Phenolsulphotransferase Activity Research Articles

Elevation of hepatic sulphotransferase activities in mice with resistance to cystic fibrosis.

The severity of intestinal disease in the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) (-/-) mice has been reported to co-segregate with gene loci which contain the genes for hydroxysteroid sulphotransferase (SULT). Because of the potential involvement of steroid hormones in CF, we investigated levels of steroid SULT activity in the livers of CFTR mice to determine whether the levels of SULT activity correlate with the occurrence or severity of CF. To elucidate the possible role of SULT activity in ameliorating the deleterious effects of CF in CFTR (-/-) mice, we determined the levels of phenol SULT (PST), hydroxysteroid SULT [dehydroepiandrosterone (DHEA)-ST] and oestrogen SULT (EST) activity in control CFTR (+/+), heterozygous CFTR (+/-) and homozygous CFTR (-/-) mice, which survive to adulthood. The level of PST activity was not significantly different between any of the groups of mice, regardless of sex or genotype. Although DHEA-ST activity was significantly higher in female mice than in male mice, there was no difference in DHEA-ST activity that could be correlated with genotype. In contrast with PST and DHEA-ST activities, we found that some male and all female adult CFTR (-/-) mice had elevated, dramatically different levels of EST from both CFTR (+/+) and CFTR (+/-) mice. Results from these SULT activity experiments were confirmed by Northern-blot analysis of mouse-liver RNA. Subsequent studies with preweanling mice revealed no differences in the levels of EST that could be correlated with genotype. Thus this study indicates that EST is elevated significantly in CFTR (-/-) mice which survive to adulthood and provides important biochemical information that EST levels may be protective in CF.

Inhibition of phenolsulphotransferase by salicylic acid: a possible mechanism by which aspirin may reduce carcinogenesis

Background—Recent epidemiological evidence has shown that chronic use of aspirin decreases susceptibility to bowel cancer. Animal studies have shown that sulphotransferase inhibitors coadministered with sulphation activated carcinogens dramatically reduce the...

Platelet phenolsulphotransferase activity in patients with dementia of Alzheimer type

Human blood platelets are more and more regarded as a reliable model of nerve cells, and some biochemical alterations found in platelet enzyme activities and receptor binding parameters may be related to analogous changes occuring in the central nervous system. Platelet phenol‐sulphotransferase activity has been evaluated in some neurological disorders, such as Parkinson's disease and headache. In the present work we determined phenolsulphotransferase activity in platelets of 36 patients with dementia of Alzheimer type, and compared such activity values with those of 36 sex‐ and age‐matched healthy controls. Patients showed a significantly higher platelet phenolsulphotransferase activity than control subjects, both with dopamine and with phenol as substrates. Moreover, a significant positive correlation was found between platelet enzyme activity and severity of illness. These findings are discussed in terms of the biochemical abnormalities found in the central nervous system of demented subjects, and in terms of the possible pathogenetic involvement of an altered phenolsulphotransferase activity (and, in general, of a modified monoamine metabolism) in such derangements of biochemical systems often occuring in dementia of Alzheimer type.

Dietary modulation of human platelet phenolsulphotransferase activity.

1. The mammalian phenolsulphotransferase enzymes are known to play a major role in both the detoxification and possibly the activation of pre-carcinogenic phenols and aromatic amines. 2. Vegetable cytosol preparations were tested in vitro for their ability to affect the sulphation of two reference compounds (rho-nitrophenol and dopamine, which are selective substrates for the phenol and monoamine forms of phenolsulphotransferase respectively), and to act as substrates for the enzymes in comparison with the same reference compounds. 3. The majority of cytosols greatly decreased (> 80%) the sulphation of either or both the reference compounds. This effect may have been due to either enzyme inhibition or substrate binding. 4. Whereas some of the cytosols were sulphated under the assay conditions, most were not. Additionally, it was found that a cytosol that decreased the sulphation of the two reference compounds was not necessarily poorly sulphated itself. 5. It is concluded that dietary factors have the potential to play a major role in modulating the sulphation detoxification pathway, and have wide ranging implications with regard to adverse drug reactions.

Reduced platelet phenolsulphotransferase activity towards dopamine and 5-hydroxytryptamine in migraine.

