Lactosylceramide beta-galactosidase in human sphingolipidoses. Evidence for two genetically distinct enzymes

In view of recent conflicting reports from two laboratories, activities of lactosylceramide beta-galactosidase were reinvestigated in detail in brains and livers of normal individuals and of patients with globoid cell leukodystrophy or GM1-gangliosidosis. Both sets of the apparently totally contradictory results were readily reproduced simply by using the different assay systems of the respective laboratories. With our own assay system, hepatic lactosylceramide beta-galactosidase appeared deficient only in Gm1-gangliosidosis, while it appeared deficient only in globoid cell leukodystrophy when the assay system of Wenger et al. (Wenger, D.A., Sattler, M., Clark, D., and McKelvey, H. (1974) Clin. Chim. Acta 56, 199-206) was used. Analyses of individual constitutents in the two assay systems revealed their complex effects on measured activities of the enzyme. The findings were strongly indicative of the existence of two genetically distinct lactosylceramide-cleaving enzymes. One enzyme (lactosylceramidase I) may be identical with galactosylceramide betal-galactosidase, and the other (lactosylceramidase II) is closely related to nonspecific 4-methylumbelliferyl beta-galactosidase. Normal human brain contains mostly lactosylceramidase I, while normal liver contains predominantly lactosylceramidase II. Lactosylceramidase I is genetically lacking globoid cell leukodystrophy, and lactosylceramidase II in GM1-gangliosidosis. Lactosylceramidase I is activated by either pure or crude taurocholate and by oleic acid and is only slightly activated by chloride ions. Lactosylceramidase II is activated by crude taurocholate but not by pure taurocholate. As activators, oleic acid is less effective and chloride more effective than for lactosylceramidase I. Citrate-phosphate buffer is more favorable to lactosylceramidase I than citrate buffer, while lactosylceramidase II responds in reverse. The standard assay system used by Wenger et al. determines almost exclusively lactosylceramidase I, while our own standard system is optimal for lactosylceramidase II and is less favorable for lactosylceramidase I. With a highly purified human hepatic beta-galactosidase preparation, exxentially free of galactosylceramide beta-galactosidase activity, lactosylceramide-cleaving activity determined by the Wenger system was less than 2 per cent of that determined by our system. If lactosylceramide beta-balactosidase assays are to be used for diagnosis of globoid cell leukodystrophy, it is absolutely essential to use an appropriate assay system in order to avoid errors of serious consequences.