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

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Abstract
Pyrazole is a strong inhibitor of liver alcohol dehydrogenase in combination with oxidized coenzyme NAD. Three different complexes of the inhibitor with the enzyme were studied by using crystallographic methods: the binary complex with pyrazole to 3.2-.ANG. resolution, the ternary complex with NAD-pyrazole to 2.9-.ANG. resolution and the ternary complex with NAD-4-iodopyrazole to 2.9-.ANG. resolution. Crystals of the binary complex are isomorphous to the apoenzyme, and pyrazole binds to the active-site Zn atom in a way analogous to imidazole. Crystals of the 2 ternary complexes are isomorphous with the ternary alcohol dehydrogenase-NADH-dimethyl sulfoxide complex. One of the N atoms of the pyrazole ring is directly bound to the active-site Zn atom with a Zn.sbd.N bond distance of 2.1 .ANG.. The other N atom is 2 .ANG. from the C4 atom of the nicotinamide ring of the coenzyme. The I 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.