Liver alcohol dehydrogenase catalytic activity

Liver alcohol dehydrogenase catalytic activity, 1017 Lossen rearrangement, 813 Lungs... 

Fig. 6 Illustration from Chin and Klinman. Increased catalytic activity of horse-liver alcohol dehydrogenase in the oxidation of benzyl alcohol to benzaldehyde by NAD, measured by cat/ M (ordinate), correlates with the Swain-Schaad exponent for the -secondary isotope effect (abscissa), for which values above about four are indicators of tunneling. This is a direct test of the hypothesis that tunneling in the action of this enzyme contributes to catalysis. As the rate increases by over two orders of magnitude and then levels off, the anomalous Swain-Schaad exponents also increase and then level off. Reproduced from Ref. 28 with the permission of the American Chemical Society.

Alcohol dehydrogenases (ADH EC 1.1.1.1), for which several X-ray structures are available ", catalyze the biological oxidation of primary and secondary alcohols via the formal transfer of a hydride anion to the oxidized form of nicotinamide adenine dinucleotide (NAD ), coupled with the release of a proton. Liver alcohol dehydrogenase (LADH) consists of two similar subunits, each of which contains two zinc sites, but only one site within each subunit is catalytically active. The catalytic zinc is coordinated in a distorted tetrahedral manner to a histidine residue, two cysteine residues and a water molecule. The remaining zinc is coordinated tetrahedrally to four cysteine residues and plays only a structural role . 

The catalytic activity of liver alcohol dehydrogenase is strongly pH dependent over a wide range. It has been well established that this pH dependence derives from the combined effects of pH on several steps in the catalytic mechanism. They are all proton equilibria involving... 

Scheme 10 Proton equilibria suggested to affect the catalytic activity of liver alcohol dehydrogenase by Kvassman and...

Enzymes as different as yeast alcohol oxidase, mushroom polyphenol oxidase, and horse-liver alcohol dehydrogenase demonstrated vastly increased enzymatic activity in several different solvents upon an increase in the water content, which always remained below the solubility limit (Zaks, 1988b). While much less water was required for maximal activity in hydrophobic than in hydrophilic solvents, relative saturation seems to be most relevant to determining the level of catalytic activity. Correspondingly, miscibility of a solvent with water is not a decisive criterion upon transition from a monophasic to a biphasic solvent system, no significant change in activity level was observed (Narayan, 1993). Therefore, the level of water essential for activity depends more on the solvent than on the enzyme. 

Kimura, E., Shionoya, M., Hoshino, A., Ikeda, T., Yamada, Y., A model for catalytically active zinc(II) ion in liver alcohol-dehydrogenase - a novel hydride transfer-reaction catalyzed by zinc (II) -macrocyclic polyamine complexes. J. Am. Chem. Soc. 1992,114, 10134-10137. 

Fig. 26. (A) Schematic diagram of one subunit of horse liver alcohol dehydrogenase. Znl is the active-site zinc. Designed by B. Furugren, from the work of Branden and colleagues [55], (B) Schematic diagram of a section through the horse liver alcohol dehydrogenase dimer. The catalytic zinc atoms are shown, with the inhibitory substrate analogue DMSO and coenzyme molecules indicated. The dimer has two active sites, each composed of parts of both subunits. From the work of Branden and colleagues [123].

Hard electrophiles like Mg(C104)2 are used to activate abiotic systems. In the enzyme liver alcohol dehydrogenase (LAD) a considerably different catalytic apparatus is present a zinc ion coordinated to two cysteines and a histidine serves as a coordinating site for the carbonyl compound/alcoholate, as illustrated in equation (10). This zinc ion has amphoteric properties consistent with the capacity to activate the reaction in both directions without being consumed, in other words to act as a catalyst. Synthetic models of this catalytically active zinc have been shown to possess some catalytic activity in analogy to the enzyme (see Section L3.3.5.1iii). 

