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M W Duffel

Publications and source records attributed to M W Duffel.

47 records · Page 3Linked to original sources
PubMed1981

On the mechanism of aryl sulfotransferase.

Aryl sulfotransferase IV (EC 2.8.2.1), purified to homogeneity from male rat liver, catalyzes the sulfation of a variety of substituted phenols, including catecholamines, tyrosine esters, and peptides containing NH2-terminal tyrosine residues. An investigation of the mechanism of the enzyme was carried out using 2-chloro-4-nitrophenol as a model substrate. Kinetic, inhibition, and binding studies with aryl sulfotransferase IV are all consistent with a random rapid equilibrium Bi Bi kinetic mechanism with two dead end product inhibitor complexes. Studies of the chemical mechanisms of the enzyme-catalyzed reaction demonstrate that electron-withdrawing substituents decrease the maximal velocity of phenol sulfation. The maximal velocity of the reaction correlates with Hammett sigma p- constants (rho = -0.25). Evidence is presented for the mechanism by which adenosine 3',5'-bisphosphate and aryl sulfotransferase catalyze the transfer of sulfate from 2-chloro-4-nitrophenyl sulfate to other phenols.

Animals↗
PubMed1979

Studies on the nature and regulation of the cellular thio:disulphide potential.

Microsomal fractions separated from homogenates of liver, kidney and corpora lutea contain a monooxygenase (dimethylaniline monooxygenase [N-oxide forming], EC 1.14.13.8) that catalyses NADPH- and oxygen-dependent oxidation of cysteamine to cystamine. The monooxygenase purified to homogeneity from hog liver also catalyses oxygenations of diverse xenobiotics, but it does not catalyse oxidation of any other physiological sulphur- or nitrogen-containing compounds. All the available evidence indicates that cysteamine is the physiological substrate for the monooxygenase, and the oxidation of this thiol to the disulphide may be a significant source of disulphide maintaining the cellular thiol:disulphide potential. The concentration of protein-low molecular weight mixed disulphide is a function of this potential. Changes in concentration of this protein-mixed disulphide reflect changes in thiol:disulphide balance. At constant substrate concentrations the potential would depend primarily on activity of the cytosol glutathione reductase (NAD(P)H: oxidized-glutathione oxidoreductase, EC 1.6.4.2) relative to that of the membrane-bound monooxygenase. In hepatic tissue from adult mice and hamsters there is a correlation between the concentration of protein-mixed disulphide and the activity of the monooxygenase relative to the reductase. Hepatic glutathione reductase is relatively constant in mice, but the monooxygenase is much higher in the female than in the male. After gonadectomy monooxygenase activity decreases in the female and increases in the male. Activities are restored to control levels by treating males with testosterone and females with progesterone. Testosterone decreases and progesterone increases activity. These two hormones apparently regulate the level of this enzyme in hepatic tissue.

Animals↗
PubMedDate not supplied

Leurosine biotransformations: an unusual ring-fission reaction catalyzed by peroxidase.

The dimeric Vinca alkaloid leurosine undergoes an unusual fission of the piperidine ring of the Iboga substructure when reacted with horseradish peroxidase and hydrogen peroxide. A preparative-scale oxidation of leurosine provided 15'-hydroxycatharinine, which was identified by infrared and proton and carbon-13 NMR spectroscopies and high-resolution mass spectrometry. A proposed pathway for the formation of 15'-hydroxycatharinine involves radical and iminium intermediates and cleavages of a putative diol. The enzymatic transformation product is 3 orders of magnitude less active than leurosine or vinblastine in vitro, in inhibiting the polymerization of tubulin.

Animals↗
PubMedDate not supplied

Influence of substrate structure on the catalytic efficiency of hydroxysteroid sulfotransferase STa in the sulfation of alcohols.

