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B Wermuth

Publications and source records attributed to B Wermuth.

At least 73 records · Page 4Linked to original sources
PubMed1988

Genotyping of human class I alcohol dehydrogenase. Analysis of enzymatically amplified DNA with allele-specific oligonucleotides.

Large inter-individual differences are noted in the susceptibility to alcohol-related problems. Part of this variation may be due to the different isoenzyme patterns of the alcohol-metabolizing enzymes and, consequently, different pharmacokinetics of alcohol degradation. We have used the polymerase chain reaction and oligonucleotide hybridization to amplify and analyze class I alcohol dehydrogenase isoenzyme-specific genomic DNA. The method unambiguously distinguishes between different allelic variants and thus provides a new means of elucidating the alcohol dehydrogenase isoenzyme pattern of humans.

Alcohol Dehydrogenase↗
PubMed1988

Degradation of aliphatic alcohols by human liver alcohol dehydrogenase: effect of ethanol and pharmacokinetic implications.

We have investigated the oxidation of low molecular weight aliphatic alcohols by Class I, II, and III alcohol dehydrogenases (ADH) isolated from human liver. These alcohols are generally present as byproducts of alcoholic beverages and referred to as alcoholic congeners. At concentrations corresponding to those in the blood after ingestion of alcoholic drinks (10-100 microM), the oxidation of propanol-1, isobutanol, 2-methylbutanol-1, and 3-methylbutanol-1 was mediated mainly by the isoenzymes of class I ADH, whereas butanol-1 was metabolized by Class I and II ADH. Class II ADH showed no activity with any of the alcohols at concentrations up to 100 microM. Lineweaver-Burk plots of the Class I ADH-catalyzed oxidation of all the congeners tested were linear in the pharmacokinetically relevant concentration range between 10 and 100 microM. Ethanol at concentrations found in the blood after moderate drinking (2.5-10 mM) caused a concentration-dependent inhibition of the congener oxidation. The experimentally determined kinetic constants were used to simulate the pharmacokinetics of propanol-1 metabolism in a multicompartment model system which accounts for first-pass elimination. The results suggest, in agreement with reported data from drinking experiments, that congener alcohols undergo considerable metabolism during the first liver passage, the extent of the first-pass metabolism depending on the ethanol dose.

1-Propanol↗
PubMed1987

Inactivation of carbonyl reductase from human brain by phenylglyoxal and 2,3-butanedione: a comparison with aldehyde reductase and aldose reductase.

Aldehyde reductase (alcohol:NADP+ oxidoreductase, EC 1.1.1.2), aldose reductase (alditol:NAD(P)+ 1-oxidoreductase, EC 1.1.1.21) and carbonyl reductase (secondary-alcohol:NADP+ oxidoreductase, EC 1.1.1.184) constitute the enzyme family of the aldo-keto reductases, a classification based on similar physicochemical properties and substrate specificities. The present study was undertaken in order to obtain information about the structural relationships between the three enzymes. Treatment of human aldehyde and carbonyl reductase with phenylglyoxal and 2,3-butanedione caused a complete and irreversible loss of enzyme activity, the rate of loss being proportional to the concentration of the dicarbonyl reagents. The inactivation of aldehyde reductase followed pseudo-first-order kinetics, whereas carbonyl reductase showed a more complex behavior, consistent with protein modification cooperativity. NADP+ partially prevented the loss of activity of both enzymes, and an even better protection of aldehyde reductase was afforded by the combination of coenzyme and substrate. Aldose reductase was partially inactivated by phenylglyoxal, but insensitive to 2,3-butanedione. The degree of inactivation with respect to the phenylglyoxal concentration showed saturation behavior. NADP+ partially protected the enzyme at low phenylglyoxal concentrations (0.5 mM), but showed no effect at high concentrations (5 mM). These findings suggest the presence of an essential arginine residue in the substrate-binding domain of aldehyde reductase and the coenzyme-binding site of carbonyl reductase. The effect of phenylglyoxal on aldose reductase may be explained by the modification of a reactive thiol or lysine rather than an arginine residue.

Alcohol Dehydrogenase↗
PubMed1987

Kinetics of carbonyl reductase from human brain.

