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Sample records for aldehyde dehydrogenase inhibitor

  1. Daidzin: a potent, selective inhibitor of human mitochondrial aldehyde dehydrogenase.

    PubMed

    Keung, W M; Vallee, B L

    1993-02-15

    Human mitochondrial aldehyde dehydrogenase (ALDH-I) is potently, reversibly, and selectively inhibited by an isoflavone isolated from Radix puerariae and identified as daidzin, the 7-glucoside of 4',7-dihydroxyisoflavone. Kinetic analysis with formaldehyde as substrate reveals that daidzin inhibits ALDH-I competitively with respect to formaldehyde with a Ki of 40 nM, and uncompetitively with respect to the coenzyme NAD+. The human cytosolic aldehyde dehydrogenase isozyme (ALDH-II) is nearly 3 orders of magnitude less sensitive to daidzin inhibition. Daidzin does not inhibit human class I, II, or III alcohol dehydrogenases, nor does it have any significant effect on biological systems that are known to be affected by other isoflavones. Among more than 40 structurally related compounds surveyed, 12 inhibit ALDH-I, but only prunetin and 5-hydroxydaidzin (genistin) combine high selectivity and potency, although they are 7- to 15-fold less potent than daidzin. Structure-function relationships have established a basis for the design and synthesis of additional ALDH inhibitors that could both be yet more potent and specific.

  2. Discovery of a novel class of covalent inhibitor for aldehyde dehydrogenases

    SciTech Connect

    Khanna, Mary; Chen, Che-Hong; Kimble-Hill, Ann; Parajuli, Bibek; Perez-Miller, Samantha; Baskaran, Sulochanadevi; Kim, Jeewon; Dria, Karl; Vasiliou, Vasilis; Mochly-Rosen, Daria; Hurley, Thomas D.

    2012-10-23

    Human aldehyde dehydrogenases (ALDHs) comprise a family of 17 homologous enzymes that metabolize different biogenic and exogenic aldehydes. To date, there are relatively few general ALDH inhibitors that can be used to probe the contribution of this class of enzymes to particular metabolic pathways. Here, we report the discovery of a general class of ALDH inhibitors with a common mechanism of action. The combined data from kinetic studies, mass spectrometric measurements, and crystallographic analyses demonstrate that these inhibitors undergo an enzyme-mediated {beta}-elimination reaction generating a vinyl ketone intermediate that covalently modifies the active site cysteine residue present in these enzymes. The studies described here can provide the basis for rational approach to design ALDH isoenzyme-specific inhibitors as research tools and perhaps as drugs, to address diseases such as cancer where increased ALDH activity is associated with a cellular phenotype.

  3. Characterization of Two Distinct Structural Classes of Selective Aldehyde Dehydrogenase 1A1 Inhibitors

    PubMed Central

    Morgan, Cynthia A.; Hurley, Thomas D.

    2015-01-01

    Aldehyde dehydrogenases (ALDH) catalyze the irreversible oxidation of aldehydes to their corresponding carboxylic acid. Alterations in ALDH1A1 activity are associated with such diverse diseases as cancer, Parkinson’s disease, obesity, and cataracts. Inhibitors of ALDH1A1 could aid in illuminating the role of this enzyme in disease processes. However, there are no commercially available selective inhibitors for ALDH1A1. Here we characterize two distinct chemical classes of inhibitors that are selective for human ALDH1A1 compared to eight other ALDH isoenzymes. The prototypical members of each structural class, CM026 and CM037, exhibit sub-micromolar inhibition constants, but have different mechanisms of inhibition. The crystal structures of these compounds bound to ALDH1A1 demonstrate that they bind within the aldehyde binding pocket of ALDH1A1 and exploit the presence of a unique Glycine residue to achieve their selectivity. These two novel and selective ALDH1A1 inhibitors may serve as chemical tools to better understand the contributions of ALDH1A1 to normal biology and to disease states. PMID:25634381

  4. Characterization of Two Distinct Structural Classes of Selective Aldehyde Dehydrogenase 1A1 Inhibitors

    SciTech Connect

    Morgan, Cynthia A.; Hurley, Thomas D.

    2015-01-29

    Aldehyde dehydrogenases (ALDH) catalyze the irreversible oxidation of aldehydes to their corresponding carboxylic acid. Alterations in ALDH1A1 activity are associated with such diverse diseases as cancer, Parkinson’s disease, obesity, and cataracts. Inhibitors of ALDH1A1 could aid in illuminating the role of this enzyme in disease processes. However, there are no commercially available selective inhibitors for ALDH1A1. Here we characterize two distinct chemical classes of inhibitors that are selective for human ALDH1A1 compared to eight other ALDH isoenzymes. The prototypical members of each structural class, CM026 and CM037, exhibit sub-micromolar inhibition constants, but have different mechanisms of inhibition. The crystal structures of these compounds bound to ALDH1A1 demonstrate that they bind within the aldehyde binding pocket of ALDH1A1 and exploit the presence of a unique Glycine residue to achieve their selectivity. Lastly, these two novel and selective ALDH1A1 inhibitors may serve as chemical tools to better understand the contributions of ALDH1A1 to normal biology and to disease states.

  5. Characterization of Two Distinct Structural Classes of Selective Aldehyde Dehydrogenase 1A1 Inhibitors

    DOE PAGES

    Morgan, Cynthia A.; Hurley, Thomas D.

    2015-01-29

    Aldehyde dehydrogenases (ALDH) catalyze the irreversible oxidation of aldehydes to their corresponding carboxylic acid. Alterations in ALDH1A1 activity are associated with such diverse diseases as cancer, Parkinson’s disease, obesity, and cataracts. Inhibitors of ALDH1A1 could aid in illuminating the role of this enzyme in disease processes. However, there are no commercially available selective inhibitors for ALDH1A1. Here we characterize two distinct chemical classes of inhibitors that are selective for human ALDH1A1 compared to eight other ALDH isoenzymes. The prototypical members of each structural class, CM026 and CM037, exhibit sub-micromolar inhibition constants, but have different mechanisms of inhibition. The crystal structuresmore » of these compounds bound to ALDH1A1 demonstrate that they bind within the aldehyde binding pocket of ALDH1A1 and exploit the presence of a unique Glycine residue to achieve their selectivity. Lastly, these two novel and selective ALDH1A1 inhibitors may serve as chemical tools to better understand the contributions of ALDH1A1 to normal biology and to disease states.« less

  6. Development of Selective Inhibitors for Aldehyde Dehydrogenases Based on Substituted Indole-2,3-diones

    PubMed Central

    2015-01-01

    Aldehyde dehydrogenases (ALDH) participate in multiple metabolic pathways and have been indicated to play a role in several cancerous disease states. Our laboratory is interested in developing novel and selective ALDH inhibitors. We looked to further work recently published by developing a class of isoenzyme-selective inhibitors using similar indole-2,3-diones that exhibit differential inhibition of ALDH1A1, ALDH2, and ALDH3A1. Kinetic and X-ray crystallography data suggest that these inhibitors are competitive against aldehyde binding, forming direct interactions with active-site cysteine residues. The selectivity is precise in that these compounds appear to interact directly with the catalytic nucleophile, Cys243, in ALDH3A1 but not in ALDH2. In ALDH2, the 3-keto group is surrounded by the adjacent Cys301/303. Surprisingly, the orientation of the interaction changes depending on the nature of the substitutions on the basic indole ring structure and correlates well with the observed structure–activity relationships for each ALDH isoenzyme. PMID:24444054

  7. Biological Evaluation and 3D-QSAR Studies of Curcumin Analogues as Aldehyde Dehydrogenase 1 Inhibitors

    PubMed Central

    Wang, Hui; Du, Zhiyun; Zhang, Changyuan; Tang, Zhikai; He, Yan; Zhang, Qiuyan; Zhao, Jun; Zheng, Xi

    2014-01-01

    Aldehyde dehydrogenase 1 (ALDH1) is reported as a biomarker for identifying some cancer stem cells, and down-regulation or inhibition of the enzyme can be effective in anti-drug resistance and a potent therapeutic for some tumours. In this paper, the inhibitory activity, mechanism mode, molecular docking and 3D-QSAR (three-dimensional quantitative structure activity relationship) of curcumin analogues (CAs) against ALDH1 were studied. Results demonstrated that curcumin and CAs possessed potent inhibitory activity against ALDH1, and the CAs compound with ortho di-hydroxyl groups showed the most potent inhibitory activity. This study indicates that CAs may represent a new class of ALDH1 inhibitor. PMID:24840575

  8. Contribution of aldehyde oxidase, xanthine oxidase, and aldehyde dehydrogenase on the oxidation of aromatic aldehydes.

