Section 4.2.2. Principles of membrane-protein crystallization

There are two principal types of membrane-protein crystals (Michel, 1983). First, one can think of forming two-dimensional crystals in the planes of the membrane and stacking these two-dimensional crystals in an ordered way with respect to up and down orientation, rotation and translation. This membrane-protein crystal type (`type I') is attractive, because it contains the membrane proteins in their native environment. It should even be possible to study lipid–protein interactions. Crystals of bacteriorhodopsin of this type have been obtained either upon slow removal of the detergent by dialysis at high ionic strength (Henderson & Shotton, 1980), or by a novel approach using lipidic bicontinuous cubic phases (Landau & Rosenbusch, 1996; Pebay-Peyroula et al., 1997; see also below). Alternatively, one can try to crystallize the membrane protein with the detergents still bound in a micellar manner. These crystals are held together via polar interactions between the polar surfaces of the membrane proteins. The detergent plays a more passive, but still critical, role. Such `type II' crystals look very much like crystals of soluble globular proteins. The same crystallization methods and equipment as for soluble globular proteins (see Chapter 4.1 ) can be used. However, the use of hanging drops is sometimes difficult, because the presence of detergents leads to a lower surface tension of the protein solution. Intermediate forms between type I and type II crystals are feasible, e.g. by fusion of detergent micelles.

The use of detergent concentrations just above the CMC of the respective detergent is recommended in order to prevent complications caused by pure detergent micelles. Unfortunately, the CMC is not constant. Normally, the CMC provided by the vendor has been determined in water at room temperature. A compilation of potentially useful detergents, their CMCs and their molecular weights is presented in Table 4.2.2.1. The CMC is generally lower at high ionic strength and at high temperatures. The presence of glycerol and similar compounds, as well as that of chaotropic agents (Midura & Yanagishita, 1995), also influences (decreases) the CMC.