Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.31 (beta-glucuronidase)
7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The adenylate cyclase activator forskolin (1-10 mumol/L) inhibited 45Ca release from parathyroid hormone (PTH; 10 nmol/L) stimulated prelabeled neonatal mouse calvaria in short term culture (24 h). This effect of forskolin was potentiated by rolipram, Ro 20-1724, and isobutyl-methylxanthine, three structurally different inhibitors of cyclic AMP phosphodiesterase. Forskolin (10 mumol/L) and calcitonin (30 mU/mL) inhibited the mobilization of stable calcium and inorganic phosphate as well as the release of the lysomal enzymes beta-glucuronidase and beta-N-acetylglucosaminidase from PTH-stimulated unlabeled bones. Osteoclasts in PTH-stimulated calvaria showed active ruffled borders with numerous membrane infoldings. Treatment of PTH-stimulated bones with forskolin and calcitonin resulted in a rapid (2 h) loss of the active ruffled border. In addition, forskolin and calcitonin induced similar changes with respect to the number and size distribution of cytoplasmic vesicles in PTH-activated osteoclasts. After 24 h, all signs of osteoclast inactivation were still prominent, whereas after 48 h of treatment with forskolin or calcitonin, the reappearance of a ruffled border on a number of osteoclasts signaled an escape from the inhibitory action of both calcitonin or forskolin. These data indicate that forskolin inhibits bone resorption by a cyclic AMP dependent mechanism and that the effect of forskolin and calcitonin on bone resorption and osteoclast morphology are comparable. These observations lend further support to the view that cyclic AMP may be an intracellular mediator of the inhibitory action of calcitonin on multinucleated osteoclasts.
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PMID:Comparison between the effects of forskolin and calcitonin on bone resorption and osteoclast morphology in vitro. 260 53

A monoclonal antibody (2C5) raised against rat liver lysosomal membranes was used to identify a 78-kD glycoprotein that is present in the membranes of both endosomes and lysosomes and, therefore, is designated endolyn-78. In cultures of rat hepatoma (Fu5C8) and kidney cells (NRK), this glycoprotein could not be labeled with [35S]methionine or with [32P]inorganic phosphate but was easily labeled with [35S]cysteine and [3H]mannose. Pulse-chase experiments and determinations of endoglycosidase H (endo H) sensitivity showed that endolyn-78 is derived from a precursor of Mr 58-62 kD that is processed to the mature form with a t1/2 of 15-30 min. The protein has a 22-kD polypeptide backbone that is detected after a brief pulse in tunicamycin-treated cells. During a chase in the presence of the drug, this is converted into an O-glycosylated product of 46 kD that despite the absence of N-linked oligosaccharides is effectively transferred to lysosomes. This demonstrates that the delivery of endolyn-78 to this organelle is not mediated by the mannose-6-phosphate receptor (MPR). Immunocytochemical experiments showed that endolyn-78 is present in the limiting membranes and the interior membranous structures of morphologically identifiable secondary lysosomes that contain the lysosomal hydrolase beta-glucuronidase, lack the MPR, and could not be labeled with alpha-2-macroglobulin at 18.5 degrees C, a temperature which prevents appearance of endocytosed markers in lysosomes. Endolyn-78 was present at low levels in the plasma membrane and in peripheral tubular endosomes, but was prominent in morphologically diverse components of the endosomal compartment (vacuolar endosomes and various types of multivesicular bodies) which acquired alpha-2-macroglobulin at 18.5 degrees C, and frequently contained substantial levels of the MPR and variable levels of beta-glucuronidase. On the other hand, the MPR was very rarely found in endolyn-containing structures that were not labeled with alpha-2-macroglobulin at the low temperature. Thus, the process of lysosomal maturation appears to involve the progressive delivery of lysosomal enzymes to various types of endosomes that may have already received some of the lysosomal membrane proteins. Although endolyn-78 would be one of the proteins added early to endosomes, other lysosomal membrane proteins may be added only to multivesicular endosomes that represent very advanced stages of maturation.
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PMID:Endolyn-78, a membrane glycoprotein present in morphologically diverse components of the endosomal and lysosomal compartments: implications for lysosome biogenesis. 265 37