    PubMed

    Panoutsopoulos, Georgios I; Kouretas, Demetrios; Beedham, Christine

    2004-10-01

    Aliphatic aldehydes have a high affinity toward aldehyde dehydrogenase activity but are relatively poor substrates of aldehyde oxidase and xanthine oxidase. In addition, the oxidation of xenobiotic-derived aromatic aldehydes by the latter enzymes has not been studied to any great extent. The present investigation compares the relative contribution of aldehyde dehydrogenase, aldehyde oxidase, and xanthine oxidase activities in the oxidation of substituted benzaldehydes in separate preparations. The incubation of vanillin, isovanillin, and protocatechuic aldehyde with either guinea pig liver aldehyde oxidase, bovine milk xanthine oxidase, or guinea pig liver aldehyde dehydrogenase demonstrated that the three aldehyde oxidizing enzymes had a complementary substrate specificity. Incubations were also performed with specific inhibitors of each enzyme (isovanillin for aldehyde oxidase, allopurinol for xanthine oxidase, and disulfiram for aldehyde dehydrogenase) to determine the relative contribution of each enzyme in the oxidation of these aldehydes. Under these conditions, vanillin was rapidly oxidized by aldehyde oxidase, isovanillin was predominantly metabolized by aldehyde dehydrogenase activity, and protocatechuic aldehyde was slowly oxidized, possibly by all three enzymes. Thus, aldehyde oxidase activity may be a significant factor in the oxidation of aromatic aldehydes generated from amines and alkyl benzenes during drug metabolism. In addition, this enzyme may also have a role in the catabolism of biogenic amines such as dopamine and noradrenaline where 3-methoxyphenylacetic acids are major metabolites.

  9. Aldehyde Dehydrogenase Inhibitors: a Comprehensive Review of the Pharmacology, Mechanism of Action, Substrate Specificity, and Clinical Application

    PubMed Central

    Koppaka, Vindhya; Thompson, David C.; Chen, Ying; Ellermann, Manuel; Nicolaou, Kyriacos C.; Juvonen, Risto O.; Petersen, Dennis; Deitrich, Richard A.; Hurley, Thomas D.

    2012-01-01

    Aldehyde dehydrogenases (ALDHs) belong to a superfamily of enzymes that play a key role in the metabolism of aldehydes of both endogenous and exogenous derivation. The human ALDH superfamily comprises 19 isozymes that possess important physiological and toxicological functions. The ALDH1A subfamily plays a pivotal role in embryogenesis and development by mediating retinoic acid signaling. ALDH2, as a key enzyme that oxidizes acetaldehyde, is crucial for alcohol metabolism. ALDH1A1 and ALDH3A1 are lens and corneal crystallins, which are essential elements of the cellular defense mechanism against ultraviolet radiation-induced damage in ocular tissues. Many ALDH isozymes are important in oxidizing reactive aldehydes derived from lipid peroxidation and thereby help maintain cellular homeostasis. Increased expression and activity of ALDH isozymes have been reported in various human cancers and are associated with cancer relapse. As a direct consequence of their significant physiological and toxicological roles, inhibitors of the ALDH enzymes have been developed to treat human diseases. This review summarizes known ALDH inhibitors, their mechanisms of action, isozyme selectivity, potency, and clinical uses. The purpose of this review is to 1) establish the current status of pharmacological inhibition of the ALDHs, 2) provide a rationale for the continued development of ALDH isozyme-selective inhibitors, and 3) identify the challenges and potential therapeutic rewards associated with the creation of such agents. PMID:22544865

  10. Alcohol dehydrogenases from Scheffersomyces stipitis involved in the detoxification of aldehyde inhibitors derived from lignocellulosic biomass conversion.

    PubMed

    Ma, Menggen; Wang, Xu; Zhang, Xiaoping; Zhao, Xianxian

    2013-09-01

    Aldehyde inhibitors such as furfural and 5-hydroxymethylfurfural (HMF) are generated from biomass pretreatment. Scheffersomyces stipitis is able to reduce furfural and HMF to less toxic furanmethanol and furan-2,5-dimethanol; however, the enzymes involved in the reductive reaction still remain unknown. In this study, transcription responses of two known and five putative alcohol dehydrogenase genes from S. stipitis were analyzed under furfural and HMF stress conditions. All the seven alcohol dehydrogenase genes were also cloned and overexpressed for their activity analyses. Our results indicate that transcriptions of SsADH4 and SsADH6 were highly induced under furfural and HMF stress conditions, and the proteins encoded by them exhibited NADH- and/or NADPH-dependent activities for furfural and HMF reduction, respectively. For furfural reduction, NADH-dependent activity was also observed in SsAdh1p and NAD(P)H-dependent activities were also observed in SsAdh5p and SsAdh7p. For HMF reduction, NADPH-dependent activities were also observed in SsAdh5p and SsAdh7p. SsAdh4p displayed the highest NADPH-dependent specific activity and catalytic efficiency for reduction of both furfural and HMF among the seven alcohol dehydrogenases. Enzyme activities of all SsADH proteins were more stable under acidic condition. For most SsADH proteins, the optimum temperature for enzyme activities was 30 °C and more than 50 % enzyme activities remained at 60 °C. Reduction activities of formaldehyde, acetaldehyde, isovaleraldehyde, benzaldehyde, and phenylacetaldehyde were also observed in some SsADH proteins. Our results indicate that multiple alcohol dehydrogenases in S. stipitis are involved in the detoxification of aldehyde inhibitors derived from lignocellulosic biomass conversion.

  11. Two clinical drugs deubiquitinase inhibitor auranofin and aldehyde dehydrogenase inhibitor disulfiram trigger synergistic anti-tumor effects in vitro and in vivo

    PubMed Central

    Liu, Ningning; Hua, Xianliang; Cai, Jianyu; Yang, Changshan; Long, Huidan; Zhao, Chong; Chen, Xin; Lan, Xiaoying; Zang, Dan; Wu, Jinjie; Li, Xiaofen; Shi, Xianping; Wang, Xuejun; Liu, Jinbao

    2016-01-01

    Inhibition of proteasome-associated deubiquitinases (DUBs) is emerging as a novel strategy for cancer therapy. It was recently reported that auranofin (Aur), a gold (I)-containing compound used clinically to treat rheumatoid arthritis, is a proteasome-associated DUB inhibitor. Disulfiram (DSF), an inhibitor of aldehyde dehydrogenase, is currently in clinical use for treating alcoholism. Recent studies have indicated that DSF can also act as an antitumor agent. We investigated the effect of combining DSF and Aur on apoptosis induction and tumor growth in hepatoma cancer cells. Here we report that (i) the combined treatment of Aur and DSF results in synergistic cytotoxicity to hepatoma cells in vitro and in vivo; (ii) Aur and DSF in combination induces caspase activation, endoplasmic reticulum (ER) stress, and reactive oxygen species (ROS) production; (iii) pan-caspase inhibitor z-VAD-FMK could efficiently block apoptosis but not proteasome inhibition induced by Aur and DSF combined treatment, and ROS is not required for Aur+DSF to induce apoptosis. Collectively, we demonstrate a model of synergism between DSF and proteasome-associated DUB inhibitor Aur in the induction of apoptosis in hepatoma cancer cells, identifying a potential novel anticancer strategy for clinical use in the future. PMID:26625200

  12. Two clinical drugs deubiquitinase inhibitor auranofin and aldehyde dehydrogenase inhibitor disulfiram trigger synergistic anti-tumor effects in vitro and in vivo.

