Studies on a sphingolipid activator protein (SAP-2) in fibroblasts from patients with lysosomal storage diseases, including Niemann-Pick disease Type C.

Sphingolipid activator protein-2 (SAP-2) has been found to stimulate the enzymatic hydrolysis of at least three sphingolipids, glucosylceramide, galactosylceramide and sphingomyelin. Using monospecific antibodies against SAP-2 the level of SAP-2 was determined in cultured skin fibroblasts by rocket immunoelectrophoresis. Extracts from 14 controls had 1.03 +/- 0.28 micrograms cross-reactive material/mg solubilized protein and extracts from 46 patients with Niemann-Pick disease Type C had 1.12 +/- 0.26. Extracts from other lysosomal storage diseases, including Gaucher disease, Krabbe disease and Niemann-Pick disease Types A, B and D, had normal or slightly elevated SAP-2 concentrations, while extracts from patients with I-Cell disease had half normal SAP-2 concentration. When the fibroblast extracts were subjected to sodium dodecylsulfate-polyacrylamide gel electrophoresis followed by electroblotting and immunochemical staining two major SAP-2 bands with estimated molecular weights of 9000 and 10000 were found. Extracts from patients with I-Cell disease showed only a faint higher molecular weight band. Isoelectric focusing followed by electroblotting and immunochemical staining demonstrated no significant difference in the charge of SAP-2 obtained from different cell lines. In this study we could not demonstrate any change in concentration, size or charge of SAP-2 in fibroblast extracts from Niemann-Pick disease Type C, and we provided evidence that SAP-2 might be subject to post-translational processing similar to that of lysosomal enzymes.