n-acetylglucosamine-6-sulfate and Tay-Sachs-Disease

4-Methylumbelliferyl-6-sulfo-2-acetamido-2-deoxy derivatives of beta-D glucopyranoside and beta-D-galactopyranoside were prepared by direct sulfation of the commonly used unsulfated derivatives. Both sulfated substrates were highly specific for hexosaminidase A, and in fractionated serum, cells, and tissue preparations, less than 2.5% of these activities were associated with hexosaminidase B and the intermediate isozyme fractions. Serum and leukocytes from patients with infantile Tay-Sachs disease, including a patient with thermolabile hexosaminidase B, had less than 2% of noncarrier activities. Carrier values were clearly separated from those of noncarriers, and no problems were encountered in utilizing sera from pregnant women. The % hexosaminidase A values as derived from the ratio between the activities toward the sulfated and unsulfated substrates in the same specimen were comparable to those obtained by the heat-inactivation method (except for subjects with thermolabile hexosaminidase B) and may be helpful in genotype determination in borderline cases.

Topics: Acetylglucosamine; beta-N-Acetylhexosaminidases; Female; Genetic Carrier Screening; Hexosaminidase A; Hexosaminidase B; Humans; Pregnancy; Substrate Specificity; Tay-Sachs Disease

The first step of the degradation of p-nitrophenyl-6-sulfo-2-acetamido-2-deoxy-beta-D-glucopyranoside and of keratan sulfate-derived oligosaccharides bearing N-acetylglucosamine-6-sulfate residues at the nonreducing end was considered to be accomplished by the action of a specific sulfatase (Kresse, H., Paschke, E., von Figura, K., Gilberg, W., and Fuchs W. (1980) Proc. Natl. Acad. Sci. U. S. A. 77, 6822-6826). In purification from human placenta, however, this activity co-chromatographed with isoenzyme A of beta-N-acetylhexosaminidase and had the same electrophoretic mobility as the latter enzyme. The activity was precipitated by a specific antiserum against beta-N-acetylhexosaminidase. A pronounced enzyme deficiency was found in Tay-Sachs and Sandhoff fibroblasts. The purified enzyme released p-nitrophenol from the chromogenic substrate as well as a second product which contained equimolar amounts of hexosamine and sulfate. This product had the same electrophoretic and chromatographic behavior as sulfated N-acetylglucosamine. It could be degraded by periodate to a smaller charged fragment. Incubation of keratan sulfate-derived oligosaccharides with beta-N-acetylhexosaminidase A analogously resulted in the liberation of N-acetylglucosamine-6-sulfate. The enzyme showed the highest affinity towards a trisulfated tetrasaccharide and exhibited a similar Km for the sulfated and the unsulfated p-nitrophenyl derivative.

Topics: Acetylglucosamine; beta-N-Acetylhexosaminidases; Cells, Cultured; Fibroblasts; Glucosamine; Hexosaminidases; Humans; Kinetics; Mucopolysaccharidoses; Sandhoff Disease; Skin; Substrate Specificity; Sulfur Radioisotopes; Tay-Sachs Disease; Tritium