scholarly journals Evidence for an UDP-glucuronic acid/phenol glucuronide antiport in rat liver microsomal vesicles

1996 ◽  
Vol 315 (1) ◽  
pp. 171-176 ◽  
Author(s):  
Gábor BÁNHEGYI ◽  
László BRAUN ◽  
Paola MARCOLONGO ◽  
Miklós CSALA ◽  
Rosella FULCERI ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Rat Liver ◽  
Glucuronic Acid ◽  
Udp Glucuronosyltransferase ◽  
Luminal Compartment ◽  
Loading Procedure ◽  
Stimulation Of

The transport of glucuronides synthesized in the luminal compartment of the endoplasmic reticulum by UDP-glucuronosyltransferase isoenzymes was studied in rat liver microsomal vesicles. Microsomal vesicles were loaded with p-nitrophenol glucuronide (5 mM), phenolphthalein glucuronide or UDP-glucuronic acid, by a freeze–thawing method. It was shown that: (i) the loading procedure resulted in millimolar intravesicular concentrations of the different loading compounds; (ii) addition of UDP-glucuronic acid (5 mM) to the vesicles released both intravesicular glucuronides within 1 min; (iii) glucuronides stimulated the release of UDP-glucuronic acid from UDP-glucuronic acid-loaded microsomal vesicles; (iv) trans-stimulation of UDP-glucuronic acid entry by loading of microsomal vesicles with p-nitrophenol glucuronide, phenolphthalein glucuronide, UDP-glucuronic acid and UDP-N-acetylglucosamine almost completely abolished the latency of UDP-glucuronosyltransferase, although mannose 6-phosphatase latency remained unaltered; (v) the loading compounds by themselves did not stimulate UDP-glucuronosyltransferase activity. This study indicates that glucuronides synthesized in the lumen of endoplasmic reticulum can leave by an antiport, which concurrently transports UDP-glucuronic acid into the lumen of the endoplasmic reticulum.

scholarly journals Mechanism of stimulation of microsomal UDP-glucuronosyltransferase by UDP-N-acetylglucosamine

1995 ◽  
Vol 305 (1) ◽  
pp. 321-328 ◽  
Author(s):  
X Bossuyt ◽  
N Blanckaert
Keyword(s):  
Endoplasmic Reticulum ◽  
Glucuronic Acid ◽  
Carrier System ◽  
Transport Pathways ◽  
Weak Inhibitor ◽  
Cytosolic Side ◽  
In Trans ◽  
Udp Glucuronosyltransferase ◽  
Carrier Systems ◽  
Stimulation Of

We propose the existence in rat liver endoplasmic reticulum (ER) of two asymmetric carrier systems. One system couples UDP-N-acetylglucosamine (UDPGlcNAc) transport to UDP-glucuronic acid (UDPGlcA) transport. When UDPGlcNAc was presented at the cytosolic side of the ER, it then acted as a weak inhibitor of UDPGlcA uptake. By contrast, UDPGlcNAc produced a forceful trans-stimulation of microsomal UDPGlcA uptake when it was present within the lumen of the ER. Likewise, cytosolic UDPGlcA strongly trans-stimulated efflux of intravesicular UDPGlcNAc, whereas cytosolic UDPGlcNAc was ineffective in trans-stimulating efflux of UDPGlcA. A second asymmetric carrier system couples UDPGlcNAc transport to UMP transport. Microsomal UDPGlcNAc influx was markedly stimulated by UMP present inside the microsomes. Such stimulation was only apparent when microsomes had been preincubated and thereby preloaded with UMP, indicating that UMP exerted its effect on UDPGlcNAc uptake by trans-stimulation from the lumenal side of the ER membrane. Contrariwise, extravesicular UMP only minimally trans-stimulated efflux of intramicrosomal UDPGlcNAc. It is widely accepted that UDPGlcNAc acts as a physiological activator of hepatic glucuronidation, but the mechanism of this effect has remained elusive. Based on our findings, we propose a model in which the interaction of two asymmetric transport pathways, i.e. UDPGlcA influx coupled to UDPGlcNAc efflux and UDPGlcNAc influx coupled to UMP efflux, combined with intravesicular metabolism of UDPGlcA, forms a mechanism that leads to stimulation of glucuronidation by UDPGlcNAc.

scholarly journals Uridine diphosphoxylose enhances hepatic microsomal UDP-glucuronosyltransferase activity by stimulating transport of UDP-glucuronic acid across the endoplasmic reticulum membrane

