ABSTRACT
Membrane proteins are membrane-embedded nanomachines that fulfill key functions such as energy conversion, solute transport, secretion, and signal transduction. The lack of structural information is related to the instability of membrane proteins in a detergent-solubilized state, making the growth of three-dimensional (3D) crystals
difficult. Two-dimensional (2D) crystals of purified membrane proteins reconstituted in the presence of lipids provide an environment close to the native one and allow the structure and function of membrane proteins to be assessed. To this end, electron crystallography is used providing 3D information at the atomic level. Atomic force microscopy allows the surface of membrane proteins to be studied at subnanometer resolution, giving information about their conformational variability that cannot be assessed by crystallographic methods. In addition, atomic force microscopy is the ideal method to directly image the topography of the native membrane. This chapter presents methods to grow 2D crystals, to record them by cryoelectron microscopy, and to extract the structural information by digital image processing. Atomic force microscopy techniques are discussed and relevant results presented. The chapter is intended to provide a compact description of the current possibilities offered by these methods, and their power is demonstrated by the examples selected.
