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:3.1.6.1 (sulfatase)
3,205 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Dihydrodiol dehydrogenase (DD; EC 1.3.1.20) purified to homogeneity from rat liver cytosol will catalyze the NAD(P)(+)-dependent oxidation of (+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (B[a]P-diol) to yield benzo[a]pyrene-7,8-dione (BPQ). To verify that BPQ is a metabolite of B[a]P-diol in rat liver, an S100 fraction was supplemented with NAD+ and NADP+, and the formation of BPQ was followed by reverse-phase HPLC. The identity of BPQ was established by co-chromatography with an authentic standard (under different solvent conditions) and by RP-HPLC using a diode-array detector which established that the metabolite shared spectral identity with BPQ. The formation of BPQ in the S100 fraction was blocked by either a competitive inhibitor (indomethacin) or a suicide substrate [1-(4-nitrophenyl)-propen-1-ol] for DD, indicating that BPQ was being formed by this enzyme. To assess the contribution of DD to the metabolism of [3H]B[a]P-diol, subcellular fractions obtained from uninduced rat liver were fortified with co-factors to optimize the activity of enzymes that would compete for this proximate carcinogen. Under these conditions, S100 fractions fortified with NAD+ and NADP+ metabolized 25% of the B[a]P-diol, producing 731 +/- 154 pmol of BPQ. In contrast, rat liver microsomes fortified with an NADPH generating system metabolize 75% of the B[a]P-diol producing 2614 +/- 379 pmoles of benzo[a]pyrene-tetrahydrotetrols. Rat liver homogenates (S10) fortified with either uridine diphosphoglucuronic acid or phosphoadenosine phosphosulfate produced 180 +/- 56 and 95 +/- 31 pmoles of conjugates respectively, which were recovered as B[a]P-diol after treatment of the aqueous phase with either beta-glucuronidase or aryl sulfatase. Of the metabolites analyzed BPQ was formed in the second largest amount. These studies show that in uninduced rat liver DD may play a significant role in the metabolism of B[a]P-diol. The metabolic fate of BPQ remains to be determined.
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PMID:Contribution of dihydrodiol dehydrogenase to the metabolism of (+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene in fortified rat liver subcellular fractions. 139 42

The sulfate conjugate of the model compound 4-methylumbelliferone was taken up and hydrolyzed considerably more rapidly by isolated hepatocytes than was the glucuronide conjugate. Using intact hepatocytes or homogenates of hepatocytes, compounds were identified that either inhibited 4-methylumbelliferyl sulfate hydrolysis via arylsulfatase or impaired its uptake into cells. For example, sodium sulfate inhibited hydrolysis of 4-methylumbelliferyl sulfate by intact hepatocytes (half-maximal inhibition, 0.1 mM) but not by homogenates, suggesting a selective action on organic sulfate uptake at the plasma membrane. In contrast, cholesterol sulfate inhibited hydrolysis of 4-methylumbelliferyl sulfate by homogenates but not by hepatocytes, consistent with the hypothesis that cholesterol sulfate does not readily enter intact cells. Compounds that inhibited hydrolysis of 4-methylumbelliferyl sulfate by both isolated hepatocytes and microsomes include sodium sulfite (half-maximal inhibition, 0.1 mM), pregnenolone sulfate (half-maximal inhibition, 1 microM), and estrone sulfate (half-maximal inhibition, 10 microM). To test whether production of sulfate conjugates could be modified by agents affecting arylsulfatase in intact hepatocytes, we examined the effects of pregnenolone sulfate on the production of 4-methylumbelliferyl sulfate from 4-methylumbelliferone. Addition of pregnenolone sulfate (100 microM) to intact cells increased rates of 4-methylumbelliferone sulfate production and decreased the fraction of 4-methylumbelliferone converted into the glucuronide. Hydrolysis of 4-methylumbelliferyl sulfate by isolated microsomes was inhibited in a dose-dependent manner by adenosine 3'-phosphate 5'-phosphosulfate (PAPS) when cytosol, a source of sulfotransferase was present. Furthermore, addition of low concentrations of PAPS (0.5 microM) to a reconstituted system of microsomes and cytosol impaired the formation of fluorescent product from 4-methylumbelliferyl sulfate until PAPS was consumed, indicating that futile cycling via arylsulfatase and sulfotransferase occurred. Subsequent futile cycling of free 4-methylumbelliferone and 4-methylumbelliferyl sulfate occurred upon repeated additions of PAPS and was prevented by sodium sulfite, an inhibitor of arylsulfatase. These results argue strongly that sulfate conjugate production within hepatocytes is regulated by futile cycling via sulfotransferase and arylsulfatase. Thus, drugs and endogenous substances that affect arylsulfatase may have marked effects on sulfate conjugate production by the liver.
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PMID:Futile cycling of a sulfate conjugate by isolated hepatocytes. 200 78