    PubMed

    Huang, Hongbiao; Liao, Yuning; Liu, Ningning; Hua, Xianliang; Cai, Jianyu; Yang, Changshan; Long, Huidan; Zhao, Chong; Chen, Xin; Lan, Xiaoying; Zang, Dan; Wu, Jinjie; Li, Xiaofen; Shi, Xianping; Wang, Xuejun; Liu, Jinbao

    2016-01-19

    Inhibition of proteasome-associated deubiquitinases (DUBs) is emerging as a novel strategy for cancer therapy. It was recently reported that auranofin (Aur), a gold (I)-containing compound used clinically to treat rheumatoid arthritis, is a proteasome-associated DUB inhibitor. Disulfiram (DSF), an inhibitor of aldehyde dehydrogenase, is currently in clinical use for treating alcoholism. Recent studies have indicated that DSF can also act as an antitumor agent. We investigated the effect of combining DSF and Aur on apoptosis induction and tumor growth in hepatoma cancer cells. Here we report that (i) the combined treatment of Aur and DSF results in synergistic cytotoxicity to hepatoma cells in vitro and in vivo; (ii) Aur and DSF in combination induces caspase activation, endoplasmic reticulum (ER) stress, and reactive oxygen species (ROS) production; (iii) pan-caspase inhibitor z-VAD-FMK could efficiently block apoptosis but not proteasome inhibition induced by Aur and DSF combined treatment, and ROS is not required for Aur+DSF to induce apoptosis. Collectively, we demonstrate a model of synergism between DSF and proteasome-associated DUB inhibitor Aur in the induction of apoptosis in hepatoma cancer cells, identifying a potential novel anticancer strategy for clinical use in the future.

  13. ALDEHYDE DEHYDROGENASES EXPRESSION DURING POSTNATAL DEVELOPMENT: LIVER VS. LUNG

    EPA Science Inventory

    Aldehydes are highly reactive molecules present in the environment, and can be produced during biotransformation of xenobiotics. Although the lung can be a major target for aldehyde toxicity, development of aldehyde dehydrogenases (ALDHs), which detoxify aldehydes, in lung has be...

  14. ALDEHYDE DEHYDROGENASES EXPRESSION DURING POSTNATAL DEVELOPMENT: LIVER VS. LUNG

    EPA Science Inventory

    Aldehydes are highly reactive molecules present in the environment, and can be produced during biotransformation of xenobiotics. Although the lung can be a major target for aldehyde toxicity, development of aldehyde dehydrogenases (ALDHs), which detoxify aldehydes, in lung has be...

  15. Kinetic study of porcine kidney betaine aldehyde dehydrogenase.

    PubMed

    Figueroa-Soto, C G; Valenzuela-Soto, E M

    2000-03-16

    Porcine kidney betaine aldehyde dehydrogenase (EC 1.2.1.8) kinetic properties were determined at low substrate concentrations. The double-reciprocal plots of initial velocity versus substrate concentration are linear and intersect at the left of the 1/v axis and showed substrate inhibition with betaine aldehyde. Studies of inhibition by NADH and dead-end analogs showed that NADH is a mixed inhibitor against NAD(+) and betaine aldehyde. AMP is competitive with respect to NAD(+) and mixed with betaine aldehyde. Choline is competitive against betaine aldehyde and uncompetitive with respect to NAD(+). The kinetic behavior is consistent with an Iso-Ordered Bi-Bi Steady-State mechanism. Copyright 2000 Academic Press.

  16. Mitochondrial aldehyde dehydrogenase and cardiac diseases

    PubMed Central

    Chen, Che-Hong; Sun, Lihan; Mochly-Rosen, Daria

    2010-01-01

    Numerous conditions promote oxidative stress, leading to the build-up of reactive aldehydes that cause cell damage and contribute to cardiac diseases. Aldehyde dehydrogenases (ALDHs) are important enzymes that eliminate toxic aldehydes by catalysing their oxidation to non-reactive acids. The review will discuss evidence indicating a role for a specific ALDH enzyme, the mitochondrial ALDH2, in combating oxidative stress by reducing the cellular ‘aldehydic load’. Epidemiological studies in humans carrying an inactive ALDH2, genetic models in mice with altered ALDH2 levels, and small molecule activators of ALDH2 all highlight the role of ALDH2 in cardioprotection and suggest a promising new direction in cardiovascular research and the development of new treatments for cardiovascular diseases. PMID:20558439

  17. Structure of aldehyde reductase in ternary complex with coenzyme and the potent 20alpha-hydroxysteroid dehydrogenase inhibitor 3,5-dichlorosalicylic acid: implications for inhibitor binding and selectivity.

    PubMed

    Carbone, Vincenzo; Chung, Roland; Endo, Satoshi; Hara, Akira; El-Kabbani, Ossama

    2008-11-01

    The structure of aldehyde reductase (ALR1) in ternary complex with the coenzyme NADPH and 3,5-dichlorosalicylic acid (DCL), a potent inhibitor of human 20alpha-hydroxysteroid dehydrogenase (AKR1C1), was determined at a resolution of 2.41A. The inhibitor formed a network of hydrogen bonds with the active site residues Trp22, Tyr50, His113, Trp114 and Arg312. Molecular modelling calculations together with inhibitory activity measurements indicated that DCL was a less potent inhibitor of ALR1 (256-fold) when compared to AKR1C1. In AKR1C1, the inhibitor formed a 10-fold stronger binding interaction with the catalytic residue (Tyr55), non-conserved hydrogen bonding interaction with His222, and additional van der Waals contacts with the non-conserved C-terminal residues Leu306, Leu308 and Phe311 that contribute to the inhibitor's selectivity advantage for AKR1C1 over ALR1.

  18. Aldehyde dehydrogenase. Covalent intermediate in aldehyde dehydrogenation and ester hydrolysis.

    PubMed Central

    Blatter, E E; Abriola, D P; Pietruszko, R

    1992-01-01

    4-trans-(NN-Dimethylamino)cinnamaldehyde (an aldehyde, DACA) and 4-trans-(NN-dimethylamino)cinnamoylimidazole (an amide, DACI) have been shown to be substrates for human aldehyde dehydrogenase (EC 1.2.1.3) which form chromophoric covalent intermediates. The spectra of covalent intermediates from both the cytoplasmic (E1) and mitochondrial (E2) isoenzymes derived from DACA and DACI were compared. The spectra were similar when either substrate was used, and also when the two isoenzymes were compared, and resembled that obtained for 4-trnas-(NN-dimethylamino)cinnamoyl-N-acetylcysteine, but differed from the spectrum of 4-trans-(NN-dimethylamino)cinnamoyl ethyl ester. After extensive digestion of the covalent intermediates from both 3H-labelled DACA and DACI with Pronase and purification, the labelled amino acid was identified as cysteine. Covalent intermediates from both DACA and DACI were also digested with trypsin, and labelled peptides were purified by ion-exchange and reverse-phase chromatography. Amino acid sequence analysis showed that the peptide comprising residues 273-307 was labelled by both DACA and DACI. The radioactive label at cysteine residues 301-303 of the primary structure could be unequivocally identified by employing the DACA derivative. Assignment of label to cysteine-302 was achieved by employing iodoacetamide-labelled E1 isoenzyme (iodoacetamide specifically labels cysteine-302), in which case there was no formation of the covalent intermediate from either DACA or DACI. In addition, cysteine-302 is the only cysteine residue conserved in all aldehyde dehydrogenases sequenced. Thus cysteine-302 is the amino acid residue that forms a covalent intermediate with both aldehyde and ester substrates. PMID:1546951

  19. Betaine aldehyde dehydrogenase isozymes of spinach

    SciTech Connect

    Hanson, A.D.; Weretilnyk, E.A.; Weigel, P.

