Crystallization of bacteriorhodopsin from bicelle formulations at room temperature

We showed previously that high-quality crystals of bacteriorhodopsin (bR) from Halobacterium salinarum can be obtained from bicelle-forming DMPC/CHAPSO mixtures at 37degreesC. As many membrane proteins are not sufficiently stable for crystallization at this high temperature, we tested whether the bicelle method could be applied at a lower temperature. Here we show that bR can be crystallized at room temperature using two different bicelle-forming compositions: DMPC/CHAPSO and DTPC/CHAPSO. The DTPC/CHAPSO crystals grown at room temperature are essentially identical to the previous, twinned crystals: space group P2(1) with unit cell dimensions of a = 44.7 Angstrom, b = 108.7 Angstrom, c = 55.8 Angstrom, beta = 113.6degrees. The room-temperature DMPC/CHAPSO crystals are untwinned, however, and belong to space group C222(1) with the following unit cell dimensions: a = 44.7 Angstrom, b = 102.5 Angstrom, c = 128.2 Angstrom. The bR protein packs into almost identical layers in the two crystal forms, but the layers stack differently. The new untwinned crystal form yielded clear density for a previously unresolved CHAPSO molecule inserted between protein subunits within the layers. The ability to grow crystals at room temperature significantly expands the applicability of bicelle crystallization.