A simple rapid method for simultaneous ethosuximide and phenobarbital assay in brain tissue, serum and urine has been developed. Extraction of samples from brain tissue and serum were performed with dichloromethane at low pH in the presence of an excess of ammonium sulfate. Glucuronide conjugates in urine samples were hydrolyzed by enzymatic cleavage with beta-glucuronidase and then extracted with dichloromethane. The extracts were analyzed using a Spherisorb 5 ODS column and a mixture of acetonitrile, methanol and phosphate buffer (21:24:55, v/v) as eluent. No interference was encountered and the method is both precise and reproducible.
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PMID:Simultaneous measurement of ethosuximide and phenobarbital in brain tissue, serum and urine by HPLC. 273 17

We studied the mechanism of gallbladder sludge formation in guinea pigs (n = 30) treated with lincomycin (80 mg/kg/day) for 7 consecutive days. At sacrifice (day 8) gallbladders of treated animals contained turbid bile, sludge and in one animal a single gallstone. The precipitates were amorphous on X-ray diffraction. Infra-red spectroscopy revealed calcium phosphate as the major component. Compared to saline-treated controls (n = 15) concentrations of total protein, total phosphate and total bilirubin in gallbladder bile were significantly increased (P less than 0.05). The increase in total phosphate was due to the inorganic component, since phospholipid phosphorus was unchanged. The relative amounts of unconjugated bilirubin and of bilirubin mono- and diconjugates in gallbladder bile were unaffected by treatment as was beta-glucuronidase activity. However, sludge was enriched in unconjugated bilirubin compared to gallbladder bile. This was most probably caused by alkaline hydrolysis of bilirubin monoconjugates. To some extent, disproportionation of bilirubin monoconjugates in bile or sludge, either in vivo or during sample preparation, might also have led to increased unconjugated pigment.
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PMID:Lincomycin treatment of guinea pigs causes formation of pigmented phosphate containing gallbladder sludge and stones. 280 56

We have utilized the adenylate cyclase stimulator, cholera toxin, as a tool to test the role of cyclic AMP as a mediator of the effects on bone resorption by the calcium-regulating hormones, parathyroid hormone (PTH) and calcitonin. The effects on bone resorption were studied in an organ culture system using calvarial bones from newborn mice. Cyclic AMP response was assayed in calvarial bone explants and isolated osteoblasts from neonatal mouse calvaria. Cholera toxin caused a dose-dependent cAMP response in calvarial bones, seen at and above approx. 1-3 ng/ml and calculated half-maximal stimulation (EC50) at 18 ng/ml. The stimulatory effect of cholera toxin could be potentiated by the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX, 0.2 mmol/l). Cyclic AMP accumulation in the bones was maximal after 4-6 h, and thereafter declined. However, activation of the adenylate cyclase was irreversible and the total amount (bone + medium) of cAMP produced, in the presence of IBMX (0.2 mmol/l), increased with time, for at least 48 h. In osteoblast-like cells cholera toxin (1 microgram/ml) stimulated the cellular levels of cAMP with a peak after 60-120 min, which could be potentiated with IBMX. The total cAMP accumulation indicated an irreversible response. In short-term bone organ cultures (at most, 24 h) cholera toxin, at and above 3 ng/ml, inhibited the stimulatory effect of PTH (10 nmol/l) on 45Ca release from prelabelled calvarial bones. The inhibitory effect of cholera toxin (0.1 microgram/ml) on 45Ca release was significant after 6 h and the calculated IC50 value at 24 h was 11.2 ng/ml. Cholera toxin (0.1 microgram/ml) also inhibited PTH-stimulated (10 nmol/l) release of Ca2+, inorganic phosphate (Pi), beta-glucuronidase, beta-N-acetylglucosaminidase and degradation of organic matrix (release of 3H from [3H]proline-labelled bones) in 24 h cultures. 45Ca release from bones stimulated by prostaglandin E2 (1 mumol/l) and 1 alpha-hydroxyvitamin D3 (0.1 mumol/l) was also inhibited by cholera toxin (0.3 microgram/ml) in 24-h cultures. The inhibitory effect of cholera toxin on bone resorption was transient, and in long-term cultures (120 h) cholera toxin caused a dose-dependent, delayed stimulation of mineral mobilization (Ca2+, 45Ca, Pi), degradation of matrix and release of the lysosomal enzymes beta-glucuronidase and beta-N-acetylglucosaminidase.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Effects of cholera toxin on cyclic AMP accumulation and bone resorption in cultured mouse calvaria. 282 May 4