    1986-04-01

    Betaine is synthesized in spinach chloroplasts via the pathway Choline ..-->.. Betaine Aldehyde ..-->.. Betaine; the second step is catalyzed by betaine aldehyde dehydrogenase (BADH). The subcellular distribution of BADH was determined in leaf protoplast lysates; BADH isozymes were separated by 6-9% native PAGE. The chloroplast stromal fraction contains a single BADH isozyme (number1) that accounts for > 80% of the total protoplast activity; the extrachloroplastic fraction has a minor isozyme (number2) which migrates more slowly than number1. Both isozymes appear specific for betaine aldehyde, are more active with NAD than NADP, and show a ca. 3-fold activity increase in salinized leaves. The phenotype of a natural variant of isozyme number1 suggests that the enzyme is a dimer.

  20. Relationships within the aldehyde dehydrogenase extended family.

    PubMed Central

    Perozich, J.; Nicholas, H.; Wang, B. C.; Lindahl, R.; Hempel, J.

    1999-01-01

    One hundred-forty-five full-length aldehyde dehydrogenase-related sequences were aligned to determine relationships within the aldehyde dehydrogenase (ALDH) extended family. The alignment reveals only four invariant residues: two glycines, a phenylalanine involved in NAD binding, and a glutamic acid that coordinates the nicotinamide ribose in certain E-NAD binary complex crystal structures, but which may also serve as a general base for the catalytic reaction. The cysteine that provides the catalytic thiol and its closest neighbor in space, an asparagine residue, are conserved in all ALDHs with demonstrated dehydrogenase activity. Sixteen residues are conserved in at least 95% of the sequences; 12 of these cluster into seven sequence motifs conserved in almost all ALDHs. These motifs cluster around the active site of the enzyme. Phylogenetic analysis of these ALDHs indicates at least 13 ALDH families, most of which have previously been identified but not grouped separately by alignment. ALDHs cluster into two main trunks of the phylogenetic tree. The largest, the "Class 3" trunk, contains mostly substrate-specific ALDH families, as well as the class 3 ALDH family itself. The other trunk, the "Class 1/2" trunk, contains mostly variable substrate ALDH families, including the class 1 and 2 ALDH families. Divergence of the substrate-specific ALDHs occurred earlier than the division between ALDHs with broad substrate specificities. A site on the World Wide Web has also been devoted to this alignment project. PMID:10210192

  1. On the role of microsomal aldehyde dehydrogenase in metabolism of aldehydic products of lipid peroxidation.

    PubMed

    Antonenkov, V D; Pirozhkov, S V; Panchenko, L F

    1987-11-30

    To elucidate a possible role of membrane-bound aldehyde dehydrogenase in the detoxication of aldehydic products of lipid peroxidation, the substrate specificity of the highly purified microsomal enzyme was investigated. The aldehyde dehydrogenase was active with different aliphatic aldehydes including 4-hydroxyalkenals, but did not react with malonic dialdehyde. When Fe/ADP-ascorbate-induced lipid peroxidation of arachidonic acid was carried out in an in vitro system, the formation of products which react with microsomal aldehyde dehydrogenase was observed parallel with malonic dialdehyde accumulation.

  2. Drosophila melanogaster alcohol dehydrogenase: mechanism of aldehyde oxidation and dismutation.

    PubMed

    Winberg, J O; McKinley-McKee, J S

    1998-02-01

    Drosophila alcohol dehydrogenase (Adh) catalyses the oxidation of both alcohols and aldehydes. In the latter case, the oxidation is followed by a reduction of the aldehyde, i.e. a dismutation reaction. At high pH, dismutation is accompanied by a small release of NADH, which is not observed at neutral pH. Previously it has been emphasized that kinetic coefficients obtained by measuring the increase in A340, i.e. the release of NADH at high pH is not a direct measure of the aldehyde oxidation reaction and these values cannot be compared with those for alcohol dehydrogenation. In this article we demonstrate that this is not entirely true, and that the coefficients phiB and phiAB, where B is the aldehyde and A is NAD+, are the same for a dismutation reaction and a simple aldehyde dehydrogenase reaction. Thus the substrate specificity of the aldehyde oxidation reaction can be determined by simply measuring the NADH release. The coefficients for oxidation and dehydrogenation reactions (phi0d and phiAd respectively) are complex and involve the constants for the dismutation reaction. However, dead-end inhibitors can be used to determine the quantitative contribution of the kinetic constants for the aldehyde oxidation and reduction pathways to the phi0d and phiAd coefficients. The combination of dead-end and product inhibitors can be used to determine the reaction mechanism for the aldehyde oxidation pathway. Previously, we showed that with Drosophila Adh, the interconversion between alcohols and aldehydes followed a strictly compulsory ordered pathway, although aldehydes and ketones formed binary complexes with the enzyme. This raised the question regarding the reaction mechanism for the oxidation of aldehydes, i.e. whether a random ordered pathway was followed. In the present work, the mechanism for the oxidation of different aldehydes and the accompanying dismutation reaction with the slow alleloenzyme (AdhS) from Drosophila melanogaster has been studied. To obtain

  3. Drosophila melanogaster alcohol dehydrogenase: mechanism of aldehyde oxidation and dismutation.

    PubMed Central

    Winberg, J O; McKinley-McKee, J S

    1998-01-01

    Drosophila alcohol dehydrogenase (Adh) catalyses the oxidation of both alcohols and aldehydes. In the latter case, the oxidation is followed by a reduction of the aldehyde, i.e. a dismutation reaction. At high pH, dismutation is accompanied by a small release of NADH, which is not observed at neutral pH. Previously it has been emphasized that kinetic coefficients obtained by measuring the increase in A340, i.e. the release of NADH at high pH is not a direct measure of the aldehyde oxidation reaction and these values cannot be compared with those for alcohol dehydrogenation. In this article we demonstrate that this is not entirely true, and that the coefficients phiB and phiAB, where B is the aldehyde and A is NAD+, are the same for a dismutation reaction and a simple aldehyde dehydrogenase reaction. Thus the substrate specificity of the aldehyde oxidation reaction can be determined by simply measuring the NADH release. The coefficients for oxidation and dehydrogenation reactions (phi0d and phiAd respectively) are complex and involve the constants for the dismutation reaction. However, dead-end inhibitors can be used to determine the quantitative contribution of the kinetic constants for the aldehyde oxidation and reduction pathways to the phi0d and phiAd coefficients. The combination of dead-end and product inhibitors can be used to determine the reaction mechanism for the aldehyde oxidation pathway. Previously, we showed that with Drosophila Adh, the interconversion between alcohols and aldehydes followed a strictly compulsory ordered pathway, although aldehydes and ketones formed binary complexes with the enzyme. This raised the question regarding the reaction mechanism for the oxidation of aldehydes, i.e. whether a random ordered pathway was followed. In the present work, the mechanism for the oxidation of different aldehydes and the accompanying dismutation reaction with the slow alleloenzyme (AdhS) from Drosophila melanogaster has been studied. To obtain

  4. A novel nicotinoprotein aldehyde dehydrogenase involved in polyethylene glycol degradation.

    PubMed

    Ohta, T; Tani, A; Kimbara, K; Kawai, F

    2005-09-01

    A gene (pegC) encoding aldehyde dehydrogenase (ALDH) was located 3.4 kb upstream of a gene encoding polyethylene glycol (PEG) dehydrogenase (pegA) in Sphingomonas macrogoltabidus strain 103. ALDH was expressed in Escherichia coli and purified on a Ni-nitrilotriacetic acid agarose column. The recombinant enzyme was a homotetramer consisting of four 46.1-kDa subunits. The alignment of the putative amino acid sequence of the cloned enzyme showed high similarity with a group of NAD(P)-dependent ALDHs (identity 36-52%); NAD-binding domains (Rossmann fold and four glycine residues) and catalytic residues (Glu225 and Cys259) were well conserved. The cofactor, which was extracted from the purified enzyme, was tightly bound to the enzyme and identified as NADP. The enzyme contained 0.94 mol NADP per subunit. The enzyme was activated by Ca(2+), but by no other metals; no metal (Zn, Fe, Mg, or Mn) was detected in the purified recombinant enzyme. Activity was inhibited by p-chloromercuric benzoate, and heavy metals such as Hg, Cu, Pb and Cd, indicating that a cysteine residue is involved in the activity. Enzyme activity was independent of N,N-dimethyl-p-nitrosoaniline as an electron acceptor. Trans-4-(N,N-dimethylamino)-cinnamaldehyde was not oxidized as a substrate, but the compound worked as an inhibitor for the enzyme, as did pyrazole. The enzyme acted on n-aldehydes C(2)-C(14)) and PEG-aldehydes. Thus the enzyme was concluded to be a novel Ca(2+)-activating nicotinoprotein (NADP-containing) PEG-aldehyde dehydrogenase involved in the degradation of PEG in S. macrogoltabidus strain 103.