Initial-rate analysis of the carbonyl reductase-catalysed reduction of menadione by NADPH gave families of straight lines in double-reciprocal plots consistent with a sequential mechanism being obeyed. The fluorescence of NADPH was increased up to 7-fold with a concomitant shift of the emission maximum towards lower wavelength in the presence of carbonyl reductase, and both NADPH and NADP+ caused quenching of the enzyme fluorescence, indicating formation of a binary enzyme-coenzyme complex. Deuterium isotope effects on the apparent V/Km values decreased with increasing concentrations of menadione but were independent of the NADPH concentration. The results, together with data from product inhibition studies, are consistent with carbonyl reductase obeying a compulsory-order mechanism, NADPH binding first and NADP+ leaving last. No significant differences in the kinetic properties of three molecular forms of carbonyl reductase were detectable.

Alcohol Oxidoreductases↗
PubMed1986

Carbonyl reductase provides the enzymatic basis of quinone detoxication in man.

Enzymes catalyzing the two-electron reduction of quinones to hydroquinones are thought to protect the cell against quinone-induced oxidative stress. Using menadione as a substrate, carbonyl reductase, a cytosolic, monomeric oxidoreductase of broad specificity for carbonyl compounds, was found to be the main NADPH-dependent quinone reductase in human liver, whereas DT-diaphorase, the principal two-electron transferring quinone reductase in rat liver, contributed a very minor part to the quinone reductase activity of human liver. Carbonyl reductase from liver was indistinguishable from carbonyl reductase previously isolated from brain (B. Wermuth, J. biol. Chem. 256, 1206 (1981] on the basis of molecular weight, isoelectric point, immunogenicity, substrate specificity and inhibitor sensitivity. The purified enzyme from liver catalyzed the reduction of a great variety of quinones. The best substrates were benzo- and naphthoquinones with short substituents, and the K-region orthoquinones of phenanthrene, benz(a)anthracene, pyrene and benzo(a)pyrene. A long hydrophobic side chain in the 3-position of the benzo- and naphthoquinones and the vicinity of a bay area or aliphatic substituent (pseudo bay area) to the oxo groups of the polycyclic compounds decreased or abolished the ability of the quinone to serve as a substrate. Non-k-region orthoquinones of polycyclic aromatic hydrocarbons were more slowly reduced than the corresponding K-region derivatives. The broad specificity of carbonyl reductase for quinones is in keeping with a role of the enzyme as a general quinone reductase in the catabolism of these compounds.

Alcohol Oxidoreductases↗
PubMed1985

Immunochemical characterization of aldo-keto reductases from human tissues.

Aldose reductase, aldehyde reductase and carbonyl reductase constitute a family of monomeric NADPH-dependent oxidoreductases with similar physical and chemical properties. Characterization of the enzymes from human tissues by immunotitration and an enzyme immunoassay indicated that, despite their apparent likeness, the three reductases do not cross-react immunochemically.

Alcohol Oxidoreductases↗
PubMed1983

Aldose and aldehyde reductase exhibit isocorticosteroid reductase activity.

In this paper we describe the reduction of corticosteroid metabolites containing the 17 beta-aldol side chain (isocorticosteroids) by aldose and aldehyde reductase from human tissues. Aldose reductase catalyzed the reduction of the aldehydes derived from cortisol and corticosterone at about the same rate, whereas aldehyde reductase preferentially acted on the aldehydes derived from 17-deoxycorticosteroids. At comparable rates of reduction the Michaelis constants for the best steroid aldehydes were one order of magnitude lower than for the hitherto best substrates. We propose that aldose and aldehyde reductase participate in the conversion of the corticosteroid ketol side chain to the glycol side chain via an aldol intermediate by the 'long loop' pathway proposed by Monder and Bradlow [(1977) J. Steroid Biochem. 8, 897-908].

Alcohol Oxidoreductases↗
PubMed1982

Purification and characterization of human-brain aldose reductase.

Aldose reductase (EC 1.1.1.21) from human brain has been purified to apparent homogeneity. The enzyme catalyzes the NADPH-dependent reduction of several physiological and xenobiotic aldehydes. Isocorticosteroids, e.g. isocortisol and isocorticosterone, are the best substrates (Km less than 1 micron), followed by aromatic and arylalkyladehydes, including biogenic aldehydes (Km = 3 - 15 microM). The activity towards aldoses is highest with glyceraldehyde (Km = 25 microM) and decreases with increasing number of carbon atoms of the sugar. Flavonoids, e.g. quercetin and rutin, inhibit aldose reductase (IC50 = 2 - 5 microM). Sulfate ions, on the other hand, stimulate the enzyme activity. Thiol-modifying reagents, e.g. 4-hydroxymercuribenzoate and iodoacetate, cause a time-dependent inactivation. Aldose reductase consists of a single polypeptide chain with a molecular weight of 38 000 and an isoelectric point of 5.9. In the presence of thiol reagents the isoelectric point is shifted to 5.1. Antibodies against aldose reductase do not cross-react with other carbonyl reductases, Nevertheless, the comparison of structural and enzymic properties of aldose reductase with those of other carbonyl reductases suggests a relationship between aldose reductase and aldehyde reductase (EC 1.1.1.2).

