In this study we demonstrate that an activator protein is required for the enzymatic degradation of membrane-bound ganglioside GM1 and that both SAP-B and the GM2 activator protein significantly enhance the degradation of the ganglioside GM1 by acid -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 observed in patients with either isolated prosaposin or isolated GM2 activator deficiency. We also demonstrate that anionic phospholipids such as bis(monoacylglycero)phosphate and phosphatidylinositol, which specifically occur in inner membranes of endosomes and in lysosomes, are essential for the activator-stimulated hydrolysis of the ganglioside GM1. Assays utilizing surface plasmon resonance spectroscopy showed that bis(monoacylglycero)phosphate increases the binding of both -galactosidase and activator proteins to substrate-carrying membranes.
A rare inherited disease, multiple sulfatase deficiency, is attributed to a defect in a posttranslational protein modification which is essential for the catalytic activity of all known sulfatases. Structure analysis of arylsulfatase A, the enzyme that cleaves sulfatide 1, shows that the modification of a cysteine residue into a formylglycine residue is essential for catalytic activity.
Eine Erbkrankheit, die multiple Sulfatasedefizienz, wird auf einen Fehler bei einer posttranslationalen Proteinmodifikation zurückgeführt. Strukturuntersuchungen an der Arylsulfatase A, die das Sulfatid 1 spaltet, ergaben, daß diese neuartige Modifikation, die Bildung eines Formylglycinrestes aus einem Cysteinrest, für die katalytische Aktivität der Sulfatasen erforderlich ist.
The interaction between glycosphingolipids and recombinant human GM2-activator was studied in a microwell binding assay. A-series gangliosides like GM3, GM2 and GM1 were strongly bound by the recombinant human GM2 activator. A weak binding was observed to GD1b and sulfatide, while neutral glycolipids were not bound. Optimal binding occurred at pH 4.2 and was inhibited by increasing concentrations of citrate buffer and NaCl. In contrast with these in vitro results the recombinant human GM2-activator is able to restore the degradation of GA2 in fibroblasts from patients with the AB variant of GM2 gangliosidosis in vivo.
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