Saposin D solubilizes anionic phospholipid-containing membranes

Saposin (Sap) D is a late endosomal/lysosomal small protein, generated toge ther with three other similar proteins, Sap A, B, and C, from the common pr ecursor, prosaposin. Although the functions of saposins such as Sap B and C are well known (Sap B promotes the hydrolysis of sulfatides and Sap C that of glucosylceramide), neither the physiological function nor the mechanism of action of Sap D are yet fully understood. We previously found that a dr amatic increase of Sap D superficial hydrophobicity, occurring at the low p H values characteristic of the late endosomal/lysosomal environment, trigge rs the interaction of the saposin with anionic phospholipid-containing vesi cles. We have presently found that, upon lipid binding, Sap D solubilizes t he membranes, as shown by the clearance of the vesicles turbidity. The resu lts of gel filtration, density gradient centrifugation, and negative staini ng electron microscopy demonstrate that this effect is due to the transform ation of large vesicles to smaller particles. The solubilizing effect of Sa p D is highly dependent on pH, the lipid/saposin ratio, and the presence of anionic phospholipids; small variations in each of these conditions marked ly influences the activity of Sap D. The present study documents the intera ction of Sap D with membranes as a complex process. Anionic phospholipids a ttract Sap D from the medium; when the concentration of the saposin on the lipid surface reaches a critical value, the membrane breaks down into recom binant small particles enriched in anionic phospholipids. Our results sugge st that the role played by Sap D is more general than promoting sphingolipi d degradation, e.g. the saposin might also be a key mediator of the solubil ization of intralysosomal/late endosomal anionic phospholipidcontaining mem branes.