  5. Aldehyde dehydrogenase is used by cancer cells for energy metabolism

    PubMed Central

    Kang, Joon Hee; Lee, Seon-Hyeong; Hong, Dongwan; Lee, Jae-Seon; Ahn, Hee-Sung; Ahn, Ju-Hyun; Seong, Tae Wha; Lee, Chang-Hun; Jang, Hyonchol; Hong, Kyeong Man; Lee, Cheolju; Lee, Jae-Ho; Kim, Soo-Youl

    2016-01-01

    We found that non-small-cell lung cancer (NSCLC) cells express high levels of multiple aldehyde dehydrogenase (ALDH) isoforms via an informatics analysis of metabolic enzymes in NSCLC and immunohistochemical staining of NSCLC clinical tumor samples. Using a multiple reaction-monitoring mass spectrometry analysis, we found that multiple ALDH isozymes were generally abundant in NSCLC cells compared with their levels in normal IMR-90 human lung cells. As a result of the catalytic reaction mediated by ALDH, NADH is produced as a by-product from the conversion of aldehyde to carboxylic acid. We hypothesized that the NADH produced by ALDH may be a reliable energy source for ATP production in NSCLC. This study revealed that NADH production by ALDH contributes significantly to ATP production in NSCLC. Furthermore, gossypol, a pan-ALDH inhibitor, markedly reduced the level of ATP. Gossypol combined with phenformin synergistically reduced the ATP levels, which efficiently induced cell death following cell cycle arrest. PMID:27885254

  6. Targeting Aldehyde Dehydrogenase 2: New Therapeutic Opportunities

    PubMed Central

    Chen, Che-Hong; Ferreira, Julio Cesar Batista; Gross, Eric R.; Mochly-Rosen, Daria

    2014-01-01

    A family of detoxifying enzymes called aldehyde dehydrogenases (ALDHs) has been a subject of recent interest, as its role in detoxifying aldehydes that accumulate through metabolism and to which we are exposed from the environment has been elucidated. Although the human genome has 19 ALDH genes, one ALDH emerges as a particularly important enzyme in a variety of human pathologies. This ALDH, ALDH2, is located in the mitochondrial matrix with much known about its role in ethanol metabolism. Less known is a new body of research to be discussed in this review, suggesting that ALDH2 dysfunction may contribute to a variety of human diseases including cardiovascular diseases, diabetes, neurodegenerative diseases, stroke, and cancer. Recent studies suggest that ALDH2 dysfunction is also associated with Fanconi anemia, pain, osteoporosis, and the process of aging. Furthermore, an ALDH2 inactivating mutation (termed ALDH2*2) is the most common single point mutation in humans, and epidemiological studies suggest a correlation between this inactivating mutation and increased propensity for common human pathologies. These data together with studies in animal models and the use of new pharmacological tools that activate ALDH2 depict a new picture related to ALDH2 as a critical health-promoting enzyme. PMID:24382882

  7. Halophile aldehyde dehydrogenase from Halobacterium salinarum.

    PubMed

    Kim, Hyo-Jeong; Joo, Won-A; Cho, Chang-Won; Kim, Chan-Wha

    2006-01-01

    Halobacterium salinarum is a member of the halophilic archaea. In the present study, H. salinarum was cultured at various NaCl concentrations (3.5, 4.3, and 6.0 M NaCl), and its proteome was determined and identificated via proteomics technique. We detected 14 proteins which were significantly down-regulated in 3.5 M and/or 6 M NaCl. Among the identified protein spots, aldehyde dehydrogenase (ALDH) was selected for evaluation with regard to its potential applications in industry. The most effective metabolism function exhibited by ALDH is the oxidation of aldehydes to carboxylic acids. The ALDH gene from H. salinarum (1.5 kb fragment) was amplified by PCR and cloned into the E. coli strain, BL21 (DE3), with the pGEX-KG vector. We subsequently analyzed the enzyme activity of the recombinant ALDH (54 kDa) at a variety of salt concentrations. The purified recombinant ALDH from H. salinarum exhibited the most pronounced activity at 1 M NaCl. Therefore, the ALDH from H.salinarum is a halophilic enzyme, and may prove useful for applications in hypersaline environments.

  8. Derivatives of cinnamic acid interact with the nucleotide binding site of mitochondrial aldehyde dehydrogenase. Effects on the dehydrogenase reaction and stimulation of esterase activity by nucleotides.

    PubMed

    Poole, R C; Bowden, N J; Halestrap, A P

    1993-04-22

    A wide variety of cinnamic acid derivatives are inhibitors of the low Km mitochondrial aldehyde dehydrogenase. Two of the most potent inhibitors are alpha-cyano-3,4-dihydroxythiocinnamamide (Ki0.6 microM) and alpha-cyano-3,4,5-trihydroxycinnamonitrile (Ki2.6 microM). With propionaldehyde as substrate the inhibition by these compounds was competitive with respect to NAD+. alpha-Fluorocinnamate was a much less effective inhibitor of the enzyme, with mixed behaviour towards NAD+, but with a major competitive component. These cinnamic acid derivatives were ineffective as inhibitors of the aldehyde dehydrogenase-catalysed hydrolysis of p-nitrophenyl acetate, but inhibited the ability of NAD+ and NADH to activate this activity. Inhibition of the stimulation of esterase activity was competitive with respect to NAD+ and NADH, and the derived Ki values were the same as for inhibition of dehydrogenase activity. NAD+, but not acetaldehyde, could elute the low Km aldehyde dehydrogenase from alpha-cyanocinnamate-Sepharose, to which the enzyme binds specifically (Poole RC and Halestrap AP, Biochem J 259: 105-110, 1989). The cinnamic acid derivatives have little effect on lactate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase or a high Km aldehyde dehydrogenase present in rat liver mitochondria. It is concluded that some cinnamic acid derivatives are potent inhibitors of the low Km aldehyde dehydrogenase, by competing with NAD+/NADH for binding to the enzyme. They are much less effective as inhibitors of other NAD(+)-dependent dehydrogenases.

  9. Human liver aldehyde dehydrogenase: coenzyme binding

    SciTech Connect

    Kosley, L.L.; Pietruszko, R.

    1987-05-01

    The binding of (U-/sup 14/C) NAD to mitochondrial (E2) and cytoplasmin(E1) aldehyde dehydrogenase was measured by gel filtration and sedimentation techniques. The binding data for NAD and (E1) yielded linear Scatchard plots giving a dissociation constant of 25 (+/- 8) uM and the stoichiometry of 2 mol of NAD bound per mol of E1. The binding data for NAD and (E2) gave nonlinear Scatchard plots. The binding of NADH to E2 was measured via fluorescence enhancement; this could not be done with E1 because there was no signal. The dissociation constant for E2 by this technique was 0.7 (+/- 0.4) uM and stoichiometry of 1.0 was obtained. The binding of (U-/sup 14/C) NADH to (E1) and (E2) was also measured by the sedimentation technique. The binding data for (E1) and NADH gave linear Scatchard plots giving a dissociation constant of 13 (+/- 6) uM and the stoichiometry of 2.0. The binding data for NADH to (E2) gave nonlinear Scatchard plots. With (E1), the dissociation constants for both NAD and NADH are similar to those determined kinetically, but the stoichiometry is only half of that found by stopped flow technique. With (E2) the dissociation constant by fluorometric procedure was 2 orders of magnitude less than that from catalytic reaction.