The sulphation of the neurotransmitters dopamine and 5-hydroxytryptamine, and of the prototypical xenobiotic 4-nitrophenol, by phenolsulphotransferases was measured in platelet homogenates prepared from a group of migraine sufferers and a group of control subjects. The activity of the M form of phenolsulphotransferase, responsible for the sulphation of dopamine and 5-hydroxytryptamine was significantly reduced in the migraine population, by 28% with dopamine as substrate and by 20% with 5-hydroxytryptamine. The activity of the P form of the enzyme towards 4-nitrophenol was the same in both groups. We also report that the selective inhibition of P form phenolsulphotransferase by red wine is much more potent than previously thought, with a 2000-fold dilution of dealcoholised red wine having the ability to inhibit sulphation by this enzyme by 50%. Our findings suggest that a reduced capacity for sulphation and inactivation of biogenic amines and catecholamines may be related to susceptibility to migraine.

Characterization of bovine tracheobronchial phenol sulphotransferase cDNA and detection of mRNA regulation by cortisol.

Phenol sulphotransferases esterify both endogenous and foreign hydroxylated aromatic compounds with sulphate. Since these enzymes participate in both hormone and drug metabolism, elucidating their regulation at both the enzymic and molecular levels may provide new understanding in several metabolic pathways. The primary structure of a bovine phenol sulphotransferase has been determined by isolation of the corresponding cDNA. Two partial bovine cDNAs were first isolated by probing a tracheal epithelial cell lambda gt11 cDNA library with a rat phenol sulphotransferase cDNA. These clones provided the sequences of the 5' and 3' ends of the predicted coding region. A contiguous cDNA was subsequently isolated by PCR using 5' and 3' oligonucleotide primers and the cDNA library as the template. The sequence of the resulting approx. 1 kbp cDNA predicted an amino acid sequence that included sequences determined for several tryptic peptides of the purified protein. Antiserum directed to a synthetic N-terminal peptide predicted by the cDNA sequence showed reactivity with the purified enzyme. High-level Trc-promoter-driven expression of the recombinant bovine enzyme was achieved in Escherichia coli. The bovine cDNA was used to determine relative steady-state levels of phenol sulphotransferase transcripts in bovine lung tissues; distal lung parenchymal RNA levels were 6-10-fold greater than those in tracheobronchial epithelium. Using a bronchial epithelial cell culture model, however, cortisol was observed to increase mRNA levels by 5-fold in both a dose- and time-dependent manner; this corresponds to previously reported glucocorticoid stimulation of phenol sulphotransferase activity in this system [Beckmann, Illig and Bartzatt (1994) J. Cell Physiol. 160, 603-610].

Effects of hypophysectomy and thyroxine on the expression of hepatic oestrogen, hydroxysteroid and phenol sulphotransferases

Sulphation in rats, and other mammals, is carried out by a family of sulphotransferase isoenzymes, which can be further subdivided into oestrogen, hydroxysteroid and phenol sulphotransferases. We have examined the effects of hypophysectomy on the activity and expression of representative members of the three major sulphotransferase sub-families in male Wistar rat liver cytosols, and have found that the different sub-families are subject to differential regulation by pituitary hormones. Our data show that in male rat liver hydroxysteroid sulphotransferases activity was increased, oestrogen sulphotransferases activity was not altered and phenol sulphotransferases activity was reduced. Further, we have studied the effect on sulphotransferase expression of administration of thyroxine and dexamethasone to hypophysectomized rats. Treatment of hypophysectomized rats with thyroxine virtually abolished oestrogen sulphotransferase activity in male rat liver but had no effect on hydroxysteroid sulphotransferase or phenol sulphotransferase activity. Treatment of hypophysectomized rats with dexamethasone had no effect on sulphotransferase activities. Quantitative immunoblot analysis of liver cytosols showed that these changes in enzyme activity were related to changes in levels of the respective enzyme proteins.

Interindividual variability of phenol- and catechol-sulphotransferases in platelets from adults and newborns.

