Transient expression of wild-type and mutant glucocerebrosidases in hybrid vaccinia expression system.
Gaucher disease, the most prevalent lysosomal storage disease, is characterised by a significant phenotypic variation caused by more than 150 mutations. In order to verify pathogenicity of mutations found in the Czech Gaucher population, the vaccinia expression system was used. The wild-type human beta-glucocerebrosidase cDNA and cDNAs carrying the mutations 72delC, 1326insT, 1263del55, S196P, N370S, L444P, G202E, D409H, T369M, L444P+V460V, and D409H+T369M were expressed in Gaucher fibroblast cell line (L444P/S107L), BSC40, and HeLa G cells. The enzymatic activity and immunological reactivity were analysed. Only beta-glucocerebrosidase-deficient fibroblasts were suitable for expression using plasmid transfection. The expressed beta-glucosidase activity of mutant glucocerebrosidases was in good correlation with the presumed severity of the mutations.
Hodanová
Katerina
K
Center for Integrated Genomics and Institute of Inherited Metabolic Disorders, First Faculty of Medicine, Charles University, Ke Karlovu 2, Praha 2, 128 08, Czech Republic.
Melková
Zora
Z
Horowitz
Mia
M
Hrebícek
Martin
M
D003001:Cloning, Molecular; D005776:Gaucher Disease / Q000201:enzymology; D005822:Genetic Vectors; D005962:Glucosylceramidase / Q000096:biosynthesis / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D014616:Vaccinia virus
D005962:Glucosylceramidase