Analysis of the multiple forms of Gaucher spleen sphingolipid activator protein 2.
Gaucher spleen sphingolipid activator protein 2 was fractionated into concanavalin A binding- and non-binding fractions. These fractions each contained several bands on non-denaturing polyacrylamide gel electrophoresis (PAGE). The two fractions were further fractionated by electroblotting the proteins from preparative gels onto nitrocellulose, staining with Ponceau S to locate the bands of protein and then eluting the protein components from the nitrocellulose. A total of ten fractions, each containing only one or two major components, was collected. All of these subfractions activated beta-glucocerebrosidase and sphingomyelinase and most subfractions also activated beta-galactocerebrosidase. The structural relationship of the bands was investigated using endoglycosidase digestions. The results indicated that the two bands with the fastest mobility on non-denaturing PAGE did not contain any carbohydrate. The remaining bands showed only limited or partial digestion with endoglycosidase H and endoglycosidase D, but were readily hydrolysed with endoglycosidase F. The products of these digestions included bands with similar mobilities to the non-carbohydrate containing bands.
Paton
B C
BC
Department of Chemical Pathology, Adelaide Children's Hospital Incorp., South Australia.
Poulos
A
A
D003208:Concanavalin A; D004591:Electrophoresis, Polyacrylamide Gel; D005698:Galactosylceramidase / Q000378:metabolism; D005776:Gaucher Disease / Q000378:metabolism*; D005962:Glucosylceramidase / Q000378:metabolism; D006023:Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D006026:Glycoside Hydrolases; D006801:Humans; D049231:Saposins; D049128:Sphingolipid Activator Proteins; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism; D013154:Spleen / Q000032:analysis*
D006023:Glycoproteins; C483345:PSAP protein, human; D049231:Saposins; D049128:Sphingolipid Activator Proteins; D003208:Concanavalin A; D013108:Sphingomyelin Phosphodiesterase; D006026:Glycoside Hydrolases; D005962:Glucosylceramidase; D005698:Galactosylceramidase