CLEAVAGE EN BLOC OF N-ACETYLGLUCOSAMINE-6-SULFATE AND N-ACETYLGALACTOSAMINE-6-SULFATE BY HEXOSAMINIDASE A: DEFICIENCY OF BOTH ACTIVITIES IN TAY-SACHS DISEASE

Abstract

Current assays of hexosaminidase A (Hex A) are either indirect (heat-inactivation method) or complex (GM₂ ganglioside substrate). The finding in Sanfilippo syndrome type D (N-acetylglucosamine-6-sulfatase deficiency) that Hex A can bypass the blockage in keratan sulfate degradation by cleavage en bloc of β-N-acetyl-glucosamine-6-sulfate, led to the present approach. 4-methyl-umbelliferyl derivatives of β-N-acetylglucosamine-6-sulfate (MUβG1cNAc-6-S) and β-N-acetylgalactosamine-6-sulfate (MUβGa1NAc-6-S) were prepared from the commonly used unsulfated derivatives. Sera (n=5), leukocytes (n=3) and fibroblasts (n=2) from patients with Tay-Sachs disease (TSD) demonstrated markedly deficient activities toward both sulfated substrates, 2-4% of control activities (n=12), and samples from obligate heterozygotes for TSD (n=4) had intermediate activity values, 43-62% of control activities. Fibroblasts (n=2) from patients with Morquio syndrome type A (N-acetylgalactosamine-6-sulfatase deficiency) had normal activities toward both sulfated substrates but the characteristic urinary excretion of chondroitin sulfate in this disease, indicates that there is no analogy with Sanfilippo syndrome type D and Hex A is incapable of cleaving en bloc βGa1NAc-6-S from the natural substrate. In contrast to natural substrates, the synthetic sulfated substrates are both specific for Hex A and thus, can provide direct and simple methods for diagnosis as well as for antenatal and carrier detection of TSD.

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