Quantitative Imaging of the Electrostatic Field and Potential Generated by a Transmembrane Protein Pore at Subnanometer Resolution

Elucidating the mechanisms by which proteins translocate small molecules and ions through transmembrane pores and channels is of great interest in biology, medicine, and nanotechnology. However, the characterization of pore forming proteins in their native state lacks suitable methods that are capable of high-resolution imaging (similar to 1 nm) while simultaneously mapping physical and chemical properties. Here we report how force distance (FD) curve-based atomic force microscopy (AFM) imaging can be applied to image the native pore forming outer membrane protein F (OmpF) at subnanometer resolution and to quantify the electrostatic field and potential generated by the transmembrane pore. We further observe the electrostatic field and potential of the OmpF pore switching on and off in dependence of the electrolyte concentration. Because electrostatic field and potential select for charged molecules and ions and guide them to the transmembrane pore the insights are of fundamental importance to understand the pore function. These experimental results establish FD-based AFM as a unique tool to image biological systems to subnanometer resolution and to quantify their electrostatic properties.