Characterization of β-galactosidase in leukocytes and fibroblasts of GM1 gangliosidosis heterozygotes compared to normal subjects

Objectives: Characterization of β-galactosidase in leukocytes and fibroblasts of heterozygotes for GM1 type I. Design and methods: Leukocyte and fibroblast β-galactosidase activity was determined fluorimetrically using 4-methylumbelliferyl-β-D-galactoside as an artificial substrate. Optimum pH, Km, Vmax and thermostability of the enzyme at 42 °C were determined. Results: The leukocyte and fibroblast enzyme of heterozygotes have an optimum pH of 4.0 and 4.2, respectively. In normal subjects, the optimum pH was 4.2 in both cells, according to previous studies. The Km of the enzyme of heterozygotes was determined to be 0.65 mM in leukocytes and 0.59 mM in fibroblasts. The Vmax was determined in 167.21 nmol/h/mg of protein in heterozygotes leukocytes and 541.2 nmol/h/mg of protein in heterozygotes fibroblasts compared to 291.7 and 1768.1 nmol/h/mg of protein in controls leukocytes and fibroblasts, respectivelly. When leukocyte and fibroblast heterozygote β-galactosidase was preincubated at 42 °C, after 80 min the residual activity was determined to be 25 to 30% of the initial activity. These results are similar to the control group. Conclusions: We have found significant differences between the two groups in some investigated parameters. Both fibroblasts and leukocytes showed a virtually similar level of reliability as source of enzyme for the detection of heterozygotes.