HIGH-RESOLUTION ATOMIC FORCE MICROSCOPY OF NATIVE MEMBRANES - Life at the Nanoscale: Atomic Force Microscopy of Live Cells - page 22

to obtain an analysable sample, including overexpression, solubilization,
puriication and crystallization. Any of these steps may represent a severe
experimental bottleneck. AFM, in contrast, while restricted to a topo-
graphical analysis of the sample at a relatively low lateral resolution (~10
Å), provides such a high SNR that single molecules can be structurally
analysed. 2 This is the feature that allows the structural analysis of membrane
proteins in a native membrane packed with several molecule species.
Figure 2.1. Comparison of techniques used to analyse membrane protein structure,
taking into consideration the number of molecules to be analysed to obtain wished
or achievable structural information, the corresponding required biochemical proce-
dures and an estimated amount of protein needed to obtain an analysable sample.
As detailed in Fig. 2.2 , AFM has a second trump that renders the
technique particularly attractive for the assessment of structure-function
relationships. AFM measures these relationships under physiological
conditions, i.e., in a physiological buffer, at room temperature and under
atmospheric pressure. This is a major breakthrough compared with other
high-resolution techniques that often demand vacuum or low-temperature
conditions. Besides this technical advantage, the above-mentioned
extraordinary SNR of AFM allows for studying more native samples than
what any other techniques can survey ( Fig. 2.2 ) . Ultimately, biologists
would like to see membrane proteins in native membranes directly on a
live cell. This has been impossible to date because of physical reasons, such