1996 ◽  
Vol 315 (1) ◽  
pp. 189-193 ◽  
Author(s):  
Xavier BOSSUYT ◽  
Norbert BLANCKAERT
Keyword(s):  
Endoplasmic Reticulum ◽  
Glucuronic Acid ◽  
Endoplasmic Reticulum Membrane ◽  
Acid Uptake ◽  
Physiological Importance ◽  
Cytosolic Side ◽  
Udp Glucuronosyltransferase ◽  
Exogenous Compounds ◽  
Stimulation Of ◽  
Er Membrane

The UDP-glucuronosyltransferase (UGT) system fulfils a pivotal role in the biotransformation of potentially toxic endogenous and exogenous compounds. Here we report that the activity of UGT in rat liver is stimulated by UDP-xylose. This stimulation was found in native microsomal vesicles as well as in the intact endoplasmic reticulum (ER) membrane, as studied in permeabilized hepatocytes, indicating the potential physiological importance of UDP-xylose in the regulation of UGT. We present evidence that UDP-xylose enhances UGT activity by stimulation of (i) the uptake of UDP-glucuronic acid across the ER membrane and (ii) the elimination of the UDP and/or UMP reaction product out of the ER lumen. UDP-xylose produced a marked trans-stimulation of microsomal UDP-glucuronic acid uptake when it was present within the lumen of the ER. When UDP-xylose was presented at the cytosolic side of the ER, it acted as a weak inhibitor of UDP-glucuronic acid uptake. Likewise, cytosolic UDP-glucuronic acid strongly trans-stimulated efflux of intravesicular UDP-xylose, whereas cytosolic UDP-xylose was inefficient in trans-stimulating efflux of UDP-glucuronic acid. Microsomal UDP-xylose influx was markedly stimulated by UMP and UDP. Such stimulation was only apparent when microsomes had been preincubated and thereby preloaded with UMP or UDP, indicating that UMP and UDP exerted their effect on UDP-xylose uptake by trans-stimulation from the luminal side of the ER membrane.

scholarly journals Two kinetically-distinct components of UDP-glucuronic acid transport in rat liver endoplasmic reticulum

1996 ◽  
Vol 1283 (2) ◽  
pp. 223-231 ◽  
Author(s):  
Eric Battaglia ◽  
Susan Nowell ◽  
Richard R. Drake ◽  
Magdalena Mizeracka ◽  
Carl L. Berg ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Rat Liver ◽  
Glucuronic Acid ◽  
Acid Transport

Nuclear membrane-bound UDPglucuronosyltransferase of rat liver

1982 ◽  
Vol 60 (10) ◽  
pp. 972-979 ◽  
Author(s):  
Jan Zaleski ◽  
Surendra K. Bansal ◽  
Teresa Gessner
Keyword(s):  
Endoplasmic Reticulum ◽  
High Activity ◽  
Rat Liver ◽  
Nuclear Membrane ◽  
Glucuronic Acid ◽  
Microsomal Enzyme ◽  
Subcellular Membrane ◽  
Membrane Bound ◽  
Specific Activities

Some properties of rat liver nuclear membrane-bound UDPglucuronosyltransferase were compared with those of the endoplasmic reticulum bound enzyme. The activity of nuclear membrane-bound UDPglucuronosyltransferase was stimulated only about 1.5-fold by Lubrol WX. Under the same conditions microsomal UDPglucuronosyltransferase was, as usual, highly activated (up to 10-fold), when 4-nitrophenol was the acceptor of glucuronic acid. Specific activities of the detergent-activated enzyme were similar in microsomal and nuclear membrane preparations, when the following aglycone substrates were used: 4-methylumbelliferone, 4-nitrophenol, 1-naphthol, phenolphthalein, and testosterone. Apparent Km values for UDP-glucuronic acid ranged between 0.15–0.25 mM for glucuronidation of 4-nitrophenol and 1-naphthol, by either Lubrol WX activated or non-activated, nuclear membrane-bound UDPglucuronosyltransferase. These values were comparable to those found for detergent activated microsomal enzyme. The results show a similarity in behavior of detergent-activated UDPglucuronosyltransferase regardless of subcellular membrane source and, therefore, suggest the association of the same glucuronosyltransferase with nuclear membrane and endoplasmic reticulum. A possible significance of the presence of high activity of this enzyme in nuclear membrane is discussed.

scholarly journals Regulation of onset of development of UDP-glucuronosyltransferase activity towards o-aminophenol by glucocorticoids in late-foetal rat liver in utero

