The Saposin-like Domain of the Plant Aspartic Proteinase Precursor Is a Potent Inducer of Vesicle Leakage

A unique feature of plant aspartic proteinase precursors is the presence of an internal domain, known as plant-specific insert, whose function is not completely understood. The three-dimensional structure of the plant-specific insert resembles that of saposin-like proteins, a group of lipid-binding proteins involved in a variety of physiological processes. Here we show that recombinant plant-specific insert is able to interact with phospholipid vesicles and to induce leakage of their contents in a pH- and lipid-dependent manner. The leakage activity is higher at pH 4.5 and requires the presence of acidic phospholipids such as phosphatidylserine. To determine whether the same effect could be observed when the plant-specific insert is part of the precursor form, procardosin A and a mutant form lacking this specific domain were produced and characterized. Procardosin A displays a similar activity profile, whereas the mutant without the plant-specific insert shows only residual activity. These findings indicate that the plant-specific insert domain of plant aspartic proteinases mediates an interaction of their precursors with phospholipid membranes and induces membrane permeabilization. It is therefore possible that the plant-specific insert, alone or in conjunction with the proteolytic activity of plant aspartic proteinases, may function either as a defensive weapon against pathogens or in late autolysis of plant cells.