An enzyme which catalyzes the transfer of sulfate from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to gastrin (G17) was identified in rat gastric mucosal cells. The enzyme activity was detected in the 105,000xg supernatant fraction. Formation of gastrin sulfate was shown by using 125I-gastrin and non-radioactive PAPS. The product was sensitive to acid hydrolysis, arylsulfatase treatment and removed by gastrin antibody, but not changed by treatments with chondro-4-sulfatase and chondro-6-sulfatase. The product had a molecular weight of 2050 daltons, close to the molecular weight of G17 sulfate, and, therefore, indicating the sulfated product is not APS derived from the degradation of PAPS. The enzyme activity showed a Km value of 5 microM for PAPS and a pH optimum of 6.0. The activity was not detected in the liver preparation.
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PMID:Enzymatic sulfation of gastrin in rat gastric mucosa. 239 84

The capacity of polynuclear aromatic hydrocarbons to elicit profound effects on the development of avian aortic atheromata has raised questions regarding the biotransformation of polynuclear aromatic hydrocarbons in the target (aortic) tissue. Results of this investigation demonstrate the capacity of aortic enzymes to affect the sulfoconjugation of 3-hydroxy-benzo(a)pyrene and describe several characteristics of the aortic sulfotransferase activity. Conjugating activities measured in avian aortic tissues were approximately 10-20% of those assayed in corresponding preparations of avian hepatic tissues under the same reaction conditions. Activities were measured in homogenates, in a series of homogenate subfractions, in whole organ cultures, in cultured aortic endothelial cells, and in cultured aortic smooth muscle cell preparations. Sulfoconjugation was localized in the cytosolic fraction and kinetics in this fraction yielded a range of apparent Km values from 9 to 16 microM (mean = 11.8 +/- 3.1, n = 4) and a range of apparent Vmax values from 281 to 457 pmol/mg of protein/30 min (mean = 360 +/- 49, n = 4). Abdominal and thoracic segments of the aorta exhibited virtually identical specific activities. Also, activities assayed in cultured aortic smooth muscle cells were similar to those measured in cultured aortic endothelial cells. Capacity to generate adenosine 3'-phosphate 5'-phosphosulfate (PAPS) appeared to limit the reaction rate as judged by comparative investigations with PAPS and a PAPS-generating system. Aortic sulfatases actively hydrolyzed benzo(a)pyrene-3-O-sulfate. The sulfatase activity appeared to partially mask sulfotransferase activities measured in organ and cell culture preparations and in particulate subfractions of cellular homogenates.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Phase II biotransformation of carcinogens/atherogens in cultured aortic tissues and cells. I. Sulfation of 3-hydroxy-benzo(a)pyrene. 287 26

The mechanism by which various chemicals induce renal cystic disease is unknown. To examine the early events in cystogenesis the ultrastructure and biochemistry of liver and kidney were analyzed after the administration of a chemical that induces renal cyst formation. Special emphasis was placed on examining potential mechanisms that would account for the observed loss of extracellular proteoglycans. Renal cystic disease was chemically induced in rats by feeding 2-amino-4,5-diphenylthiazole (DPT) for up to 4 weeks. After 4 days of feeding, DPT had induced a 4-fold increase in total urine output relative to diet-restricted control groups. Both groups maintained, but did not gain, weight during the feeding schedule. Cyst formation was localized to the medullary collecting tubules. Relative to diet-restricted controls, rats fed DPT exhibited diminished renal and hepatic catalase activity, but elevated activity for UDP-glucuronosyltransferase. Medulla showed an increase in the specific activities of the enzymes galactosyltransferase and sulfatase B. These enzymological findings correlated with ultrastructural observations of a loss of peroxisomes, proliferation of endoplasmic reticulum and enlargement of the golgi apparatus. Serum and urinary levels of inorganic sulfate were significantly increased in DPT-fed rats relative to controls. Tissue levels of UDP-glucuronic acid and adenosine 3'-phosphate 5'-phosphosulfate were not depressed by DPT feeding. Thus, DPT-induced cyst formation and loss of staining for glycosaminoglycans does not involve gross depletions of UDP-glucuronic acid and adenosine 3'-phosphate 5'-phosphosulfate, mutual cosubstrates for Phase II drug conjugation reactions and glycosaminoglycan synthesis.
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PMID:Diphenylthiazole-induced changes in renal ultrastructure and enzymology: toxicologic mechanisms in polycystic kidney disease? 311 18