  10. Salivary aldehyde dehydrogenase: activity towards aromatic aldehydes and comparison with recombinant ALDH3A1.

    PubMed

    Giebułtowicz, Joanna; Wolinowska, Renata; Sztybor, Anna; Pietrzak, Monika; Wroczyński, Piotr; Wierzchowski, Jacek

    2009-07-02

    A series of aromatic aldehydes was examined as substrates for salivary aldehyde dehydrogenase (sALDH) and the recombinant ALDH3A1. Para-substituted benzaldehydes, cinnamic aldehyde and 2-naphthaldehydes were found to be excellent substrates, and kinetic parameters for both salivary and recombinant ALDH were nearly identical. It was demonstrated that for the fluorogenic naphthaldehydes the only produced reaction product after incubation in saliva is the carboxylate.

  11. Inhibition of porcine kidney betaine aldehyde dehydrogenase by hydrogen peroxide.

    PubMed

    Rosas-Rodríguez, Jesús A; Figueroa-Soto, Ciria G; Valenzuela-Soto, Elisa M

    2010-01-01

    Renal hyperosmotic conditions may produce reactive oxygen species, which could have a deleterious effect on the enzymes involved in osmoregulation. Hydrogen peroxide was used to provoke oxidative stress in the environment of betaine aldehyde dehydrogenase in vitro. Enzyme activity was reduced as hydrogen peroxide concentration was increased. Over 50% of the enzyme activity was lost at 100 μM hydrogen peroxide at two temperatures tested. At pH 8.0, under physiological ionic strength conditions, peroxide inhibited the enzyme. Initial velocity assays of betaine aldehyde dehydrogenase in the presence of hydrogen peroxide (0-200 μM) showed noncompetitive inhibition with respect to NAD(+) or to betaine aldehyde at saturating concentrations of the other substrate at pH 7.0 or 8.0. Inhibition data showed that apparent V(max) decreased 40% and 26% under betaine aldehyde and NAD(+) saturating concentrations at pH 8.0, while at pH 7.0 V(max) decreased 40% and 29% at betaine aldehyde and NAD(+) saturating concentrations. There was little change in apparent Km(NAD) at either pH, while Km(BA) increased at pH 7.0. K(i) values at pH 8 and 7 were calculated. Our results suggest that porcine kidney betaine aldehyde dehydrogenase could be inhibited by hydrogen peroxide in vivo, thus compromising the synthesis of glycine betaine.

  12. DEVELOPMENTAL EXPRESSION OF ALDEHYDE DEHYDROGENASE IN RAT: A COMPARISON OF LIVER AND LUNG DEVELOPMENT

    EPA Science Inventory

    Metabolism is one of the major determinants for age-related susceptibility changes to chemicals. Aldehydes are highly reactive molecules present in the environment and can be produced during biotransformation of xenobiotics. Aldehyde dehydrogenases (ALDH) are important in aldehyd...

  13. DEVELOPMENTAL EXPRESSION OF ALDEHYDE DEHYDROGENASE IN RAT: A COMPARISON OF LIVER AND LUNG DEVELOPMENT

    EPA Science Inventory

    Metabolism is one of the major determinants for age-related susceptibility changes to chemicals. Aldehydes are highly reactive molecules present in the environment and can be produced during biotransformation of xenobiotics. Aldehyde dehydrogenases (ALDH) are important in aldehyd...

  14. Inhibition effects of furfural on alcohol dehydrogenase, aldehyde dehydrogenase and pyruvate dehydrogenase.

    PubMed Central

    Modig, Tobias; Lidén, Gunnar; Taherzadeh, Mohammad J

    2002-01-01

    The kinetics of furfural inhibition of the enzymes alcohol dehydrogenase (ADH; EC 1.1.1.1), aldehyde dehydrogenase (AlDH; EC 1.2.1.5) and the pyruvate dehydrogenase (PDH) complex were studied in vitro. At a concentration of less than 2 mM furfural was found to decrease the activity of both PDH and AlDH by more than 90%, whereas the ADH activity decreased by less than 20% at the same concentration. Furfural inhibition of ADH and AlDH activities could be described well by a competitive inhibition model, whereas the inhibition of PDH was best described as non-competitive. The estimated K(m) value of AlDH for furfural was found to be about 5 microM, which was lower than that for acetaldehyde (10 microM). For ADH, however, the estimated K(m) value for furfural (1.2 mM) was higher than that for acetaldehyde (0.4 mM). The inhibition of the three enzymes by 5-hydroxymethylfurfural (HMF) was also measured. The inhibition caused by HMF of ADH was very similar to that caused by furfural. However, HMF did not inhibit either AlDH or PDH as severely as furfural. The inhibition effects on the three enzymes could well explain previously reported in vivo effects caused by furfural and HMF on the overall metabolism of Saccharomyces cerevisiae, suggesting a critical role of these enzymes in the observed inhibition. PMID:11964178

  15. Enantioselective oxidation of aldehydes catalyzed by alcohol dehydrogenase.

    PubMed

    Könst, Paul; Merkens, Hedda; Kara, Selin; Kochius, Svenja; Vogel, Andreas; Zuhse, Ralf; Holtmann, Dirk; Arends, Isabel W C E; Hollmann, Frank

    2012-09-24

    Teaching old dogs new tricks: Alcohol dehydrogenases (ADHs) may be established redox biocatalysts but they still are good for a few surprises. ADHs can be used to oxidize aldehydes, and this was demonstrated by the oxidative dynamic kinetic resolution of profens. In the presence of a suitable cofactor regeneration system, this reaction can occur with high selectivity. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Human aldehyde dehydrogenase-catalyzed oxidation of ethylene glycol ether aldehydes.

    PubMed

    Gross, Aaron; Ong, Ta Ren; Grant, Rainer; Hoffmann, Todd; Gregory, Daniel D; Sreerama, Lakshmaiah

    2009-03-16

    Ethylene glycol ethers (EGEs) are primary alcohols commonly used as solvents in numerous household and industrial products. Exposure to EGEs has been correlated with delayed encephalopathy, metabolic acidosis, sub-fertility and spermatotoxicity in humans. In addition, they also cause teratogenesis, carcinogenesis, hemolysis, etc., in various animal models. Metabolism EGEs parallels ethanol metabolism, i.e., EGEs are first converted to 2-alkoxy acetaldehydes (EGE aldehydes) by alcohol dehydrogenases, and then to alkoxyacetic acids by aldehyde dehydrogenases (ALDHs). The acid metabolite of EGEs is considered responsible for toxicities associated with EGEs. The role of human ALDHs in EGE metabolism is not clear; accordingly, we have investigated the ability of five different human ALDHs (ALDH1A1, ALDH2, ALDH3A1, ALDH5A1 and ALDH9A1) to catalyze the oxidation of various EGE aldehydes. The EGE aldehydes used in this study were synthesized via Swern oxidation. All of the human ALDHs were purified from human cDNA clones over-expressing these enzymes in E. coli. The ALDHs tested, so far, differentially catalyze the oxidation of EGE aldehydes to their corresponding acids (K(m) values range from approximately 10 microM to approximately 20.0mM). As judged by V(max)/K(m) ratios, short-chain alkyl-group containing EGE aldehydes are oxidized to their acids more efficiently by ALDH2, whereas aryl- and long-chain alkyl-group containing EGE aldehydes are oxidized to their acid more efficiently by ALDH3A1. Given the product of ALDH-catalyzed reaction is toxic, this process should be considered as a bio-activation (toxification) process.

  17. Effects of herbal infusions, tea and carbonated beverages on alcohol dehydrogenase and aldehyde dehydrogenase activity.

