Nanomechanical properties of proteins and membranes depend on loading rate and electrostatic interactions

Knowing the dynamic mechanical response of tissue, cells, membranes, proteins, nucleic acids, and carbohydrates to external perturbations is important to understand various biological and biotechnological problems. Atomic force microscopy (AFM)-based approaches are the most frequently used nanotechnologies to determine the mechanical properties of biological samples that range in size from microscopic to (sub)nanoscopic. However, the dynamic nature of biomechanical properties has barely been addressed by AFM imaging. In this work, we characterizethe viscoelastic properties of the native light-driven proton pump bacteriorhodopsin of the purple membrane of Halobacterium salinarum. Using force-distance curve (F-D)-based AFM we imaged purple membranes while force probing their mechanical response over a wide range of loading rates (from ∼0.5 to 100 μN/s). Our results show that the mechanical stiffness of protein and membrane increases with the loading rate up to a factor of 10 (from ∼0.3 to 3.2 N/m). In addition, the electrostatic repulsion between AFM tip and sample can alter the mechanical stiffness measured by AFM up to ∼60% (from ∼0.8 to 1.3 N/m).These findings indicate that the mechanical response of membranes and proteins and probably of other biomolecular systems should be determined at different loading rates to fully understand their properties.