1977 ◽  
Vol 168 (3) ◽  
pp. 507-511 ◽  
Author(s):  
G J Wishart ◽  
G J Dutton
Keyword(s):  
Rat Liver ◽  
Alimentary Tract ◽  
Normal Development ◽  
In Utero ◽  
Transferase Activity ◽  
Foetal Rat ◽  
Udp Glucuronosyltransferase ◽  
Foetal Lung ◽  
Precocious Development ◽  
Stimulation Of

1. A precocious development of UDP-glucuronosyltransferase activity (EC 2.4.1.17) towards o-aminophenol is demonstrated in 15-17 day foetal rat liver in utero after dexamethasone administration to the mother. 2. This stimulation of liver transferase activity in utero is directly proportional to the dose of dexamethasone infected. 3. Precocious development of transferase activity in utero can also be effected with the natural glucocorticoid cortisol by multiple injections of large amounts of this hormone into the mother. 4. Transferase activity towards o-aminophenolin foetal lung, kidney and upper alimentary tract can also be precociously stimulated by dexamethasone in 17-day foetuses in utero. 5. Natural development of hepatic transferase activity between days 18 and 20 of gestation is retarded after foetal hypophysectomy by decapitation in utero. 6. Overall glucuronidation of o-aminophenol, as observed in foetal rat liver, is also precociously stimulated by dexamethasone. 7. From this and from evidence previously presented we suggest that glucocorticoids, which are known to increase in rat foetuses between days 17 and 20 of gestation, trigger the normal development in utero of hepatic transferase activity towards o-aminophenol which occurs at that time. We also suggest that these hormones are responsible for the rise in activity of the enzyme in foetal lung, kidney and upper alimentary tract which occurs during the same gestational period.

scholarly journals Diazobenzenesulphonate selectively abolishes stimulation of glucuronidation by UDP-N-acetylglucosamine

1980 ◽  
Vol 192 (3) ◽  
pp. 971-974 ◽  
Author(s):  
B Haeger ◽  
R de Brito ◽  
T Hallinan
Keyword(s):  
Endoplasmic Reticulum ◽  
Guinea Pig ◽  
Glucuronic Acid ◽  
Significant Inhibition ◽  
Modifying Agent ◽  
Stimulation Of

1. Basal rates of glucuronidation of oestrone (guinea pig) or of 4-nitrophenol (rat or guinea pig) were not significantly altered in sealed liver microsomal vesicles, treated with the membrane-impermeant protein-modifying agent diazobenzenesulphonate at 0.5-1.0 mM. 2. Contrarily, diazobenzenesulphonate abolished the normal stimulation of glucuronidation by UDP-N-acetylglucosamine. 3. Ultrasonication to increase microsomal permeability activated glucuronidation by 680-750% and permitted significant inhibition by diazobenzenesulphonate. 4. These findings are consistent with a model wherein glucuronyltransferases are embedded in the luminal leaflet of the endoplasmic reticulum and access of UDP-glucuronic acid to the transferases is facilitated by transmembrane carriers, which are stimulated by UDP-N-acetylglucosamine and are available to diazobenzenesulphonate; ultrasonication serves to permit access of diazobenzenesulphonate to glucuronyltransferases themselves, resulting in inhibition of their activity.

scholarly journals Glucuronidation of the environmental oestrogen bisphenol A by an isoform of UDP-glucuronosyltransferase, UGT2B1, in the rat liver

1999 ◽  
Vol 340 (2) ◽  
pp. 405-409 ◽  
Author(s):  
Hiroshi YOKOTA ◽  
Hidetomo IWANO ◽  
Mari ENDO ◽  
Tsutomu KOBAYASHI ◽  
Hiroki INOUE ◽  
...  
Keyword(s):  
Bisphenol A ◽  
Rat Liver ◽  
Northern Blot Analysis ◽  
Glucuronic Acid ◽  
Liver Microsomes ◽  
Male Rat ◽  
Rat Liver Microsomes ◽  
Crystalline Compound ◽  
Udp Glucuronosyltransferase

