Large scale purification of an almond oleosin using an organic solvent procedure

Oleosins are small proteins which are specific to oil-bodies, the plant organelles specialized in oil storage. These proteins are thought to stabilize the structure and to prevent the oil droplets from coalescing. Since oleosins are insoluble in water, they are usually purified either as aggregates in water or as polypeptides solubilized with SDS. These conditions raise major problems for performing biophysical studies, however. We have taken advantage of the solubility of almond oleosin in chloroform to purify it by differential extraction using solvent mixtures. Detergent-free oleosin was purified to apparent homogeneity with a purification yield of 20 % using almond meal as starting material. An oil-body suspension was reconstituted by sonicating the solvent-purified oleosin with phospholipids and triacylglycerols. Protease protection experiments suggest that oleosins are inserted into the reconstituted oil-body as they are in native organelles. This simple purification procedure allows to obtain a detergent-free solution of oleosin easy to handle which can be used for oil-body reconstitution and biophysical studies by the Wilhemly plate method. (C) 2001 Editions scientifiques et medicales Elsevier SAS.