Biochemical Pharmacology of Pyrazoles

Deis, Frank H.; Lester, David

doi:10.1007/978-1-4684-3450-7_18

Cited by 9 publications

(4 citation statements)

References 133 publications

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“…Pyrazole is a potent inhibitor of LADH. The structure of the LADH-N AD+-pyrazole complex is of interest not only because of possible implications for the catalytic mechanism of LADH but also because of its potential medical importance in the treatment of alcoholism, as well as methanol and ethylene glycol poisoning (Deis & Lester, 1979). In the presence of NAD"1", pyrazole forms a tight ternary complex with LADH which has a Kx of 0.2 µ (Theorell et al, 1969) and a lifetime of 25 s at pH 7 (Shore & Gilleland, 1970).…”

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Structure of the liver alcohol dehydrogenase-NAD+-pyrazole complex as determined by nitrogen-15 NMR spectroscopy

Becker¹,

Roberts²

1984

Biochemistry

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The structures of the liver alcohol dehydrogenase (LADH)-NAD+-pyrazole and LADH-NAD+-4-ethylpyrazole complexes were investigated by 15N nuclear magnetic resonance (NMR) spectroscopy. 15N chemical shifts were obtained for 15N-labeled inhibitors and 15N-labeled coenzyme bound in the ternary enzyme complexes. The structures of the two inhibitor complexes appear to be very similar. 15N NMR studies of model pyrazole-zinc chloride complexes were carried out to determine the effect of zinc complexation on pyrazole chemical shifts. The N1 nicotinamide chemical shift of the coenzyme of the LADH-NAD+-pyrazole complex demonstrates that the NAD+ is converted to a dihydronicotinamide derivative in the complex. The N1 chemical shift of the pyrazole in the ternary complex is consistent with covalent bond formation between pyrazole N1 and the nicotinamide ring of the coenzyme. The N2 chemical shift of the pyrazole in the ternary complex indicates that the nucleus of this nitrogen is about 40 ppm more shielded than those of the N2 nitrogens of typical pyrazoles. Such shielding is expected as the result of direct complexation of N2 to the active-site zinc. Shift comparisons with zinc-pyrazole complexes indicate a high degree of inner-sphere coordination of the pyrazole N2 to the active-site zinc in the ternary complex.

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Structure of the liver alcohol dehydrogenase-NAD+-pyrazole complex as determined by nitrogen-15 NMR spectroscopy

Becker¹,

Roberts²

1984

Biochemistry

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“…One of the nitrogen atoms of the pyrazole ring is directly bound to the I^razole and its derivatives have been extensively studied as inhibitors for liver alcohol dehydrogenase. They have been used in steady-state and fast kinetic experiments (Yonetani, 1963;Brand et al, 1967;Shore & Gilleland, 1970; Theorell & Tatemoto, 1971;McFarland & Bernhard, 1972; Jacobs et al, 1974;Reynolds & McKinley-McKee, 1975; Luisi et al, 1975;McFarland et al, 1977;De Traglia et al, 1977;Andersson, P., et al, 1981), in purification of the enzyme (Andersson et al, 1974;Lange & Vallee, 1976), and in experiments with enzyme where the active-site metal has been exchanged (Maret et al, 1980;Makinen & Yim, 1981), as well as for NMR (Bobsein & Meyers, 1981;Andersson, I., et al, 1981) and metabolic studies (Lester et al, 1968;Blomstrand & Theorell, 1970;Rydberg et al, 1972;Deis & Lester, 1979; Lieber, 1977). Some derivatives of pyrazole are the strongest inhibitors so far found for the enzyme (Tolf, 1981).…”

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Pyrazole binding in crystalline binary and ternary complexes with liver alcohol dehydrogenase

