Lysosomal enzyme hexosaminidase

Methods for the assay of hexosaminidase A and total hexosaminidase activities in dried blood spots on filter paper offer considerable advantages for screening (Chamoles et al., 2002). The deficient activity of the lysosomal enzymes hexosaminidase A and total hexosaminidase (hexosaminidase A plus B) are usually measured in plasma or extracts of leukocytes. To tubes containing a 3-mm-diameter blood spot, elution liquid and substrate solution were added. After incubation at 37°C, the amount of hydrolyzed product was compared with a calibrator to allow the quantification of enzyme activity. The method was proven reliable even after storage for up to 38 months at room temperature. For total hexosaminidase, the substrate was 4-methyl-umbelliferyl-2-acetamido-... 

The biochemical defect In Tay-Sachs disease is an inherited deficiency of -hexosaminidase, a lysosomal enzyme responsible for hydrolysis ofGM2 ganglioside, which accumulates abnormally in the lyso-somes. 

Several laboratories have studied the assimilation of specific lysosomal enzymes using as model systems skin fibroblasts deficient in the enzyme under study. The underlying mechanism for the translocation of lysosomal enzymes was hypothesized to involve binding of carbohydrate-containing recognition markers to specific cell surface receptors (1 5). In support of this hypothesis Hickman, Shapiro, and Neufeld (16J found that treatment of N-acetyl-B-hexosaminidase with periodate under conditions that dTd not affect enzymatic activity prevented the efficient assimilation of this enzyme by Sandhoff fibroblasts. Additionally, Kresse and von Figura (1 7) found that treatment of f -acetyl-a-hexosaminidase with B-galactosidase reduced the assimilation of this enzyme by San-filippo B fibroblasts. 

Many tissues from patients with I-cell disease exhibit normal levels of intracellular lysosomal enzymatic activities (e.g., brain, liver, kidney, and spleen). In the liver, lysosomal enzyme levels are normal except for diminished P-galactosidase and elevated P-hexosaminidase, P-xylosidase.and a-galactosidase. This fairly spe-... 

Brain lysosomes are deficient in the enzyme hexosaminidase A. The enzyme has two subunits, A and B. Because of a lack of the enzyme, hydrolysis of the terminal N-acetylgalactosamine from the... 

Tay-Sachs disease is a fatal genetic disorder where harmful amounts of lipids called ganglioside accumulate in the nerve cells and brains of those affected. Infants with this disorder appear normal for the first several months of life, and then as the lipids distend the nerve cells and brain cells, progressive deterioration occurs the child becomes blind, deaf, and eventually unable to swallow. Tay-Sachs disease occurs mainly in Jewish children of Eastern European descent, and death from bronchopneumonia usually occurs by age 3 to 4 years. A reddish spot on the retina also develops, and symptoms first appear around 6 months of age. It is a lysosomal storage disorder with insufficient activity of the enzyme hexosaminidase A, which catalyzes the biodegradation of the gangliosides. The diagnosis is made by the clinical suspicion and serum hexosaminidase level. Currently there is no treatment available for this disease. 

Present evidence suggests that proteolysis of the precursor form of lysosomal enzymes is critical for targeting [43,44]. Studies using portions of the cloned Dictyostelium P-hexosaminidase [45] fused to yeast invertase as a reporter have been unable to determine... 

O-GlcNAcase with even higher selectivities of up to 9 x 105-fold over the lysosomal hexosaminidases.306,307 The A/-(3-thiopropanoyl) derivative 145 in complex with recombinant enzyme (PDB 2XPK) showed the acyl residue in a selectivity pocket, which distinguishes the enzyme from the human lysosomal hexosaminidases, which feature more shallow pockets that cannot accommodate the extended acyl moiety. 

Tay-Sachs disease A genetic disease that is a result of a deficiency in hexosaminidase A (P-A-acetylhexosaminidase), an enzyme that is involved in the degradation of ganghosides in the lysosome. The disease is prevalent in Jewish children of Eastern European descent and leads to a buildup of ganghoside in nerve cells of the brain and neuronal dysfunchon. 

Defective hexosaminidase enzyme leads to accumulation of excess sphingolipids in the lysosomes of neurons, impairing neural development. 

Fig. 17.4 Simplified structure of a complex N-linked glycan and the sites of action of B-hexosaminidase. Glycoproteins are degraded by specific enzymes from both directions. Lysosomal catabolism is terminated when an A-acetylglucosamine residue is encountered by the catalytically impaired enzyme B-hexosaminidase, and thus the glycoprotein-derived oligosaccharide in the box accumulates in Sandhoff patients.

Many of the chaperons used have been imino sugars because of their tight-binding properties to the active site and at sub-inhibitory concentrations produce an enhancement of enzyme that may be sufficient to degrade the lysosomally stored material. For the neuronopathic disorders few candidates have been identified but a recent chemical library screen for potential hexosaminidase chaperons has revealed additional molecular frameworks that could be used that are outside the imino sugar structural motif (Tropak et al., 2007). 

Since exO /S-jV-acetyl-D-hexosaminidases show little or no activity tow ard appropriate sulfated hexosaminides, a major difficulty with the pathway in Fig. 4 for the degradation of the choiidroitin sulfates has been the apparent absence of demonstrable oligosaccharide-sulfatase activities in mammalian tissues (Dodgson and Lloyd, 1968). It now appears that suitable sulfatases do exist in lysosomes, and that the enzymic pathway in Fig. 4c occurs in mammalian tissues (Aronson and Davidson, 1968). 

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