Comparative investigation of PdR by usual and ultrafine atomic force microscopy

Abstract

Atomic force microscopy is one of the most perspective methods for determination of the structure of proteins and their complexes. The vertical resolution of this method is about 0.1 nm, which is close to X-ray resolution. At the same time, the lateral resolution, determining by broadening effect of a standard AFM probe, is not very high—about 20–50 nm, depending on probe geometry. Naturally, the probe tip broadening effect leads to substantial enhancement of measured protein volume. In this study, a comparative analysis of sizes of the protein putidaredoxin reductase (PdR) obtained by the use of two AFM probe types, standard and supersharp, was undertaken. Usage of standard probes enabled to correctly measure the height of PdR while the volume of this protein was measured with considerably (more than one order) enhancement. It was shown that application of supersharp AFM probes results in the lowering of measured protein height; at the same time, the measured protein volume is more exact and appears to be close to RSA data. Therefore, to obtain exact data on protein volume and height, these two parameters should be measured by use of both supersharp probes and standard geometry probes.