Sulfotransferase a (STa) is an isoform of hydroxysteroid (alcohol) sulfotransferase that catalyzes the formation of sulfuric acid esters from both endogenous and xenobiotic alcohols. Among its various functions in toxicology, STa is the major form of hepatic sulfotransferase in the rat that catalyzes the formation of genotoxic and carcinogenic sulfuric acid esters from hydroxymethyl polycyclic aromatic hydrocarbons. The goal of the present study was to elucidate fundamental quantitative relationships between substrate structure and catalytic activity of STa that would be applicable to these and other xenobiotics. We have modified previous procedures for purification of STa in order to obtain sufficient amounts of homogeneous enzyme for determination of kcat/Km values, a quantitative measure of catalytic efficiency. We determined the catalytic efficiency of STa with benzyl alcohol and eight benzylic alcohols that were substituted with n-alkyl groups (CnH2n + 1, where n = 1-8) in the para position, and the optimum value for kcat/Km in these reactions was obtained with n-pentylbenzyl alcohol. Correlations between logarithms of kcat/Km values and logarithms of partition coefficients revealed that hydrophobicity of the substrate was a major factor contributing to the catalytic efficiency of STa. Primary n-alkanols (CnH(2n+1)OH, where n = 3-16) exhibited an optimum kcat/Km for C9-C11 and a linear decrease in vmax of the reaction for C3-C14; 15- and 16-carbon n-alkanols were not substrates for STa. These results indicated limits to the length of the extended carbon chain in substrates. Such limits may also apply to hydroxysteroids, since cholesterol was inactive as either substrate or inhibitor of STa. Furthermore, the importance of steric effects on the catalytic efficiency of STa was also evident with a series of linear, branched, and cyclic seven-carbon aliphatic alcohols. In conclusion, our results provide fundamental quantitative relationships between substrate structure and catalytic efficiency that yield insight into the specificity of STa for both endogenous and xenobiotic alcohols.

9,10-Dimethyl-1,2-benzanthracene↗
PubMedDate not supplied

Enzymatic aspects of the phenol (aryl) sulfotransferases.

The sulfotransferases that are active in the metabolism of xenobiotics represent a large family of enzymes that catalyze the transfer of the sulfuryl group from 3'-phosphoadenosine 5'-phosphosulfate to phenols, to primary and secondary alcohols, to several additional oxygen-containing functional groups, and to amines. Restriction of this review to the catalytic processes of phenol or aryl sulfotransferases does not really narrow the field, because these enzymes have overlapping specificity, not only for specific compounds, but also for multiple functional groups. The presentation aims to provide an overview of the wealth of phenol sulfotransferases that are available for study but concentrates on the enzymology of rat and human enzymes, particularly on the predominant phenol sulfotransferase from rat liver. The kinetics and catalytic mechanism of the rat enzyme is extensively reviewed and is compared with observations from other sulfotransferases.

Animals↗
PubMedDate not supplied

Inhibition of aryl sulfotransferase by carboxylic acids.

Aryl sulfotransferase (AST) IV catalyzes the 3'-phosphoadenosine-5'-phosphosulfate-dependent formation of sulfuric acid esters of a wide range of phenols, benzylic alcohols, hydroxamic acids, catecholamines, tyrosine carboxylesters, and peptides with N-terminal tyrosines. The objective of this investigation was to determine whether aryl carboxylic acids could act either as substrates or inhibitors for AST IV. These studies were conducted with AST IV that was purified to homogeneity from male Sprague-Dawley rats. Although none of the carboxylic acids tested were substrates for AST IV, they did competitively inhibit the enzyme with 1-naphthalenemethanol as substrate at pH 7.0. 1-Naphthoic acid, 2-naphthoic acid, and salicylic acid were particularly effective inhibitors of the sulfotransferase. The distance of the carboxyl group from the aromatic ring influenced the inhibitory capability of the carboxylic acids examined. The Kis for 1-naphthylacetic acid and 2-naphthylacetic acid as inhibitors of AST IV-catalyzed sulfation of 1-naphthalenemethanol were approximately 10-fold higher than those of the corresponding naphthoic acids. A substituted 2-naphthylacetic acid derivative, naproxen, also inhibited the aryl sulfotransferase. Preliminary studies indicate that aryl carboxylic acids also inhibit sulfation of phenols catalyzed by AST IV. 2-Naphthoic acid inhibited the sulfation of 2-naphthol catalyzed by AST IV at pH 5.5 with a Ki of 260 microM. In addition to these results with the homogeneous sulfotransferase, inhibition of aryl sulfotransferase activity by carboxylic acids was also observed in rat hepatic 100,000 g supernatant fractions. Thus, aryl carboxylic acids represent a new class of inhibitors for AST IV.

Animals↗