Activities of a cathepsin B-like cysteine proteinase have previously been observed to correlate with the malignancy of several animal and human tumors. Plasma membrane fractions of some of these tumors have been found to be enriched in cathepsin B-like activity. We have determined the subcellular distribution of this enzyme and three additional lysosomal hydrolases (cathepsin H, beta-hexosaminidase, and beta-glucuronidase) in normal murine liver and six metastatic variants of the B16 melanoma. The tissues were fractionated initially by differential centrifugation followed by Percoll density gradient centrifugation of the light mitochondrial fraction. Two fractions were obtained: an L-2 fraction enriched in all four lysosomal hydrolases; and an L-1 fraction enriched in a marker enzyme for the plasma membrane. Cathepsin B-like and beta-hexosaminidase activities, but not the other hydrolase activities, were also found to be enriched in the L-1 fractions of the metastatic B16 tumors. We explored the nature of the association of the cathepsin B-like activity with the plasma membrane using fractions from the spontaneously metastatic B16 amelanotic melanoma. Activity could not be dissociated from the plasma membrane fraction by washing with a physiological salt solution suggesting that it was not adsorbed to this fraction nonspecifically, nor could it be displaced by mannose 6-phosphate or other sugars which compete for binding to the known lysosomal receptors. High salt concentrations, low concentrations of the mild detergent saponin, mild acidification, or phosphatidylinositol-specific phospholipase C did not elute the cathepsin B-like activity. However, activity was eluted by exposure to 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, a detergent used in the purification of integral membrane proteins. The B16 amelanotic melanoma plasma membrane-associated cathepsin B-like activity had a slightly higher pH optimum and was resistant to inactivation by neutral pH and to inhibition by three low molecular weight inhibitors of cysteine proteinases. The Ki values for inhibition by leupeptin and stefin A were 20-fold higher. The presence of a cathepsin B-like cysteine proteinase at the surface of metastatic tumor cells, particularly in a form which can retain activity at physiological pH and retain activity in the presence of extracellular proteinase inhibitors, may contribute to the focal dissolution of the extracellular matrix observed at sites of contact with invading tumor cells.
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PMID:Properties of a plasma membrane-associated cathepsin B-like cysteine proteinase in metastatic B16 melanoma variants. 282 39

A high-performance liquid chromatographic method for the determination of triazolam and its main metabolites (1-hydroxymethyltriazolam and 4-hydroxytriazolam) in human urine has been developed. After hydrolysis with beta-glucuronidase, the unchanged drug and its metabolites were extracted with a Sep-Pak C18 cartridge and further purified by a Sep-Pak silica cartridge. The extract was chromatographed on a reversed-phase column using a mobile phase consisting of methanol-10 mM phosphate buffer, pH 8 (65:35) and UV detection at 220 nm. The overall recoveries of triazolam, 1-hydroxymethyltriazolam and 4-hydroxytriazolam were ca. 88, 75 and 75%, respectively, and the detection limits of these compounds were 5 ng/ml, using a 10-ml specimen.
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PMID:High-performance liquid chromatographic determination of triazolam and its metabolites in human urine. 283 Feb 92