Adrenal Cortex Hormones↗
PubMed1981

Purification and properties of an NADPH-dependent carbonyl reductase from human brain. Relationship to prostaglandin 9-ketoreductase and xenobiotic ketone reductase.

A nonspecific NADPH-dependent carbonyl reductase from human brain (formerly designated as aldehyde reductase 1; Ris, M. M., and von Wartburg, J. P. (1973) Eur. J. Biochem. 37, 69-77) has been purified to homogeneity. The enzyme reduces a number of biologically and pharmacologically active carbonyl compounds. Quinones, e.g. menadione, ubiquinone, and tocopherolquinone are the best substrates, followed by aldehydes containing an activated carbonyl moiety, e.g. 4-nitrobenzaldehyde or methylglyoxal. The enzyme also reduces ketones, e.g. prostaglandins of the E and A class, the anthracycline antibiotic daunorubicin and 3-ketosteroids. During catalysis the pro 4S hydrogen atom of the nicotinamide ring of NADPH is transferred to the substrate. Flavonoids, e.g. quercetin and rutin, indomethacin, ethacrynic acid, and dicoumarol inhibit the enzyme activity. 4-Hydroxymercuribenzoate and iodoacetate inactivate the enzyme. NADPH and substrate do not protect against the loss of activity. Carbonyl reductase consists of a single polypeptide chain with a molecular weight of 30,000. The native enzyme occurs in three molecular forms with similar substrate specificity and inhibitor sensitivity. The isoelectric points of the three enzyme species are 6.95, 7.85, and 8.5. In the presence of coenzyme the isoelectric points are shifted to 5.2 to 5.9. The comparison of structural and enzymic features of carbonyl reductase with other monomeric oxidoreductases suggests a close relationship of carbonyl reductase with prostaglandin 9-keto-reductase and xenobiotic ketone reductase.

Alcohol Oxidoreductases↗
PubMed1980

Characterization of a mitochondrial NADH-dependent nitro reductase from rat brain.

Rat brain mitochondria contain an NADH-linked nitro reductase that converts various aromatic nitro compounds, including the anti-schistosomal agent niridazole, into the hydroxylamine metabolites. The enzyme is tightly bound to the inner membrane and its activity is measurable only after disrupting the mitochondria. Triton X-100 (1%) and sonication partially solubilize the enzyme. The molecular weight determined by gel filtration is approx. 200 000. The temperature optima for the membrane-bound and for the solubilized enzyme are at 35 and 30 degrees C, respectively. The pH optimum for the membrane-bound enzyme is 9.2. NAD+ and 4-hydroxymercuribenzoate decrease the enzyme activity. Oxygen, carbon monoxide, cyanide, rotenone, barbiturates, chlorpromazine, dicumarol and chelating agents have no effect on the activity. The subcellular localization, substrate specificity and sensitivity to inhibitors distinguish the mitochondrial nitro reductase from the corresponding microsomal and cytosolic enzymes.

Animals↗
PubMed1980

Kinetic studies on NADPH-linked aldehyde reductase from human liver.

The mechanism of D-glucuronate reduction by human liver NADPH-dependent aldehyde reductase was investigated. At pH 7.4 the Km values for NADPH, NADP+, D-glucuronate and L-gulonate were 2.2 microM, 6 microM, 3.2 mM and 6 mM, respectively. Product inhibition studies in the forward direction (reduction of glucuronate) gave a competitive pattern for the inhibition of NADPH oxidation by NADP+ and non-competitive patterns for the other three inhibitions. In the backward direction all patterns appeared to be competitive. Deuterium isotope effects were dependent on the concentration of D-glucuronate and decreased to unity at infinite concentrations of D-glucuronate. Our findings suggest for aldehyde reductase a kinetic mechanism with sequential ordered binding of NADPH and D-glucuronate and random dissociation of NADP+ and L-gulonate.

Aldehyde Oxidoreductases↗