1 Phenol- and catechol- sulphotransferase activities were measured with p-nitrophenol and dopamine as substrates in platelets obtained from 100 newborns and 100 healthy adults. 2 Mean +/- (s.d.) estimates of catechol sulphotransferase activity were 7.07 +/- 5.93 (adult) and 13.3 +/- 6.42 (newborn) pmol min(-1) mg(-1) protein, respectively (P < 0.001). The coefficients of variation were 84% (adult) and 48% (newborn). The frequency distribution of sulphotransferase activity was symmetric and did not deviate significantly from normality in newborn platelets, but was positively skewed in adult platelets. 3 Mean +/- (s.d.) estimates of phenol sulphotransferase activity were 3.01 +/- 3.21 (adult) and 4.80 +/- 4.34 (newborn) pmol min(-1) mg(-1), respectively (P < 0.001). The coefficients of variation were 107% (adult) and 90% (newborn). The frequency distribution of sulphotransferase activity was positively skewed in both newborn and adult platelets. 4 Since sulphotransferase activity in platelets is well-expressed at birth and a prenatal development of sulphotransferase has been described in mid-gestational human foetal liver, the newborn should be able to sulphate drugs.

Sensitive assays for the determination of monoamine oxidase and phenol sulphotransferase activity in small tissue samples

A method to assay phenol Sulphotransferase (PST) and monoamine oxidase (MAO) in brain (anterior pituitary gland, hypothalamus) and liver specimens as small as 4 mg is described. The specimens were homogenized (sonicated) in various volumes of a buffer, the smallest being 100 μL, to obtain the homogenates. MAO assay was carried out using 30 μL of the homogenate and for PST assay, 30 μL of either the homogenate or, in the case of liver, the supernatant (100,000 × g for 60 min). The radiolabeled products of the enzymatic reactions were separated from the radiolabeled substrates by high-pressure liquid chromatography (HPLC) and the radioactivity of the eluted products measured directly by a radioisotope detector coupled to the HPLC system. The constraint of the assay protocol was not the weight of the specimens but the volume of buffer used in the preparation of the homogenate. Although 100 μL was a convenient working volume, the tissue can also, with care, be sonicated in a 50 μL buffer. With extremely small specimens, weighed fractions of the specimens could be sonicated directly in the control and experimental incubation mixtures bypassing the preparation of the homogenate. Thus, the overall method offers, for the first time, a reliable and adaptable means for measuring MAO and PST in small to extremely small tissue specimens.

Cytosolic phenol and steroid sulphotransferase activities are decreased in a sex-dependent manner in streptozotocin-induced diabetic rats

Cytosolic phenol and steroid sulphotransferase activities are decreased in a sex-dependent manner in streptozotocin-induced diabetic rats

Correlation of phenol sulphotransferase activities in the liver and platelets of rat

Phenol sulphotransferase (PST) activity in rat platelet cytosol for p-nitrophenol (PNP) sulphation was found to have a similar dependence on PNP concentration and thermostability to that in rat liver cytosol. The activities of PST isoenzyme for the sulphation in the microM range of PNP in rat platelets and rat liver were significantly correlated. Thus, measurement of PST activity in platelets could be a useful and practical method for predicting this activity in liver.

Sulphotransferase-mediated activation of the carcinogen 5-hydroxymethyl-chrysene: Species and sex differences in tissue distribution of the enzyme activity and a possible participation of hydroxysteroid sulphotransferases

Sulphation of the carcinogen 5-hydroxymethyl-chrysene (5-HCR) to the active metabolite 5-HCR sulphate occurred at significant rates in all of hepatic cytosols prepared from the male and female experimental animals, rats, mice, guinea-pigs and hamsters. The 5-HCR-sulphating activity was also found in kidney cytosols of all the experimental animals used, while their activities were much less than those of hepatic cytosols. In the male mice, the enzyme activity of testis was higher than any other examined tissue. Small intestine and adrenal of male and female guinea-pigs had relatively high enzyme activities. Small enzyme activities were also found in a variety of extrahepatic tissues of some of these animals. Marked species and sex differences (female ⪢ male in the rat and mouse) were observed in the hepatic enzyme activity. In the female rat liver which showed the highest 5-HCR-sulphating activity among the examined tissues of all the animals, a typical hydroxysteroid sulphotransferase inhibitor, dehydroepiandrosterone (DHA) sulphate (1 mM), potently and competitively inhibited the sulphation of 5-HCR as well as that of DHA, a typical substrate for hydroxysteroid sulphotransferases. On the contrary, the phenol sulphotransferase inhibitors, pentachlorophenol and 2,6-dichloro-4-nitrophenol, had only a little effect on these enzyme activities even at a concentration of 50 μM that showed a potent inhibition of the phenol sulphotransferase activity. These results suggest that 5-HCR be sulphated in the female rat liver by hydroxysteroid sulphotransferases, but not by phenol sulphotransferases.

