Studies on the Isozyme of β-Glucuronidase

Sadahiro, Ryuzo; Takanashi, Seiji; Kawada, Minoru

doi:10.1093/oxfordjournals.jbchem.a128156

Cited by 17 publications

(4 citation statements)

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“…The microsomal i-glucuronidase in mouse kidney (Ide & Fishman, 1969) and rat liver Ganschow & Bunker, 1970) also differs from the lysosomal ,-glucuronidase in that a larger proportion occurs in the bound state. Moreover, it is more cationic than the lysosomal ,-glucuronidase on polyacrylamide-gel electrophoresis (Ganschow & Bunker, 1970;Mameli et al, 1972) and more basic than the latter on ion-exchange chromatography (Sadahiro et al, 1965). The soluble fl-N-acetylhexosaminidase in human spleen is exclusively in the acidic form, whereas the lysosomal and microsomal enzymes occur in both the basic and acidic form (Robinson & Stirling, 1968).…”

Section: Characterization Of Acid Hydrolases In Subcellular Fractionsmentioning

confidence: 99%

Physicochemical modifications of lysosomal hydrolases during intracellular transport

Goldstone

Koenig

1973

Biochemical Journal

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1. The following fractions were prepared from rat kidney and characterized ultrastructurally, biochemically and enzymically: (a) an ordinary rough microsomal (RM(1)) fraction; (b) a special rough microsomal (RM(2)) fraction enriched seven- to nine-fold in acid hydrolases over the homogenate; (c) a smooth microsomal (SM) fraction; (d) a Golgi (GM) fraction enriched 2.5-fold in acid hydrolases and 10-, 15- and 20-fold in sialyltransferase, N-acetyl-lactosamine synthetase and galactosyltransferase respectively; (e) a lysosomal (L) fraction enriched 15- to 23-fold in acid hydrolases. The frequency of Golgi sacs and tubules seen in the electron microscope and the specific activity of the three glycosyltransferases in these fractions increased in the order: RM(2)

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Section: Characterization Of Acid Hydrolases In Subcellular Fractionsmentioning

confidence: 99%

Physicochemical modifications of lysosomal hydrolases during intracellular transport

Goldstone

Koenig

1973

Biochemical Journal

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“…Multiple forms of fi-glucuronidase were demonstrated initially by careful chromatography on DEAE-cellulose (Moore & Lee, 1960;Sadahiro et al, 1965;Plapp & Cole, 1967;Aoshima & Sakurai, 1969;Okochi et al, 1968;Delvin & Gianetto, 1970), and subsequently by gel electrophoresis (Lundin & Allison, 1966). Attempts to demonstrate functional and electrophoretic differences between microsomal and lysosomal enzyme in mouse tissues were initially unsuccessful (Ganschow & Paigen, 1967), but later very slight electrophoretic differences were detected (Ganschow & Paigen, 1968;Ganschow & Bunker, 1970).…”

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confidence: 99%

Rabbit β-glucuronidase. Purification and properties, and the existence of multiple forms

Dean¹

1974

Biochemical Journal

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1. beta-Glucuronidase (EC 3.2.1.31) was purified from rabbit liver by a procedure involving autolysis, (NH(4))(2)SO(4) fractionation, chromatography on DEAE-cellulose and hydroxyapatite, gel filtration, sedimentation in a sucrose gradient, and isoelectric focusing. 2. Electron microscopy revealed ferritin as the major contaminant in later stages of purification and also showed aggregates of enzyme molecules. Particular attention was paid to the removal of ferritin. 3. The purified enzyme was homogeneous in polyacrylamide-gel electrophoresis both in non-dissociating conditions and in the presence of sodium dodecyl sulphate, and in Ouchterlony gel diffusion and immunoelectrophoresis against polyspecific antisera. 4. Sedimentation in sucrose gradients gave a molecular weight of 300000, whereas gel filtration indicated 440000. 5. Subunits of 75000 molecular weight were observed in gel electrophoresis in the presence of sodium dodecyl sulphate and in gel filtration in the presence of urea. 6. The K(m) value for p-nitrophenyl beta-d-glucuronide was 0.6mm, and the enzyme was extremely sensitive to lactone inhibitors. It was also inhibited by Hg(2+) ions. 7. Multiple forms were observed in the pure enzyme by isoelectric focusing, with pI values of 4.5-5.8. Subunits showed similar heterogeneity. The origin of the multiple forms was investigated in detail, and the possibility of artifact generation largely excluded. Some of the forms of lowest pI disappeared after neuraminidase digestion. The nature of the residual heterogeneity remains to be elucidated.

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“…However, such an inhibitor might be either extremely effective or present in sufficient concentration as to resist being swamped by even large quantities of enzyme. Alternate pos sibilities are that there is more than one active site on a molecule of ß-glucuronidase or that there are isozymes [24] with different spec ificity for different substrates, inhibitors or modifiers. It is not possible on the basis of experiments done thus far to distinguish between these alternate hypotheses to account for some of the effects which have been observed.…”

Section: Discussionmentioning

confidence: 99%

“…It is not possible on the basis of experiments done thus far to distinguish between these alternate hypotheses to account for some of the effects which have been observed. The demonstration of multiple sites or isozymes [24] would certainly help to clarify some of the questions which remain to be answered.…”

Section: Discussionmentioning

confidence: 99%

Urinary Inhibitors of ß-Glucuronidase

Kushinsky¹,

Chen²

1967

Enzymol Biol Clin

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A study has been made of some of the properties of the inhibitors of ß-glucuronidase in human urine. The inhibitory properties were determined by comparing the effect on the activity of ß-glucuronidase of urine pre-treated in many ways with that of a control containing no urine. A significant though variable fraction of the inhibition is caused by rapidly hydrolyzing competitive substrates of normal dietary or therapeutic origin. Other inhibitors such as 1,4-saccharolactone probably account for a substantially greater fraction of the inhibition than do the competitive substrates. The inhibition by 1,4-saccharolactone is a function of the pH, temperature and time of equilibration of the medium. Saccharolactone can he separated from the conjugated estrogens in urine by gel-filtration on Sephadex. The extent which non-substrate, competitive inhibitors or competitive substrates reduce the activity of ß-glucuronidase is a function of the inhibitor constant (K(I)) or the Michaelis constant (Km) respectively and the concentrations of these substances. On prolonged standing the activity of ß-glucuronidase is reduced more in the presence of urine than in the absence of urine. This phenomenon probably is due at least partially to the slow formation of saccharolactone from endogenous saccharic acid in urine.

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Studies on the Isozyme of β-Glucuronidase

Cited by 17 publications

References 0 publications

Physicochemical modifications of lysosomal hydrolases during intracellular transport

Physicochemical modifications of lysosomal hydrolases during intracellular transport

Rabbit β-glucuronidase. Purification and properties, and the existence of multiple forms

Urinary Inhibitors of ß-Glucuronidase

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