beta-Glucuronidases purified from human hepatoma and from normal liver could serve as a substrate for a cAMP-dependent protein kinase. The rate of phosphorylation reaction of the hepatoma beta-glucuronidase was rapid, whereas that of the normal liver beta-glucuronidase was slow and much lower. Stoichiometry of phosphorylation was 4.3 and 0.46 mol of phosphate/mol of the beta-glucuronidase from the hepatoma and normal liver, respectively. Tryptic peptide mapping of 32P-labeled beta-glucuronidase from hepatoma identified two distinct phosphopeptides (X and Y). The peptide from hepatoma hydrolase was phosphorylated predominantly at the X, while the peptide Y was the major phosphopeptide in the hydrolase of normal liver. Analysis of phosphoamino acids revealed two sites, phosphoserine and phosphothreonine. beta-Glucuronidase from hepatoma consisted of a major subunit with molecular mass of 64,000 (64 kDa) and a minor subunit with 76 kDa, whereas the hydrolase from normal liver had almost exclusively 64 kDa subunit. 32P-labeled beta-glucuronidase indicated that the 64 kDa subunit was phosphorylated both in hepatoma and normal liver beta-glucuronidases.
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PMID:Phosphorylation of beta-glucuronidases from human normal liver and hepatoma by cAMP-dependent protein kinase. 283 20

beta-Glucuronidase purified from human hepatocellular carcinoma consisted of a major subunit with molecular weight of 64,000 (64K-Da) and a minor 76K-Da subunit, whereas the hydrolase from normal liver had almost exclusively 64K-Da subunit. beta-Glucuronidase from the hepatoma and normal liver could serve as a substrate for a cAMP-dependent protein kinase. The rate of phosphorylation reaction of the hepatoma beta-glucuronidase was rapid, whereas that of the normal liver beta-glucuronidase was slow and much lower. Stoichiometry of beta-glucuronidase was 4.3 mol and 0.46 mol of phosphate per mol of the beta-glucuronidase from the hepatoma and normal liver, respectively. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of 32P-labeled beta-glucuronidase indicated that the 64K-dalton subunit was phosphorylated both in hepatoma and normal liver beta-glucuronidase. Tryptic peptide mapping of 32P-labeled beta-glucuronidase from hepatoma identified two distinct phosphopeptides (X and Y). The peptide from hepatoma hydrolase was phosphorylated predominantly at the X, while the peptide Y was the major phosphopeptide in the hydrolase of normal liver. Two-dimensional analysis of phosphoamino acids revealed two sites, phosphoserine and phosphothreonine.
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PMID:[Cancer-associated alterations of human hepatocellular carcinoma beta-glucuronidase--study on phosphorylation by 3', 5'-cyclic AMP dependent-protein kinase]. 283 6

Endocytosis of human spleen beta-glucuronidase by human fibroblasts can be completely impaired by the competitive inhibitor mannose 6-phosphate or by pretreatment with acid phosphatase or endoglycosidases H or F. However, endocytosis of bovine spleen and liver beta-glucuronidase is partially impaired by the same treatments, suggesting that the bovine enzyme contains two endocytosis recognition markers located in separate enzyme domains. The mannose 6-phosphate recognition marker seems to be responsible for approximately 23% of the bovine enzyme endocytosis. The existence of two lysosomal endocytosis systems in human fibroblasts is supported by the following facts: (a) the rate of endocytosis of mannose 6-phosphate-containing human beta-glucuronidase was not affected by the presence of high levels of the bovine enzyme (which has only the other marker). (b) Anti-215K mannose 6-phosphate receptor antibodies selectively impair the endocytosis of the beta-glucuronidase containing mannose 6-phosphate. (c) Weak bases exert a differential effect on human and bovine endocytosis. beta-Glucuronidase internalized by either system is targeted to secondary lysosomes of human beta-glucuronidase-deficient fibroblasts, where it is able to degrade accumulated glycosaminoglycans. These results suggest that human fibroblasts have two different and independent endocytic systems for targeting of acid hydrolases to lysosomes.
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PMID:Adsorptive endocytosis of lysosomal enzymes by human fibroblasts: presence of two different functional systems that deliver an acid hydrolase to lysosomes. 291 51


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