Platelet phenolsulphotransferase activity and 'abdominal migraine'.

Low platelet phenolsulphotransferase activity has been reported in adult patients with dietary sensitive migraine. Platelet activity of this enzyme was therefore measured in children having 'abdominal migraine' with probable dietary trigger and in controls. No significant difference was found in activity between the two groups. There was no significant correlation between platelet phenolsulphotransferase activity and age.

Determination of phenol sulphotransferase activity by high-performance liquid chromatography

Determination of phenol sulphotransferase activity by high-performance liquid chromatography

Phenolsulphotransferase activity in the developing mosquito ( Aedes togoi)

Phenolsulphotransferase (PST) activity was measured with N-acetyldopamine (NADA) and harmol as substrates in the larvae, pupae and adult mosquito ( Aedes togoi). Only the newly emerged pupae showed high PST activity 1–4 hr after pupation. PST activity could also be measured in each individual pupa, with the female exhibiting significantly higher specific activity (30 ± 3.7 pmol NADA [ 35S]sulphate/min per mg protein) than the males (13.6 ± 2.9 pmol NADA [ 35S]sulphate/min per mg protein). The optimum pH for the PST reaction was 9.0. The K m values for [ 35S]PAP were 0.55 and 2.5 μM when measured with NADA and harmol as acceptors, respectively; the corresponding K m values for these two substrates were 2.61 and 16.1 μM. Studies with 2,6-dichloro-4-nitrophenol showed a dose-dependent inhibition of PST. Sulphate conjugation of NADA from ATP and sodium [ 35S]sulphate was also demonstrated with pupal extracts, with pH optimum between 8.6 and 9.0. The specific activity of this overall sulphate conjugation, measured in the female pupal extract was 5.08 pmol NADA [ 35S]sulphate/min per mg protein and 1.68 pmol harmol [ 35S]sulphate/min per mg protein. The importance and possible function of sulphate conjugation of NADA in insects is discussed.

Assay of phenol sulphotransferase in human blood

An assay of phenol sulphotransferase in human blood with 4-hydroxy-3-metho-xyphenylethylene glycol as partial substrate is described. The pH optimum with this substrate is 8.0. To demonstrate the heterogeneity of the enzyme in blood an alternative partial substrate, 4-methylumbelliferone, at an optimal pH of 7.4, is used. Enzyme activity is linear with time up to at least 60 minutes of incubation. With 4-hydroxy-3-methoxyphenylethylene glycol as the substrate a linear relation between phenol sulphotransferase activity and enzyme concentration exists. Phenol sulphotransferase activity shows no correlation with age or sex of healthy subjects. Varying ratios of the enzyme activities toward both substrates support the idea of two phenol sulphotransferases in blood. As nevertheless a significant correlation exists between both sulphotransferase activities we conclude that neither substrate is sulphated exclusively by one of the two enzymes. Because preparative procedures for platelets may lead to considerable losses of specific subpopulations with relatively low or high enzyme activity, one of the advantages of using whole blood is that the activity in the whole platelet population is determined.

Human platelet phenolsulphotransferase M and P: substrate specificities and correlation with in vivo sulphoconjugation of paracetamol and salicylamide.

Human platelet phenolsulphotransferase exists in two functional forms. M and P. In this study the substrate specificity of the two forms has been further delineated by correlating activities in different individuals with various substrates. m-Tyramine, noradrenaline, adrenaline, 5-hydroxytryptamine, p-hydroxyamphetamine, isoprenaline, salbutamol and l-naphthol were all specific substrates for the M form of the enzyme. Paracetamol, a mixed substrate, was predominantly metabolized by the M form. Salicylamide at 5 microM was a substrate for the P form but became and M substrate at higher concentration. Phenol itself, a specific substrate for phenolsulphotransferase P at 10 microM, also became an M substrate at 1 mM concentration. These substrate specificities were confirmed with the selective inhibitor, dichloronitrophenol. In this study, we measured phenolsulphotransferase activity in platelets from 13 individuals selected on the basis of their wide variation in ability to sulphoconjugate paracetamol and salicylamide in vivo. There was no significant relationship between the in vivo pattern with either drug and the activity of platelet phenolsulphotransferase assayed with paracetamol or salicylamide respectively.