In vitro O-methylation of 4-hydroxyestrone monosulfates has been examined by means of high-performance liquid chromatography with electrochemical detection. When 4-hydroxyestrone or its 3-sulfate was incubated with rat liver homogenate in the presence of S-adenosyl-L-methionine, the 4-methyl ether was formed in twelve times larger amount than the 3-methyl ether. 4-Hydroxyestrone 4-sulfate served for O-methylation to much less extent as a substrate namely, only a small amount of the 4-methyl ether was formed. Enzymic sulfation of 4-hydroxyestrone with rat liver 105000 g supernatant fortified with adenosine 3'-phosphate 5'-phosphosulfate provided solely catechol estrogen 4-sulfates. The participation of catechol O-methyltransferase and aryl sulfatase in the formation of guaiacol estrogens has been discussed.
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PMID:In vitro O-methylation of 4-hydroxyestrone monosulfates. 712 41

Net sulfation of 4-methylumbelliferone in intact hepatocytes is regulated, in part, by substrate cycling between sulfotransferases (SULT) and arylsulfatases (ARS). Thus, ARS have the potential to influence rates of net sulfate conjugation of a variety of compounds in intact cells via interaction with SULT. Unlike ARSA and ARSB, which are lysosomal, steroid sulfate sulfatase (ARSC, also known as STS) is localized exclusively in the endoplasmic reticulum (ER). The present study was designed to assess the existence and extent of substrate cycling between steroids and their sulfate conjugates through ARSC and SULT, and also to initiate studies of the topology of the catalytic site of ARSC in the rat liver ER. Addition of rat liver microsomes to cytosol and 3'-phosphoadenosine 5'-phosphosulfate (PAPS) reduced rates of sulfation of dehydroepiandrosterone (DHEA) by SULT, and similarly hydrolysis of DHEA sulfate (DHEAS) was reduced when recombinant human hydroxysteroid SULT was added to rat liver microsomes in the presence of PAPS. There was no evidence for ARSC latency in the presence of detergent at either 4 or 37 degrees C, indicating that facilitated transport of steroid sulfates across the ER membrane may not be required for ARSC activity. The effect of proteases on ARSC activity in intact and disrupted microsomes was determined and compared with effects on components of the glucose-6-phosphatase system known to be localized on the lumenal and cytoplasmic surfaces of the ER. In contrast to the components of the glucose-6-phosphatase system, activity of ARSC in both intact and disrupted microsomes was substantially more resistant to protease inactivation. Our results indicate that substrate cycling of steroids and their sulfates does occur, and suggest that the active site of ARSC may be located within the ER membrane.
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PMID:Microsomal steroid sulfatase: interactions with cytosolic steroid sulfotransferases. 956 44

