Biochemical and molecular characterization of novel mutations in GLB1 and NEU1 in patient cells with lysosomal storage disorders.

Lysosomes are cytoplasmic compartments that contain many acid hydrolases and play critical roles in the metabolism of a wide range of macromolecules. Deficiencies in lysosomal enzyme activities cause genetic diseases, called lysosomal storage disorders (LSDs). Many mutations have been identified in the genes responsible for LSDs, and the identification of mutations is required for the accurate molecular diagnoses. Here, we analyzed cell lines that were derived from two different LSDs, GM1 gangliosidosis and sialidosis. GM1 gangliosidosis is caused by mutations in the GLB1 gene that encodes β-galactosidase. A lack of β-galactosidase activity leads to the massive accumulation of GM1 ganglioside, which results in neurodegenerative pathology. Mutations in the NEU1 gene that encodes lysosomal sialidase cause sialidosis. Insufficient activity of lysosomal sialidase progressively increases the accumulation of sialylated molecules, and various clinical symptoms, including mental retardation, appear. We sequenced the entire coding regions of GLB1 and NEU1 in GM1 gangliosidosis and sialidosis patient cells, respectively. We found the novel mutations p.E186A in GLB1 and p.R347Q in NEU1, as well as many other mutations that have been previously reported. We also demonstrated that patient cells containing the novel mutations showed the molecular phenotypes of the corresponding disease. Further structural analysis suggested that these novel mutation sites are highly conserved and important for enzyme activity.