    PubMed

    Li, Sha; Gan, Li-Qin; Li, Shu-Ke; Zheng, Jie-Cong; Xu, Dong-Ping; Li, Hua-Bin

    2014-01-01

    Various alcoholic beverages containing different concentrations of ethanol are widely consumed, and excessive alcohol consumption may result in serious health problems. The consumption of alcoholic beverages is often accompanied by non-alcoholic beverages, such as herbal infusions, tea and carbonated beverages to relieve drunk symptoms. The aim of this study was to supply new information on the effects of these beverages on alcohol metabolism for nutritionists and the general public, in order to reduce problems associated with excessive alcohol consumption. The effects of 57 kinds of herbal infusions, tea and carbonated beverages on alcohol dehydrogenase and aldehyde dehydrogenase activity were evaluated. Generally, the effects of these beverages on alcohol dehydrogenase and aldehyde dehydrogenase activity are very different. The results suggested that some beverages should not be drank after excessive alcohol consumption, and several beverages may be potential dietary supplements for the prevention and treatment of problems related to excessive alcohol consumption.

  18. Monitoring of fatty aldehyde dehydrogenase by formation of pyrenedecanoic acid from pyrenedecanal

    PubMed Central

    Keller, Markus A.; Watschinger, Katrin; Golderer, Georg; Maglione, Manuel; Sarg, Bettina; Lindner, Herbert H.; Werner-Felmayer, Gabriele; Terrinoni, Alessandro; Wanders, Ronald J. A.; Werner, Ernst R.

    2010-01-01

    Fatty aldehyde dehydrogenase (EC 1.2.1.48) converts long-chain fatty aldehydes to the corresponding acids. Deficiency in this enzyme causes the Sjogren Larsson Syndrome, a rare inherited disorder characterized by ichthyosis, spasticity, and mental retardation. Using a fluorescent aldehyde, pyrenedecanal, and HPLC with fluorescence detection, we developed a novel method to monitor fatty aldehyde dehydrogenase activity by quantification of the product pyrenedecanoic acid together with the substrate pyrenedecanal and possible side products, such as aldehyde adducts. As shown with recombinant enzymes, pyrenedecanal showed a high preference for fatty aldehyde dehydrogenase compared with other aldehyde dehydrogenases. The method allowed detection of fatty aldehyde dehydrogenase activity in nanogram amounts of microsomal or tissue protein and microgram amounts of Sjogren Larsson syndrome patients' skin fibroblast protein. It could successfully be adapted for the analysis of fatty aldehyde dehydrogenase activity in gel slices derived from low-temperature SDS-PAGE, showing that fatty aldehyde dehydrogenase activity from solubilized rat liver microsomes migrates as a dimer. Thus, monitoring of pyrenedecanoic acid formation from pyrenedecanal by HPLC with fluorescence detection provides a robust and sensitive method for determination of fatty aldehyde dehydrogenase activity. PMID:19965611

  19. Functional Specialization of Maize Mitochondrial Aldehyde Dehydrogenases1

    PubMed Central

    Liu, Feng; Schnable, Patrick S.

    2002-01-01

    The maize (Zea mays) rf2a and rf2b genes both encode homotetrameric aldehyde dehydrogenases (ALDHs). The RF2A protein was shown previously to accumulate in the mitochondria. In vitro import experiments and ALDH assays on mitochondrial extracts from rf2a mutant plants established that the RF2B protein also accumulates in the mitochondria. RNA gel-blot analyses and immunohistolocation experiments revealed that these two proteins have only partially redundant expression patterns in organs and cell types. For example, RF2A, but not RF2B, accumulates to high levels in the tapetal cells of anthers. Kinetic analyses established that RF2A and RF2B have quite different substrate specificities; although RF2A can oxidize a broad range of aldehydes, including aliphatic aldehydes and aromatic aldehydes, RF2B can oxidize only short-chain aliphatic aldehydes. These two enzymes also have different pH optima and responses to changes in substrate concentration. In addition, RF2A, but not RF2B or any other natural ALDHs, exhibits positive cooperativity. These functional specializations may explain why many species have two mitochondrial ALDHs. This study provides data that serve as a basis for identifying the physiological pathway by which the rf2a gene participates in normal anther development and the restoration of Texas cytoplasm-based male sterility. For example, the observations that Texas cytoplasm anthers do not accumulate elevated levels of reactive oxygen species or lipid peroxidation and the kinetic features of RF2A make it unlikely that rf2a restores fertility by preventing premature programmed cell death. PMID:12481049

  20. A coniferyl aldehyde dehydrogenase gene from Pseudomonas sp. strain HR199 enhances the conversion of coniferyl aldehyde by Saccharomyces cerevisiae.

    PubMed

    Adeboye, Peter Temitope; Olsson, Lisbeth; Bettiga, Maurizio

    2016-07-01

    The conversion of coniferyl aldehyde to cinnamic acids by Saccharomyces cerevisiae under aerobic growth conditions was previously observed. Bacteria such as Pseudomonas have been shown to harbor specialized enzymes for converting coniferyl aldehyde but no comparable enzymes have been identified in S. cerevisiae. CALDH from Pseudomonas was expressed in S. cerevisiae. An acetaldehyde dehydrogenase (Ald5) was also hypothesized to be actively involved in the conversion of coniferyl aldehyde under aerobic growth conditions in S. cerevisiae. In a second S. cerevisiae strain, the acetaldehyde dehydrogenase (ALD5) was deleted. A prototrophic control strain was also engineered. The engineered S. cerevisiae strains were cultivated in the presence of 1.1mM coniferyl aldehyde under aerobic condition in bioreactors. The results confirmed that expression of CALDH increased endogenous conversion of coniferyl aldehyde in S. cerevisiae and ALD5 is actively involved with the conversion of coniferyl aldehyde in S. cerevisiae. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  1. Non-P450 aldehyde oxidizing enzymes: the aldehyde dehydrogenase superfamily

    PubMed Central

    Marchitti, Satori A; Brocker, Chad; Stagos, Dimitrios; Vasiliou, Vasilis

    2009-01-01

    Background Aldehydes are highly reactive molecules. While several non-P450 enzyme systems participate in their metabolism, one of the most important is the aldehyde dehydrogenase (ALDH) superfamily, composed of NAD(P)+-dependent enzymes that catalyze aldehyde oxidation. Objective This article presents a review of what is currently known about each member of the human ALDH superfamily including the pathophysiological significance of these enzymes. Methods Relevant literature involving all members of the human ALDH family was extensively reviewed, with the primary focus on recent and novel findings. Conclusion To date, 19 ALDH genes have been identified in the human genome and mutations in these genes and subsequent inborn errors in aldehyde metabolism are the molecular basis of several diseases, including Sjögren-Larsson syndrome, type II hyperprolinemia, γ-hydroxybutyric aciduria and pyridoxine-dependent seizures. ALDH enzymes also play important roles in embryogenesis and development, neurotransmission, oxidative stress and cancer. Finally, ALDH enzymes display multiple catalytic and non-catalytic functions including ester hydrolysis, antioxidant properties, xenobiotic bioactivation and UV light absorption. PMID:18611112

  2. Reversible, partial inactivation of plant betaine aldehyde dehydrogenase by betaine aldehyde: mechanism and possible physiological implications.

    PubMed

    Zárate-Romero, Andrés; Murillo-Melo, Darío S; Mújica-Jiménez, Carlos; Montiel, Carmina; Muñoz-Clares, Rosario A

    2016-04-01

    In plants, the last step in the biosynthesis of the osmoprotectant glycine betaine (GB) is the NAD(+)-dependent oxidation of betaine aldehyde (BAL) catalysed by some aldehyde dehydrogenase (ALDH) 10 enzymes that exhibit betaine aldehyde dehydrogenase (BADH) activity. Given the irreversibility of the reaction, the short-term regulation of these enzymes is of great physiological relevance to avoid adverse decreases in the NAD(+):NADH ratio. In the present study, we report that the Spinacia oleracea BADH (SoBADH) is reversibly and partially inactivated by BAL in the absence of NAD(+)in a time- and concentration-dependent mode. Crystallographic evidence indicates that the non-essential Cys(450)(SoBADH numbering) forms a thiohemiacetal with BAL, totally blocking the productive binding of the aldehyde. It is of interest that, in contrast to Cys(450), the catalytic cysteine (Cys(291)) did not react with BAL in the absence of NAD(+) The trimethylammonium group of BAL binds in the same position in the inactivating or productive modes. Accordingly, BAL does not inactivate the C(450)SSoBADH mutant and the degree of inactivation of the A(441)I and A(441)C mutants corresponds to their very different abilities to bind the trimethylammonium group. Cys(450)and the neighbouring residues that participate in stabilizing the thiohemiacetal are strictly conserved in plant ALDH10 enzymes with proven or predicted BADH activity, suggesting that inactivation by BAL is their common feature. Under osmotic stress conditions, this novel partial and reversible covalent regulatory mechanism may contribute to preventing NAD(+)exhaustion, while still permitting the synthesis of high amounts of GB and avoiding the accumulation of the toxic BAL.

