Section 4.2.4. The `small amphiphile concept'

From the arguments and observations presented above, it is evident that the size and shape of the detergent micelle are very important in membrane-protein crystallization. Detergent micelles can be made smaller (and their curvatures changed) when small amphiphilic molecules like heptane-1,2,3-triol are added (Timmins et al., 1991; Gast et al., 1994). These compounds form mixed micelles with detergents. When 10% (w/v) heptane-1,2,3-triol is added to 1% solutions of N,N-dimethyldodecylamine-N-oxide in water, the number of detergent molecules per micelle decreases from 69 to 34, ∼23 heptane-1,2,3-triol molecules are incorporated and the radius of the micelle is reduced from 20.9 to 16.9 Å (Timmins et al., 1994). This so-called `small amphiphile concept' has been used successfully to crystallize bacterial photosynthetic reaction centres (Michel, 1982, 1983; Buchanan et al., 1993), bacterial light-harvesting complexes (Michel, 1991; Koepke et al., 1996) and other membrane proteins (see Table 4.2.1.1). The light-harvesting complexes from the purple bacterium Rhodospirillum molischianum yield an astonishing number of different crystal forms, but only one diffracts to high resolution. A large amount of heptane-1,2,3-triol had to be added to obtain this crystal form. As a result, heptane-1,2,3-triol itself reached supersaturation during crystallization of the protein. Normally, the protein crystallizes first and heptane-1,2,3-triol later. When heptane-1,2,3-triol crystals form, the protein crystals crack and eventually redissolve. A likely explanation is that the concentration of solubilized heptane-1,2,3-triol is reduced when it forms crystals and the mixed detergent–heptane-1,2,3-triol micelles lose heptane-1,2,3-triol and take up detergent molecules from solution. The micelles expand and the crystals crack. This behaviour caused severe problems when searching for heavy-atom derivatives. Small amphiphiles work well with the N,N-dimethylalkylamine-N-oxides and alkylglucopyranosides as detergents, but not with alkylmaltosides. The most successful small amphiphiles are heptane-1,2,3-triol and benzamidine.