Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.23.5 (cathepsin D)
 4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The kinetic properties of UDP-N-acetylglucosamine:glycoprotein N-acetylglucosamine-1-phosphotransferase (GlcNAc-phosphotransferase) partially purified from the soil amoeba Acanthamoeba castellanii have been studied. The transferase phosphorylated the lysosomal enzymes uteroferrin and cathepsin D 3-90-fold better than nonlysosomal glycoproteins and 16-83-fold better than a Man9GlcNAc oligosaccharide. Deglycosylated uteroferrin was a potent competitive inhibitor of the phosphorylation of intact uteroferrin (Ki of 48 microM) but did not inhibit the phosphorylation of RNase B or the simple sugar alpha-methylmannoside. Deglycosylated RNase (RNase A) did not inhibit the phosphorylation of RNase B or uteroferrin. These results indicate that purified amoeba GlcNAc-phosphotransferase recognizes a protein domain present on lysosomal enzymes but absent in most nonlysosomal glycoproteins. The transferase also exhibited a marked preference for oligosaccharides containing mannose alpha 1,2-mannose sequences, but this cannot account for the high affinity binding to lysosomal enzymes. A. castellanii extracts do not contain detectable levels of N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase, the second enzyme in the biosynthetic pathway for the mannose 6-phosphate recognition marker. We conclude that A. castellanii does not utilize the phosphomannosyl sorting pathway despite expression of very high levels of GlcNAc-phosphotransferase.
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PMID:Characterization of UDP-N-acetylglucosamine:glycoprotein N-acetylglucosamine-1-phosphotransferase from Acanthamoeba castellanii. 131 74

To determine the cause of the increased content of carbohydrate-bound phosphate in tumour lysosomal hydrolases, the activity and kinetics in human hepatocellular carcinoma of two enzymes involved in the formation of mannose-6-phosphate in lysosomal hydrolases UDP-GlcNAc: lysosomal enzyme GlcNAc alpha l-phosphotransferase (GlcNAc-phosphotransferase) and phosphodiester glycosidase were studied. The activity level of the phosphotransferase with artificial and natural substrates was elevated (P less than 0.025 and P less than 0.001, respectively) in hepatoma compared to that in uninvolved tissue, while the phosphodiester glycosidase of hepatoma was at a level similar to that of the uninvolved tissue. To verify a previous observation that cathepsin D of human hepatoma contained increased GlcNAc-phosphomannose, the protease was examined for carbohydrate phosphorylation by the GlcNAc-phosphotransferase. The protease from normal human liver was much more phosphorylated than hepatoma protease, confirming the previous observation. The predominant phosphorylation of the protease occurred in one of two major heavy subunits, with some phosphorylation in one of two minor light subunits.
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PMID:Elevated carbohydrate phosphotransferase activity in human hepatoma and phosphorylation of cathepsin D. 164 48

The uncovering ratio of phosphate groups in lysosomal enzymes is defined as the percentage of phosphomonoester groups in the oligosaccharide side chains based on the sum of phosphomonoester and phosphodiester groups. Using a new procedure for the specific and complete hydrolysis of uncovered phosphomonoester groups in denatured immunoprecipitates of human cathepsin D, we show that the uncovering ratio varies between different forms of the enzyme and may be used as an indicator of the maturation of its carbohydrate side chains. The uncovering ratio in the total (cellular and secreted) cathepsin D from U937 promonocytes is greater than 95%. It is only slightly decreased in cells incubated in the presence of 1 alpha,25-dihydroxycholecalciferol, in which the rate of synthesis of cathepsin D is several times higher than in the control cells. In U937 cells and also in fibroblasts, the uncovering is nearly complete in intermediate and mature forms of the intracellular cathepsin D but less extensive in the intracellular and secreted precursor. In both cell types, incubation with 10 mM NH4Cl results in a decrease in the uncovering ratio of total cathepsin D. However, the activity of the uncovering enzyme, N-acetylglucosamine-1-phosphodiester alpha-N-acetylglucosaminidase, as determined with UDP-N-acetylglucosamine is not affected with up to 60 mM NH4Cl. Our results suggest that NH4Cl, in addition to its known effects on the acidic-pH-dependent functions of lysosomal compartments and of mannose-6-phosphate receptors, impairs the processing or transport of lysosomal enzyme precursors at, or proximally to, the site of the uncovering of their mannose-6-phosphate residues.
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PMID:Suppression of the 'uncovering' of mannose-6-phosphate residues in lysosomal enzymes in the presence of NH4Cl. 216 47

Two enzymes (N-acetylglucosamine-1-phosphotransferase and phosphodiester glycosidase) involved in formation of mannose-6-phosphate at lysosomal hydrolases were studied for the activity and kinetics in human hepatocellular carcinoma. The activity level of the phosphotransferase with an artificial substrate was elevated (p less than 0.025) in hepatoma compared to that in normal liver, while the phosphodiester glycosidase of hepatoma was in a similar level with that of control. The elevation was more remarkable with a physiological substrate, cathepsin D. (P less than 0.001). Since cathepsin D from normal liver was previously demonstrated to contain less phosphomannose compared to the hepatoma protease, the protease was investigated for carbohydrate phosphorylation by the phosphotransferase. The liver protease was much more phosphorylated than the hepatoma protease, endorsing the previous observation. The predominant phosphorylation of the protease occurred in heavy subunit.
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PMID:[Carbohydrate phosphotransferase in human hepatoma and phosphorylation of cathepsin D]. 217 73

Four Baluch siblings with mucolipidosis type III (pseudo-Hurler polydystrophy) are described. The patients had features commonly found in mucolipidosis III, including claw hands, joint stiffness, aortic valve involvement and radiological dysostosis multiplex. However, intelligence was normal, there were no eye abnormalities on slit-lamp examination and skin elasticity was normal. Many lysosomal enzymes were elevated in serum and diminished in cultured fibroblasts, although the findings for beta-galactosidase were atypical. Assays for the two enzymes involved in formation of the phosphomannose recognition marker revealed normal activity of the phosphotransferase with alpha-methylmannoside as an acceptor, and normal activity of the phosphodiester glycosidase. Metabolic labelling of fibroblasts with 32P followed by immunoprecipitation of cathepsin D, electrophoresis and fluorography showed that this enzyme was not labelled in the patients' cells, although some label was detected in the secreted precursor polypeptide. The data are consistent with the assumption that activity of the phosphotransferase is low towards lysosomal enzymes as substrates, and that the patients belong to complementation group C.
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PMID:A mild form of mucolipidosis type III in four Baluch siblings. 813 3