Degradation of membrane-bound ganglioside GM1 - Stimulation by bis(monoacylglycero)phosphate and the activator proteins SAP-B and GM2-AP

According to our hypothesis (Furst, W,, and Sandhoff, K, (1992) Biochim, Bi ophys. Acta 1126, 1-16) glycosphingolipids of the plasma membrane are diges ted after endocytosis as components of intraendosomal and intralysosomal ve sicles and membrane structures. The lysosomal degradation of glycosphingoli pids with short oligosaccharide chains by acid exohydrolases requires small , non-enzymatic cofactors, called sphingolipid activator proteins (SAPs), A total of five activator proteins have been identified as follows: namely t he saposins SAP-A, -B, -C, and -D, which are derived from the single chain SAP-precursor protein (prosaposin), and the GM2 activator protein. A defici ency of prosaposin results in the storage of ceramide and sphingolipids wit h short oligosaccharide head groups. The loss of the GM2 activator protein blocks the degradation of the ganglioside GM2. The enzymatic hydrolysis of the ganglioside GM1 is catalyzed by beta -galactosidase, a water-soluble ac id exohydrolase, The lack of ganglioside GM1 accumulation in patients suffe ring from either prosaposin or GM2 activator protein deficiency has led to the hypothesis that SAPs are not needed for the hydrolysis of the gangliosi de GM1 in vivo, In this study we demonstrate that an activator protein is r equired for the enzymatic degradation of membrane-bound ganglioside GM1 and that both SAP-B and the GM2 activator protein significantly enhance the de gradation of the ganglioside GM1 by acid beta -galactosidase in a liposomal , detergent-free assay system. These findings offer a possible explanation for the observation that no storage of the ganglioside GM1 has been observe d in patients with either isolated prosaposin or isolated GM2 activator def iciency. We also demonstrate that anionic phospholipids such as bis(monoacy lglycero)phosphate and phosphatidylinositol, which specifically occur in in ner membranes of endosomes and in lysosomes, are essential for the activato r-stimulated hydrolysis of the ganglioside GM1. Assays utilizing surface:pl asmon resonance spectroscopy showed that bis(monoacylglycero)phosphate incr eases the binding of both beta -galactosidase and activator proteins to sub strate-carrying membranes.