Eklund¹,

Samama²,

Wallen³

1982

Biochemistry

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Pyrazole is a strong inhibitor of liver alcohol dehydrogenase in combination with oxidized coenzyme NAD+. We have studied three different complexes of the inhibitor with the enzyme by using crystallographic methods: (1) the binary complex with pyrazole to 3.2-A resolution, (2) the ternary ternary complex with NAD+-4-iodopyrazole to 2.9-A resolution. Crystals of the binary complex are isomorphous to the apoenzyme, and pyrazole binds to the active-site zinc atom in a way analogous to imidazole. Crystals of the two ternary complexes are isomorphous with the ternary alcohol dehydrogenase-NADH-dimethyl sulfoxide complex. One of the nitrogen atoms of the pyrazole ring is directly bound to the active-site zinc atom with a Zn-N bond distance of 2.1A. The other nitrogen atom is 2 A from the C4 atom of the nicotinamide ring of the coenzyme. The iodine atom in 4-iodopyrazole is located in the hydrophobic substrate cleft. The effect of substitutions on the pyrazole ring are discussed in relation to the structure of the active site and substrate pocket. Pyrazole derivatives with long alkyl chains bound in the 4 position are outstanding inhibitors, and this property is related to the topography of the hydrophobic substrate cleft. The conformation of the oxidized coenzyme in the ternary complexes is essentially the same as that of the reduced coenzyme NADH in the NADH-dimethyl sulfoxide complex.

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“…A dose of 1 mmole/kg inhibits elimination of a dose of ethanol of 20 mmole/kg by 87%. 4-Methylpyrazole has low toxicity and more specificity than pyrazole [29]. It does not appreciably inhibit rat liver catalase or cytochrome P-450, as the latter has an in vitro K i of 5.7 mM [30, 31].…”

Section: Methodsmentioning

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Contribution of liver alcohol dehydrogenase to metabolism of alcohols in rats

Plapp

Leidal

Murch

et al. 2015

Chemico-Biological Interactions

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The kinetics of oxidation of various alcohols by purified rat liver alcohol dehydrogenase (ADH) were compared with the kinetics of elimination of the alcohols in rats in order to investigate the roles of ADH and other factors that contribute to the rates of metabolism of alcohols. Primary alcohols (ethanol, 1-propanol, 1-butanol, 2-methyl-1-propanol, 3-methyl-1-butanol) and diols (1,3-propanediol, 1,3-butanediol, 1,4-butanediol, 1,5-pentanediol) were eliminated in rats with zero-order kinetics at doses of 5–20 mmole/kg. Ethanol was eliminated most rapidly, at 7.9 mmole/kg•h. Secondary alcohols (2-propanol-d7, 2-propanol, 2-butanol, 3-pentanol, cyclopentanol, cyclohexanol) were eliminated with first order kinetics at doses of 5–10 mmole/kg, and the corresponding ketones were formed and slowly eliminated with zero or first order kinetics. The rates of elimination of various alcohols were inhibited on average 73% (55% for 2-propanol to 90% for ethanol) by 1 mmole/kg of 4-methylpyrazole, a good inhibitor of ADH, indicating a major role for ADH in the metabolism of the alcohols. The Michaelis kinetic constants from in vitro studies (pH 7.3, 37 °C) with isolated rat liver enzyme were used to calculate the expected relative rates of metabolism in rats. The rates of elimination generally increased with increased activity of ADH, but a maximum rate of 6 ± 1 mmole/kg•h was observed for the best substrates, suggesting that ADH activity is not solely rate-limiting. Because secondary alcohols only require one NAD+ for the conversion to ketones whereas primary alcohols require two equivalents of NAD+ for oxidation to the carboxylic acids, it appears that the rate of oxidation of NADH to NAD+ is not a major limiting factor for metabolism of these alcohols, but the rate-limiting factors are yet to be identified.

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Biochemical Pharmacology of Pyrazoles

Cited by 9 publications

References 133 publications

Structure of the liver alcohol dehydrogenase-NAD+-pyrazole complex as determined by nitrogen-15 NMR spectroscopy

Structure of the liver alcohol dehydrogenase-NAD+-pyrazole complex as determined by nitrogen-15 NMR spectroscopy

Pyrazole binding in crystalline binary and ternary complexes with liver alcohol dehydrogenase

Contribution of liver alcohol dehydrogenase to metabolism of alcohols in rats

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