The intestinal bioavailability and biotransformation of 3-hydroxybenzo(a)pyrene, a major metabolite of benzo(a)pyrene in many animal species, was investigated in an in situ isolated intestinal preparation from the channel catfish, and in vitro with preparations of catfish intestine and blood. 3-Hydroxybenzo(a)pyrene was a good substrate for adenosine 3'-phosphate 5'-phosphosulfate (PAPS)-sulfotransferase and UDP-glucuronosyltransferase in cytosol or microsomes prepared from intestinal mucosa. The benzo(a)pyrene-3-glucuronide and 3-sulfate conjugates were only very slowly hydrolyzed by intestinal beta-glucuronidase and sulfatase. The K(m) values for PAPS-sulfotransferase and UDP-glucuronosyltransferase were 0.4 and 1 microM, respectively, and V(max) were 1.61 +/- 1.08 nmol benzo(a)pyrene-3-sulfate/min/mg of cytosolic protein and 1.08 +/- 0.54 nmol benzo(a)pyrene-3-glucuronide/min/mg of microsomal protein. Hydrolytic enzyme activities were three orders of magnitude slower. In the in situ intestinal preparation, [(3)H]3-hydroxybenzo(a)pyrene was readily metabolized to the glucuronide and sulfate conjugates. After 1 h of incubation of 2 or 20 microM [(3)H]3-hydroxybenzo(a)pyrene in the in situ preparation, the luminal contents contained 3-hydroxybenzo(a)pyrene, benzo(a)pyrene-3,6-dione, benzo(a)pyrene-3-sulfate, and benzo(a)pyrene-3-glucuronide. Mucosal samples contained these components, as well as some unextractable material. The blood contained mainly benzo(a)pyrene-3-sulfate and an as yet unidentified metabolite of 3-hydroxybenzo(a)pyrene bound to hemoglobin. Some, but not all, blood samples contained small amounts of 3-hydroxybenzo(a)pyrene, benzo(a)pyrene-3-glucuronide, and benzo(a)pyrene-3,6-dione. These studies demonstrate the rapid phase 2 conjugation of a phenolic benzo(a)pyrene metabolite in intestinal mucosa, and the transfer of the phase 2 sulfate and glucuronide conjugates to blood.
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PMID:Intestinal bioavailability and biotransformation of 3-hydroxybenzo(a)pyrene in an isolated perfused preparation from channel catfish, Ictalurus punctatus. 1130 39

Heparan sulfate d-glucosaminyl 3-O-sulfotransferases (3-OSTs) catalyze the transfer of sulfate from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to position 3 of the glucosamine residue of heparan sulfate and heparin. A sixth member of the human 3-OST family, named 3-OST-5, was recently reported (Xia, G., Chen, J., Tiwari, V., Ju, W., Li, J.-P., Malmstrom, A., Shukla, D., and Liu, J. (2002) J. Biol. Chem. 277, 37912-37919). In the present study, we cloned putative catalytic domain of the human 3-OST-5 and expressed it in insect cells as a soluble enzyme. Recombinant 3-OST-5 only exhibited sulfotransferase activity toward heparan sulfate and heparin. When incubated heparan sulfate with [35S]PAPS, the highest incorporation of35S was observed, and digestion of the product with a mixture of heparin lyases yielded two major35S-labeled disaccharides, which were determined as DeltaHexA-GlcN(NS,3S,6S) and DeltaHexA(2S)-GlcN(NS,3S) by further digestion with 2-sulfatase and degradation with mercuric acetate. However, when used heparin as acceptor, we identified a highly sulfated disaccharide unit as a major product. This had a structure of DeltaHexA(2S)-GlcN(NS,3S,6S). Quantitative real-time PCR analysis revealed that 3-OST-5 was highly expressed in fetal brain, followed by adult brain and spinal cord, and at very low or undetectable levels in the other tissues. Finally, we detected a tetrasulfated disaccharide unit in bovine intestinal heparan sulfate. To our knowledge, this is the first report to describe not only the natural occurrence of tetrasulfated disaccharide unit but also the enzymatic formation of this novel structure.
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PMID:Characterization of a heparan sulfate 3-O-sulfotransferase-5, an enzyme synthesizing a tetrasulfated disaccharide. 1274 Mar 61

A sensitive fluorometric assay was developed for alcohol sulfotransferase (AST). This was the first continuous fluorometric assay reported for AST. It used 3'-phosphoadenosine 5'-phosphosulfate regenerated from 3-phosphoadenosine 5'-phosphate by a recombinant phenol sulfotransferase (PST) using 4-methylumbelliferyl sulfate as the sulfuryl group donor. The recombinant PST did not use the alcohol substrate under the designed condition, and the sensitivity for AST activity was found to be comparable to that of radioactive assay as reported in the literature. The change of fluorescence intensity of 4-methylumbelliferone corresponded directly to the amount of active AST and was sensitive enough to measure nanogram or picomole amounts of the enzyme activity. This fluorometric assay was used to determine the activities of AST as purified form and in crude extracts of pig liver, rat liver, and Escherichia coli. Some properties of human dehydroepiandrosterone sulfotransferase were determined by this method and were found to be comparable to published data. Under similar assay conditions, the contaminated activities of arylsulfatase in crude extracts were also determined. This method not only is useful for the routine and detailed kinetic study of this important class of enzymes but also has the potential for the development of a high-throughput procedure using microplate reader.
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PMID:Fluorometric assay for alcohol sulfotransferase. 1576 10


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