Platelet phenolsulphotransferase activity in Parkinson's disease.

Mean platelet phenolsulphotransferase activity was significantly increased in L-dopa-treated parkinsonian patients compared with controls. The degree of rise was associated with dose of and duration of treatment with L-dopa and may thus represent an adaptive increase to circulating dopamine. Alternatively, the increase may reflect changes supervening with duration of illness.

Sulphate conjugation of p-hydroxytriamterene by platelet phenol sulphotransferase: assay conditions and correlation with metabolism in man.

1 Sulphate conjugation catalyzed by phenol sulphotransferase (PST) is an important pathway in the metabolism of many drugs including triamterene. Variations in PST activity in an easily obtained tissue such as the platelet might reflect individual differences in the sulphate conjugation in other organs and tissues. Human platelets contain at least two forms of PST, a thermolabile (TL) form for which dopamine is a substrate and a thermostable (TS) form for which low concentrations of p-nitrophenol serve as a substrate. 2 p-OH-triamterene, the major metabolite of triamterene, is conjugated with sulphate in vivo. p-OH-triamterene was a substrate for platelet PST with an apparent Michaelis-Menten value of 26 microM. Thermal stability studies indicated that p-OH-triamterene was a substrate for only the TS form of platelet PST. 3 When platelet homogenates from 29 individual subjects were tested, there was a significant correlation between PST activities measured with 4 microM p-nitrophenol and with p-OH-triamterene (r = 0.985, P less than 0.0001) but not between activities measured with dopamine and with p-OH-triamterene (r = 0.023, P greater than 0.2). These results confirmed that p-OH-triamterene was a substrate for only the TS form of human platelet PST. 4 The same 29 subjects were treated with 1 mg/kg of triamterene orally. 24-h urinary excretions of triamterene, p-OH-triamterene and p-OH-triamterene sulphate averaged 15.3%, 6.3% and 78.4%, respectively, of the total of triamterene plus measured metabolites excreted. The excretion of triamterene plus the two metabolites averaged 43.1 +/- 2.6% (mean +/- s.e. mean) of the ingested dose. There was not a significant correlation between the proportion of p-OH-triamterene excreted as sulphate conjugate and the activities of either the TS or TL forms of platelet PST activity.

Alpha-Methyldopa, alpha-methyldopamine an alpha-methylnoradrenaline: substrates for the thermolabile form of human platelet phenol sulphotransferase.

1 Sulphate conjugation catalyzed by phenol sulphotransferase (PST) is an important pathway in the catabolism of alpha-methyldopa (MD). Variations in PST activity in an easily obtained tissue such as the human platelet might reflect individual differences in the sulphate conjugation of MD in other organs and tissues. There are at least two forms of human platelet PST, a thermolabile form for which dopamine is a substrate and a thermostable form for which low concentrations of phenol can serve as a substrate. 2 MD, alpha-methyldopamine (MDA) and alpha-methylnoradrenaline (MNA) were tested as substrates for human platelet PST. All three were substrates for the thermolabile form of the enzyme and none were substrates for the thermostable form of PST. Apparent Michaelis-Menten (Km) values for MD, MDA and MNA were 5.5, 0.014 and 0.28 mM, respectively. Apparent Km values for 3'-phosphoadenosine-5'-phosphosulphate, the sulphate donor for the reaction, were 0.08, 0.13 and 0.10 microM, respectively, for the three catechol substrates. The pH optima for the reaction were 7.5 for MD and 6.5 for both MDA and MNA. 3 When platelet homogenates from 20 individual subjects were tested, there were significant correlations between PST activities measured with dopamine and those measured with MD, MDA and MNA (r = 0.54, 0.98 and 0.93, P less than 0.02, less than 0.001, and less than 0.001, respectively), but not between activities measured with low concentrations of phenol and those measured with MD, MDA and MNA (r = 0.021, 0.045 and 0.046, respectively). There results were also compatible with the conclusion that MD, MDA and MNA were substrates for the thermolabile form of platelet PST. 4 These observations will make it possible to test the hypothesis that variations in the activity of the thermolabile form of platelet PST may reflect individual differences in the sulphate conjugation of MD, MDA and MNA.