Bisphenol A, an environmental oestrogenic chemical, was found to conjugate highly with glucuronic acid in male rat liver microsomes studied in vitro. In the various isoforms tested (1A1, 1A3, 1A5, 1A6, 1A7 and 2B1), glucuronidation of bisphenol A and of diethylstilboestrol, a synthetic crystalline compound possessing oestrogenic activity and known to be glucuronidated by liver microsomes, was catalysed by an isoform of UDP-glucuronosyltransferase (UGT), namely UGT2B1, which glucuronidates some endogenous androgens. UGT activity towards bisphenol A in liver microsomes and in UGT2B1 expressed in yeast AH22 cells (22.9 and 0.58 nmol/min per mg of microsomal proteins respectively) was higher than that towards diethylstilboestrol (75.0 and 4.66 pmol/min per mg of microsomal proteins respectively). UGT activities towards both bisphenol A and diethylstilboestrol were distributed mainly in the liver but were also observed at substantial levels in the kidney and testis. Northern blot analysis disclosed the presence of UGT2B1 solely in the liver, and about 65% of the male rat liver microsomal UGT activities towards bisphenol A were absorbed by the anti-UGT2B1 antibody. These results indicate that bisphenol A, in male rat liver, is glucuronidated by UGT2B1, an isoform of UGT.

Translocon pores in the endoplasmic reticulum are permeable to small anions

2006 ◽  
Vol 291 (3) ◽  
pp. 511-517 ◽  
Author(s):  
Beáta Lizák ◽  
Ibolya Czegle ◽  
Miklós Csala ◽  
Angelo Benedetti ◽  
József Mandl ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Active Sites ◽  
Liver Microsomes ◽  
Endoplasmic Reticulum Membrane ◽  
Rat Liver Microsomes ◽  
Microsomal Enzymes ◽  
Active Centers ◽  
Udp Glucuronosyltransferase ◽  
Luminal Compartment

Contribution of translocon peptide channels to the permeation of low molecular mass anions was investigated in rat liver microsomes. Puromycin, which purges translocon pores of nascent polypeptides, creating additional empty pores, raised the microsomal uptake of radiolabeled UDP-glucuronic acid, while it did not increase the uptake of glucose-6-phosphate or glutathione. The role of translocon pores in the transport of small anions was also investigated by measuring the effect of puromycin on the activity of microsomal enzymes with intraluminal active sites. The mannose-6-phosphatase activity of glucose-6-phosphatase and the activity of UDP-glucuronosyltransferase were elevated upon addition of puromycin, but glucose-6-phosphatase and β-glucuronidase activities were not changed. The increase in enzyme activities was due to a better access of the substrates to the luminal compartment rather than to activation of the enzymes. Antibody against Sec61 translocon component decreased the activity of UDP-glucuronosyltransferase and antagonized the effect of puromycin. Similarly, the addition of the puromycin antagonist anisomycin or treatments of microsomes, resulting in the release of attached ribosomes, prevented the puromycin-dependent increase in the activity. Mannose-6-phosphatase and UDP-glucuronosyltransferase activities of smooth microsomal vesicles showed higher basal latencies that were not affected by puromycin. In conclusion, translationally inactive, ribosome-bound translocons allow small anions to cross the endoplasmic reticulum membrane. This pathway can contribute to the nonspecific substrate supply of enzymes with intraluminal active centers.

scholarly journals Evidence from rat liver nuclear preparations that latency of microsomal UDP-glucuronosyltransferase is associated with vesiculation

1980 ◽  
Vol 186 (3) ◽  
pp. 687-691 ◽  
Author(s):  
G J Wishart ◽  
D J Fry
Keyword(s):  
Electron Microscopy ◽  
Nuclear Envelope ◽  
Rat Liver ◽  
Glucuronic Acid ◽  
Udp Glucuronosyltransferase

1. Nuclear, nuclear-envelope and microsomal preparations were prepared from rat liver, and their purity and morphology monitored by electron microscopy. 2. UDP-glucuronosyltransferase activity in microsomal preparations, but not in standard nuclear or nuclear-envelope preparations, displays latency from the criterion of being enhanced (‘activated’) by a range of detergents or the endogenous activator UDP-N-acetyl-glucosamine. 3. Nuclear preparations resemble activated rather than native microsomal preparations in failing to transfer glucuronic acid from 4-nitrophenyl glucuronide to 2-aminophenol. 4. Electron microscopy indicates that membranes of nuclear preparations and of our standard nuclear-envelope preparations remain, as in vivo, in a cisternal arrangement, whereas those of microsomal preparations are vesiculated. 5. In nuclear-envelope preparations in which vesiculation has been encouraged, the transferase can be activated by detergents. 6. We suggest that latency of UDP-glucuronosyltransferase results from vesiculation of membranes during preparation and that the latency of the microsomal transferase is largely a preparative artefact.

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