Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.1.1.1 (alcohol dehydrogenase)
9,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Kinetic substituent effects have been used to examine the catalytic reaction profile of xylose reductase from the yeast Candida tenuis, a representative aldo/keto reductase of primary carbohydrate metabolism. Michaelis-Menten parameters (k(cat) and K(m)) for NADH-dependent enzymic aldehyde reductions have been determined using a homologous series of benzaldehyde derivatives in which substituents in meta and para positions were employed to systematically perturb the properties of the reactive carbonyl group. Kinetic isotope effects (KIEs) on k(cat) and k(cat)/K(m) for enzymic reactions with meta-substituted benzaldehydes have been obtained by using NADH (2)H-labelled in the pro-R C4-H position, and equilibrium constants for the conversion of these aldehydes into the corresponding alcohols (K(eq)) have been measured in the presence of NAD(H) and enzyme. Aldehyde dissociation constants (K(d)) and the hydride transfer rate constant (k(7)) have been calculated from steady-state rate and KIE data. Quantitative structure-activity relationship analysis was used to factor the observed substituent dependence of k(cat)/K(m) into a major electronic effect and a productive positional effect of the para substituent. k(cat)/K(m) (after correction for substituent position) and K(eq) obeyed log-linear correlations over the substituent parameter, Hammett sigma, giving identical slope values (rho) of +1.4 to +1.7, whereas the same Hammett plot for logK(d) yielded rho=-1.5. This leads to the conclusion that electron-withdrawing substituents facilitate the reaction and increase binding to about the same extent. KIE values for k(cat) (1.8) and k(cat)/K(m) (2.7), and likewise k(7), showed no substituent dependence. Therefore, irrespective of the observed changes in reactivity over the substrate series studied no shift in the character of the rate-limiting transition state of hydride transfer occurred. The signs and magnitudes of rho values suggest this transition state to be product-like in terms of charge development at the reactive carbon. Structure-reactivity correlations reveal active-site homologies among NADPH-specific and dual NADPH/NADH-specific yeast xylose reductases and across two aldo/keto reductase families in spite of the phylogenetic separation of the host organisms producing xylose reductase (family 2B) and aldehyde reductase (family 1A).
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PMID:Catalytic reaction profile for NADH-dependent reduction of aromatic aldehydes by xylose reductase from Candida tenuis. 1200 38

The relationship between substrate mobility and catalysis was studied with wild-type and Phe93Ala (F93A) horse liver alcohol dehydrogenase (ADH). Wild-type ADH binds 2,3,4,5,6-pentafluorobenzyl alcohol in one position as shown by X-ray results, and (19)F NMR shows five resonances for the fluorines of the bound alcohol. The two meta-fluorines exchange positions with a rate constant of about 4 s(-1), indicating that mobility (ring flipping) of the benzyl alcohol is relatively restricted. The wild-type enzyme binds 2,3-difluorobenzyl alcohol in two alternative conformations that are related by a ring flip and a small translation of the fluorinated benzene ring, and the (19)F NMR spectrum shows three resonances for the two bound fluorines, consistent with the two orientations. Phe-93 interacts with the bound benzyl alcohols, and the F93A substitution decreases the rate constants for hydride transfer for benzyl alcohol oxidation and benzaldehyde reduction by 7.4- and 130-fold, respectively. The structure of F93A ADH crystallized with NAD(+) and 2,3,4,5,6-pentafluorobenzyl alcohol is similar to the structure of the wild-type enzyme complex except that the pentafluorobenzyl alcohol is not found in one position. The (19)F NMR spectrum of the F93A ADH-NAD(+)-pentafluorobenzyl alcohol complex shows three resonances for the bound fluorines. Line shape analysis of the spectrum suggests the bound pentafluorobenzyl ring undergoes rapid ring-flipping at about 20 000 s(-1). The F93A substitution greatly increases the mobility of the benzyl alcohol but modestly and differentially decreases the probability that the substrate is preorganized for hydride transfer.
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PMID:Mobility of fluorobenzyl alcohols bound to liver alcohol dehydrogenases as determined by NMR and X-ray crystallographic studies. 1250 Dec 6

