P-Hexosaminidases

The crystal structure of human P-hexosaminidase B (GH 20) in its complex with these inhibitors (GlcNAc-thiazoline 140 PDB 1NP0 GalNAc-isofagomine 139 PDB 1NOW) revealed a very similar picture, with an activated water molecule above the P face ready to attack the anomeric center. 300... 

The best known and the commonest sphingolipi-dosis is Tay-Sachs disease.366-368 Several hundred cases have been reported since it was first described in 1881. A terrible disease, it is accompanied by mental deterioration, blindness, paralysis, dementia, and death by the age of three. About 15 children a year are born in North America with this condition, and the world figure must be 5-7 times this. The defect is in the a subunit of the (3-hexosaminidase A (point 7 in Fig. 20-10)366,366a with accumulation of ganglioside GM2- Somewhat less severe forms of the disease are caused by different mutations in the same gene369 or in a protein activator. Sandhoff disease, which resembles Tay-Sachs disease, is caused by a defect in the (3 subunit, which is present in both P-hexosaminidases A and B.368 Mutant "knockout" mice that produce only ganglioside GM3 as the major ganglioside in their central nervous system die suddenly from seizures if they hear a loud sound. This provides further evidence of the essential nature of these components of nerve membranes.3693... 

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-... 

Sandhoff disease G M2 gangliosidosis p-subunit p-hexosaminidase 5ql3... 

Hultberg, B., Isaksson, A., Berglund, M., Moberg, A.-L. Serum p-hexosaminidase isoenzymze a sensitive marker for alcohol abuse. Alcohol. Clin. Exp. Res. 1991 15 549 - 552... 

Initial studies with mast cell cultures or granules demonstrated that mast cells appear to have several important effects on connective tissue elements. Subba Rao et al. (1983) showed that cultured rat embryonic skin fibroblasts phagocytose rat mast cell granules added to the culture medium or released by co-culmred mast cells, and that this is followed by secretion of collagenase and p-hexosaminidase. Similarly Yoffe et al. (1984) demonstrated that granules derived from mast cells purified from dog mastocytomas induced a 10- to 50-fold stimulation of collagenase production by human... 

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... 

Burg, J., Banerjee, A., and Sandhoff, K., Activating proteins for ganglioside GM2 degradation by P-hexosaminidase isozymes in tissue extracts from different species. Biol. Chem. Hoppe-Seyler. 366, 887-891 (1985). 

In our laboratory an authentic in vitro cellular model for Sandhoff disease has been generated upon treatment of RAW264.7 murine macrophages with an inhibitor of p-hexosaminidase (Boomkamp and Butters, unpublished data). High performance liquid chromatography (HPLC) analyses of extracted GSL oligosaccharides show that GA2 predominantly accumulates in inhibitor-treated RAW cells as opposed to a minor but significant increase in GM2 levels (Fig. 17.7). This difference in elevation is possibly due to the... 

The specific interaction between p-hexosaminidase A and the Gm2 activator has also been taken into account in the development of assay systems for ganglioside Gm2 hydrolysis by extracts of cultured skin fibroblasts (Erzberger et al., 1980 Conzelmann et al., 1983). With such assays the residual activities in cells from patients with different variants of Gm2 gangliosidosis could be determined very precisely (Conzelmann et al., 1983). 

Since, as in simple Michaelis-Menten kinetics, the reaction rate depends linearly on the enzyme concentration, the accuracy of activator determinations depends on the precision with which the enzyme concentration is known. Although it is not necessary to employ pure enzymes for the purpose of activator quantification, problems may arise if several isoenzymes exist of which only one is able to degrade the glycolipid substrate in the presence of the activator but all of them hydrolyze the artificial substrates employed for measuring the enzyme. This is, for example, the case for a-galactosidase, p-hexosaminidase, and arylsulfatase. In these cases separation of the isoenzymes, say by ion-exchange chromatography, is required. 

Gi/mol), 2.5 p,g bovine serum albumin, 100 mU P-hexosaminidase A, and up to 25 p.1 of the suitably diluted activator sample in a total volume of 40 (il mM citrate buffer, pH 4.0, and is incubated for 1 -4 h at 37°C. Samples are then transferred to an ice bath and loaded onto 1 ml columns of DEAE-cellulose (in Pasteur pip>ettes) that have been washed with distilled water. Liberated [ H]A-acetylgalactosamine is eluted with 2 x 1 ml of a 1 mM aq A-acetylgalactosamine solution. The combined effluents are collected in scintillation vials, and after addition of 10 ml scintillation fluid, their radioactivity is measured. Blanks run with water instead of activator solution are subtracted. 

Sphingomyelinase Ceramidase P-Glucocerebrosidase P-Galactosyl hydrolase P-Hexosaminidase A P-Galactosidase a-Galactosidase P-Hexosaminidases A and B Galactocerebrosidase Arylsulfatase A... 

Granberg, M., Fowler, C. J., and Jacobsson, S. O. P. (2001). Effects of the caimabimimetic fatty acid derivatives 2-arachidonoylglycerol, anandamide, palmitoylethanolamide and methanandamide upon IgE-dependent antigen-induced p-hexosaminidase, serotonin and TNF a release from rat RBL-2H3 basophilic leukaemia cells. Naunyn Schmiedebergs Arch. Pharmacol. 364, (y6-l i. 

---