed AFM single-molecule force spectroscopy.
• To enable a more complete investigation of kinetic properties by extending the measurable range of force loading rates.

Main Methods:

• Development of a theoretical approach to incorporate hydrodynamic effects into AFM data analysis.
• Experimental validation using mechanical unfolding of a multi-domain protein.
• Measurements conducted at a high pulling speed of 30 microm/s to demonstrate the method's efficacy.

Main Results:

• The proposed approach allows for the correct evaluation of AFM force measurements across a wider range of pulling speeds.
• Hydrodynamic effects, previously limiting high-speed measurements, can now be accurately accounted for.
• Successful experimental demonstration at 30 microm/s confirms the method's applicability.

Conclusions:

• The developed method significantly extends the utility of AFM for single-molecule force spectroscopy by enabling accurate measurements at higher loading rates.
• This advancement facilitates a deeper understanding of molecular dynamics and energy landscapes.
• The findings pave the way for more comprehensive studies of biomolecular mechanics and interactions.