A transcriptionally active region has been identified in the 5' flanking region of the alcohol dehydrogenase gene of the crenarchaeon Sulfolobus solfataricus through the evaluation of the activity of putative transcriptional regulators and the role of the region upstream of the gene under specific metabolic circumstances. Electrophoretic mobility shift assays with crude extracts revealed protein complexes that most likely contain TATA box-associated factors. When the TATA element was deleted from the region, binding sites for both DNA binding proteins, such as the small chromatin structure-modeling Sso7d and Sso10b (Alba), and transcription factors, such as the repressor Lrs14, were revealed. To understand the molecular mechanisms underlying the substrate-induced expression of the adh gene, the promoter was analyzed for the presence of cis-acting elements recognized by specific transcription factors upon exposure of the cell to benzaldehyde. Progressive dissection of the identified promoter region restricted the analysis to a minimal responsive element (PAL) located immediately upstream of the transcription factor B-responsive element-TATA element, resembling typical bacterial regulatory sequences. A benzaldehyde-activated transcription factor (Bald) that specifically binds to the PAL cis-acting element was also identified. This protein was purified from heparin-fractionated extracts of benzaldehyde-induced cells and was shown to have a molecular mass of approximately 16 kDa. The correlation between S. solfataricus adh gene activation and benzaldehyde-inducible occupation of a specific DNA sequence in its promoter suggests that a molecular signaling mechanism is responsible for the switch of the aromatic aldehyde metabolism as a response to environmental changes.
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PMID:Transcriptional regulation of the gene encoding an alcohol dehydrogenase in the archaeon Sulfolobus solfataricus involves multiple factors and control elements. 1281 87

We report here the ability of an alcohol dehydrogenase (ADH) ribozyme to reduce a benzaldehyde. While the ribozyme was initially evolved in vitro based on the activity for the NAD+-dependent oxidation of the benzyl alcohol, we found that this ADH ribozyme is also capable of reducing the aldehyde in the presence of NADH and Zn2+. The rate acceleration gained by ribozyme catalysis was more than 6 orders of magnitude larger than the spontaneous reaction. Although the reversibility of phosphordiester and acyl transfer reactions catalyzed by ribozymes was known, that of other chemical reactions has not been well established. This study has demonstrated the reversibility of a hydride transfer chemistry catalyzed by the ADH ribozyme. Most interestingly, the ribozyme shares many features with the protein ADHs, e.g., reversibility and NADH/Zn2+ dependence.
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PMID:Reduction of an aldehyde by a NADH/Zn2+ -dependent redox active ribozyme. 1509 68

Cinnamyl alcohol dehydrogenases (CAD; 1.1.1.195) catalyse the reversible conversion of p-hydroxycinnamaldehydes to their corresponding alcohols, leading to the biosynthesis of lignin in plants. Outside of plants their role is less defined. The gene for cinnamyl alcohol dehydrogenase from Helicobacter pylori (HpCAD) was cloned in Escherichia coli and the recombinant enzyme characterized for substrate specificity. The enzyme is a monomer of 42.5 kDa found predominantly in the cytosol of the bacterium. It is specific for NADP(H) as cofactor and has a broad substrate specificity for alcohol and aldehyde substrates. Its substrate specificity is similar to the well-characterized plant enzymes. High substrate inhibition was observed and a mechanism of competitive inhibition proposed. The enzyme was found to be capable of catalysing the dismutation of benzaldehyde to benzyl alcohol and benzoic acid. This dismutation reaction has not been shown previously for this class of alcohol dehydrogenase and provides the bacterium with a means of reducing aldehyde concentration within the cell.
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PMID:Characterization of cinnamyl alcohol dehydrogenase of Helicobacter pylori. An aldehyde dismutating enzyme. 1572 Mar 99

An aldehyde dehydrogenase gene, designated phnN, was isolated from a genome library of the 1,4-dimethylnaphthalene-utilizing soil bacterium, Sphingomonas sp. 14DN61. Escherichia coli expressing the phnN gene converted 1,4-dihydroxymethylnaphthalene to 1-hydroxymethyl-4-naphthoic acid. The putative amino acid sequence of the phnN gene product had 31-42% identity with those of NAD(+)-dependent short-chain aliphatic aldehyde dehydrogenases and a secondary alcohol dehydrogenase. The NAD(P)(+)-binding site and two consensus sequences involved in the active site for aldehyde dehydrogenase are conserved among these proteins. The PhnN enzyme purified from recombinant E. coli showed broad substrate specificity towards various aromatic aldehydes, i.e., 1- and 2-naphaldehydes, cinnamaldehyde, vanillin, syringaldehyde, benzaldehyde and benzaldehydes substituted with a hydroxyl, methyl, methoxy, chloro, fluoro, or nitro group were converted to their corresponding carboxylic acids. Interestingly, E. coli expressing phnN was able to biotransform a variety of not only aromatic aldehydes, but also aromatic alcohols to carboxylic acids.
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PMID:Characterization of Sphingomonas aldehyde dehydrogenase catalyzing the conversion of various aromatic aldehydes to their carboxylic acids. 1581 42

