MEMBRANE PERMEABILIZING ACTIVITY OF PATHOGENESIS-RELATED PROTEIN LINUSITIN FROM FLAX SEED

Linusitin is a 25-kDa pathogenesis-related (PR) protein of class 5 iso lated from flax seeds. It has been proposed that PR-5 proteins exert t heir antifungal activity by permeabilizing fungal membranes. Using a f luorescent method that has been used for antimicrobial proteins other than PR proteins, we tested this hypothesis. The method is based on th e release of the dye calcein from the inside of either small or large unilamellar lipid vesicles. The amount of calcein release induced by t he protein was studied as a function of protein and lipid concentratio n and of membrane composition. All the results could be accounted for with an available statistical model, The model predicts that calcein r elease from the vesicles is a result of the tetrameric protein aggrega tion. Whether this aggregate corresponds to a transmembrane pore or to another protein complex that perturbs the membrane remains to be esta blished. The lipid composition of the vesicles affected the permeabili zation activity of linusitin. An increase of activity was observed by increasing the content of negatively charged phospholipids in the vesi cles, inclusion of sterols into the membrane, or decreasing the pH of the solution. The possible roles of the observed changes in permeabili zing activity in actual plant-fungus interactions are discussed.