Membrane-protein crystallization in cubo: temperature-dependent phase behaviour of monoolein-detergent mixtures.

Abstract

The lipidic cubic phase of monoolein has proved to be a matrix well suited to the production of three-dimensional crystals of membrane proteins. It consists of a single continuous bilayer, which is contorted in three-dimensional space and separates two distinct water channels. It has previously been proposed that on the addition of precipitants, membrane proteins embedded in the cubic phase migrate through the matrix to nucleation sites and that this process is dependent upon the stability of the lipidic cubic phase. Here, the effect of detergent type (C(8)-C(12) glucosides, C(8)-C(12) maltosides and C(7) thioglucoside) and concentration (1-3x the critical micelle concentration; CMC) on cubic phase stability are reported in the form of the temperature-dependent phase behaviour (268-313 K) in 40% aqueous solution. The results are tabulated to show the best monoolein (MO)-detergent mixtures, mixing temperatures and crystallization temperatures identified. Monoolein-detergent mixtures suited for low-temperature in cubo crystallization of temperature-sensitive proteins are also reported for the first time. These mixtures can be prepared at low temperatures (mixed at <or=288 K) and remain stable at 277 K for a period of at least one Month. They include MO-heptyl thioglucoside (1x and 3x CMC), MO-nonyl glucoside (3x CMC), MO-octyl maltoside (3x CMC), MO-nonyl maltoside (1x CMC) and MO-decyl maltoside (1x CMC).