We describe the three-dimensional structure of sinapyl alcohol dehydrogenase (SAD) from Populus tremuloides (aspen), a member of the NADP(H)-dependent dehydrogenase family that catalyzes the last reductive step in the formation of monolignols. The active site topology revealed by the crystal structure substantiates kinetic results indicating that SAD maintains highest specificity for the substrate sinapaldehyde. We also report substantial substrate inhibition kinetics for the SAD-catalyzed reduction of hydroxycinnamaldehydes. Although SAD and classical cinnamyl alcohol dehydrogenases (CADs) catalyze the same reaction and share some sequence identity, the active site topology of SAD is strikingly different from that predicted for classical CADs. Kinetic analyses of wild-type SAD and several active site mutants demonstrate the complexity of defining determinants of substrate specificity in these enzymes. These results, along with a phylogenetic analysis, support the inclusion of SAD in a plant alcohol dehydrogenase subfamily that includes cinnamaldehyde and benzaldehyde dehydrogenases. We used the SAD three-dimensional structure to model several of these SAD-like enzymes, and although their active site topologies largely mirror that of SAD, we describe a correlation between substrate specificity and amino acid substitution patterns in their active sites. The SAD structure thus provides a framework for understanding substrate specificity in this family of enzymes and for engineering new enzyme specificities.
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PMID:Structural and kinetic basis for substrate selectivity in Populus tremuloides sinapyl alcohol dehydrogenase. 1582 7

Although higher initial rates of phenylacetyl carbinol formation were observed in fermentations containing a high starting benzaldehyde level, a massive reduction in yeast viability was observed resulting in early cessation of production formation. Pulse feeding to maintain lower benzaldehyde concentrations resulted in a lower initial reaction rate, but prolonged yeast viability and the biotransformation. This resulted in higher overall product tilers. As benzaldehyde concentration was increased, yeast growth rate was reduced (0.5 g/L), inhibited (1-2 g/L), or cell viability reduced (3 g/L). Benzaldehyde appeared to alter the cell permeability barrier to substrates and products. Reductions in yeast biomass levels and especially protein and lipid content were observed during the biotransformation. The effects of benzaldehyde and reaction products on yeast pyruvate decarboxylase and alcohol dehydrogenase stability were determined. Homogenized yeast cells produced similar phenylacetyl carbinol levels to whole yeast only if supplemented with thiamine pyrophosphate and magnesium.
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PMID:Biotransformation of benzaldehyde by Saccharomyces cerevisiae: characterization of the fermentation and toxicity effects of substrates and products. 1858 85

Whole cells of Saccharomyces cerevisiae analyzed the conversion of benzaldehyde to benzyl alcohol in aqueous-organic biphasic media. Reaction rate increased dramatically as moisture content of the solvent was increased in the range 0% to 2%. The highest biotransformation rates were observed when hexane was used as organic solvent. Benzaldehyde was also converted to benzyl alcohol by a cell-free crude extract in biphasic systems containing hexane, although the rate of product formation was much lower. Mutant strains of S. cerevisiae lacking some or all of the ADH isoenzymes, ADH I, II, and III, manifested similar rates for bioconversion of benzaldehyde to benzyl alcohol in both aqueous and two-phase systems. In general, conversion rates observed in aqueous media were 2 to 3 times higher than those observed in hexane containing 2% moisture.
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PMID:Reductive biotransformation by wild type and mutant strains of Saccharomyces cerevisiae in aqueous-organic solvent biphasic systems. 1860 Oct 21

The capacities of yeast wild-type and mutants strains known to lack specific ADH isoenzymes to produce L-phenylacetyl carbinol (PAC) and benzyl alcohol in biotransformation trials were also investigated. Pyruvate decarboxylase activity, responsible for PAC formation and ADH activity, which can participate in reduction of benzaldehyde to benzyl alcohol, was also determined in each strain. In addition, the capacity of each strain to produce ethanol was investigated. Mutant strains lacking all of the isoenzymes, ADH-I, ADH-II, and ADH-III, still exhibited some ADH activity and were capable of production of benzyl alcohol and ethanol.
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PMID:Production of L-phenylacetyl carbinol by biotransformation: product and by-product formation and activities of the key enzymes in wild-type and ADH isoenzyme mutants of Saccharomyces cerevisiae. 1860 7


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