Certain transition metal complexes exhibit activating properties and act with turnover on the metal center analogously to the catalytically active zinc ion in the active center of liver alcohol dehydrogenase. Various chiral europium shift reagents, for example Eu(hfc)3, induce reduction of (9b) by 1,4-dihydroni-cotinamides. Turnovers of about 100 are obtained on the metal complexes and methyl mandelate is formed with enantiomeric excesses of 25-44%. ... 

Fig. 11. Schematic representation of the interactions between the substrate, coenzyme, and the active site residues in horse liver alcohol dehydrogenase. Not shown are the interactions between Arg-47 and the pyrophosphate backbone, and Asp-49, which forms a salt bridge with His-57, another ligand of the zinc atom. Because of the close proximity to residues having obvious catalytically important functions, alterations in the interactions between the coenzyme and Ser-48 and His-51 that are anticipated from the binding of acyclo-NAD could readily cause the observed changes in substrate specificity. Based on Ref. 38.

Metalloenzymes are enzymes that have a tightly bound metal ion. These metal ions are normally incorporated into the enzymes during enzyme synthesis, and removal of the metal ions often results in the complete denat-uration of the enzyme. These metal ions may contribute either to the structure or the catalytic mechanism of a metalloenzyme. Lor example, horse liver alcohol dehydrogenase contains two tightly bound zinc ions (Zn ). The first zinc ion is structural it is bound to four cysteine side chains and is essential to maintain the structural integrity of the enzyme. The second zinc ion is catalytic it is bonnd to the side chains belonging to two cysteines and one histidine at the active site of the enzyme, and it participates in the catalytic cycle of the enzyme. 

The data cited thus far indicate that both alkaline phosphatase and liver alcohol dehydrogenase contain heterogeneous populations of metal atoms of the same species. In both instances, only two of the zinc atoms native to the enzyme appear to be involved in enzymatic activity. The remaining metal atoms do not have a catalytic role but appear to influence the quaternary structure of the protein, although the details of the manner in which this is accomplished are as yet uncertain. These observations have induced us to reexamine the possible effects of metals on structure in other proteins, including those having only single chains. 

Yeast and mammalian alcohol dehydrogenases differ in substrate specificity and catalytic activity. The yeast enzyme is more specific for acetaldehyde and ethanol, but mammalian enzymes have a broad substrate specificity, and even with primary alcohols maximum activity is not observed with ethanol. Because of the large amount of alcohol dehydrogenase present in human liver and its role in alcohol metabolism in man, human liver alcohol dehydrogenase is of particular interest. It was first purified by Wartburg et and crystallized by Mourad and Woronick. Human liver alcohol dehydrogenase is a dimer with subunit structures analogous to those of horse liver, and each subunit probably contains two zinc atoms. Several different types of human ADH have been isolated. with minor variations in amino acid... 

Investigations of Catalytic Mechanism of Active-Site Co II)-Substituted Liver Alcohol Dehydrogenase (NADH)... 

Zinc ion is essential for the catalytic activities of both yeast and liver alcohol dehydrogenase. Until recently, model systems have been notably unsuccessful in accounting for the participation of Zn(II) in the enzyme-catalyzed oxidoreductive interconversion of aldehyde and alcohol. The studies of Creighton and Sigman (20) and of Shinkai and Bruice (21, 22) conclusively demonstrate that Lewis (general) acid catalysis by Zn + (and other divalent metal ions) can effectively promote aldehyde reduction by the reduced 1,4-dihydropyridine moiety. 

This was the first example of a reduction of an aldehyde by an NADH analogue in a nonenzymatic system. If 1-propyl-4,4-dideuterionicotinamide is used, monodeuterated l,10-phenanthroline-2-carbinol is produced. This result demonstrates that the product is formed by direct hydrogen transfer from the reduced coenzyme analogue. It strengthens the view that coordination or proximity of the carbonyl to the zinc ion is probably important in the enzymatic catalysis. Zn(II) ion is known to be essential for the catalytic activity of horse liver alcohol dehydrogenase (280). 

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