  3. Increased salivary aldehyde dehydrogenase 1 in non-reticular oral lichen planus.

    PubMed

    Mansourian, Arash; Shanbehzadeh, Najmeh; Kia, Seyed Javad; Moosavi, Mahdieh-Sadat

    2017-01-01

    Oral lichen planus is a potentially malignant disorder. One of the malignant transformation markers is cancer stem cells. One of the proposed marker for the detection of cancer stem cells's in head and neck cancer is aldehyde dehydrogenase. Recently it is shown that aldehyde dehydrogenase 1 expression in tissue samples is associated with oral lichen planus malignant transformation. This study evaluates salivary aldehyde dehydrogenase 1 in oral lichen planus. Thirty patients and 30 age and sex-matched healthy volunteers were recruited. Oral lichen planus was diagnosed based on the modified World Health Organization criteria. Subjects in the case group were divided into reticular and non-reticular forms. Unstimulated salivary samples were collected at 10-12 AM. Saliva concentrations of aldehyde dehydrogenase 1 were measured by ELISA. The differences between aldehyde dehydrogenase levels in the oral lichen planus group compared with the control group were not significant but aldehyde dehydrogenase in non-reticular oral lichen planus was significantly higher than that of the reticular form. This is a cross-sectional study, thus longitudinal studies in oral lichen planus may present similar or different results. The mechanism of malignant transformation in oral lichen planus is not defined. Previous analyses revealed that the aldehyde dehydrogenase 1 expression is significantly correlated with increased risk of transformation. This finding is consistent with our results because in the erosive and ulcerative forms of oral lichen planus, which have an increased risk of transformation, salivary aldehyde dehydrogenase 1 was overexpressed. A higher salivary aldehyde dehydrogenase level in non-reticular oral lichen planus can be a defensive mechanism against higher oxidative stress in these groups. Aldehyde dehydrogenase may be one of the malignant transformation markers in oral lichen planus. Further studies are needed for introducing aldehyde dehydrogenase as a prognostic

  4. Aldehyde Dehydrogenases in Cellular Responses to Oxidative/electrophilic Stress

    PubMed Central

    Singh, Surendra; Brocker, Chad; Koppaka, Vindhya; Ying, Chen; Jackson, Brian; Matsumoto, Akiko; Thompson, David C.; Vasiliou, Vasilis

    2013-01-01

    Reactive oxygen species (ROS) are continuously generated within living systems and the inability to manage ROS load leads to elevated oxidative stress and cell damage. Oxidative stress is coupled to the oxidative degradation of lipid membranes, also known as lipid peroxidation. This process generates over 200 types of aldehydes, many of which are highly reactive and toxic. Aldehyde dehydrogenases (ALDHs) metabolize endogenous and exogenous aldehydes and thereby mitigate oxidative/electrophilic stress in prokaryotic and eukaryotic organisms. ALDHs are found throughout the evolutionary gamut, from single celled organisms to complex multicellular species. Not surprisingly, many ALDHs in evolutionarily distant, and seemingly unrelated, species perform similar functions, including protection against a variety of environmental stressors like dehydration and ultraviolet radiation. The ability to act as an ‘aldehyde scavenger’ during lipid peroxidation is another ostensibly universal ALDH function found across species. Up-regulation of ALDHs is a stress response in bacteria (environmental and chemical stress), plants (dehydration, salinity and oxidative stress), yeast (ethanol exposure and oxidative stress), Caenorhabditis elegans (lipid peroxidation) and mammals (oxidative stress and lipid peroxidation). Recent studies have also identified ALDH activity as an important feature of cancer stem cells. In these cells, ALDH expression helps abrogate oxidative stress and imparts resistance against chemotherapeutic agents such as oxazaphosphorine, taxane and platinum drugs. The ALDH superfamily represents a fundamentally important class of enzymes that significantly contributes to the management of electrophilic/oxidative stress within living systems. Mutations in various ALDHs are associated with a variety of pathological conditions in humans, underscoring the fundamental importance of these enzymes in physiological and pathological processes. PMID:23195683

  5. [The role of hepatic and erythrocyte aldehyde dehydrogenase in the development of burn toxemia in rats].

    PubMed

    Solov'eva, A G

    2009-01-01

    The study was designed to examine catalytic properties of non-specific aldehyde dehydrogenase from rat liver and erythrocyte as the main markers of endogenous intoxication after burn. Enzymatic activity was assayed from changes in the rate of NADH synthesis during acetaldehyde oxidation. Burn was shown to decrease it both in the liver and in erythrocytes which resulted in the accumulation of toxic aldehydes and the development of intoxication. Simultaneous fall in alcohol dehydrogenase and lactate dehydrogenase activities is supposed to contribute to the decrease of aldehyde dehydrogenase activity as a result of thermal injury.

  6. A red-shifted fluorescent substrate for aldehyde dehydrogenase

    PubMed Central

    Minn, Il; Wang, Haofan; Mease, Ronnie C.; Byun, Youngjoo; Yang, Xing; Wang, Julia; Leach, Steven D.; Pomper, Martin G.

    2014-01-01

    Selection of cells positive for aldehyde dehydrogenase (ALDH) activity from a green fluorescent background is difficult with existing reagents. Here we report a red-shifted fluorescent substrate for ALDH, AldeRed 588-A, for labeling viable ALDHpos cells. We demonstrate that AldeRed 588-A successfully isolates ALDHhi human hematopoietic stem cells from heterogeneous cord blood mononuclear cells. AldeRed 588-A can be used for multi-color applications to fractionate ALDHpos cells in the presence of green fluorophores including the ALDEFLUOR™ reagent and cells expressing eGFP. AldeRed 588-A stains ALDHpos murine pancreatic centroacinar and terminal duct cells, as visualized by fluorescent microscopy. AldeRed588-A provides a useful tool to select stem cells or study ALDH within a green fluorescent background. PMID:24759454

  7. Inactivation of porcine kidney betaine aldehyde dehydrogenase by hydrogen peroxide.

    PubMed

    Rosas-Rodríguez, Jesús A; Valenzuela-Soto, Elisa M

    2011-05-30

    Concentrated urine formation in the kidney is accompanied by conditions that favor the accumulation of reactive oxygen species (ROS). Under hyperosmotic conditions, medulla cells accumulate glycine betaine, which is an osmolyte synthesized by betaine aldehyde dehydrogenase (BADH, EC 1.2.1.8). All BADHs identified to date have a highly reactive cysteine residue at the active site, and this cysteine is susceptible to oxidation by hydrogen peroxide. Porcine kidney BADH incubated with H(2)O(2) (0-500 μM) lost 25% of its activity. However, pkBADH inactivation by hydrogen peroxide was limited, even after 120 min of incubation. The presence of coenzyme NAD(+) (10-50 μM) increased the extent of inactivation (60%) at 120 min of reaction, but the ligands betaine aldehyde (50 and 500 μM) and glycine betaine (100 mM) did not change the rate or extent of inactivation as compared to the reaction without ligand. 2-Mercaptoethanol and dithiothreitol, but not reduced glutathione, were able to restore enzyme activity. Mass spectrometry analysis of hydrogen peroxide inactivated BADH revealed oxidation of M278, M243, M241 and H335 in the absence and oxidation of M94, M327 and M278 in the presence of NAD(+). Molecular modeling of BADH revealed that the oxidized methionine and histidine residues are near the NAD(+) binding site. In the presence of the coenzyme, these oxidized residues are proximal to the betaine aldehyde binding site. None of the oxidized amino acid residues participates directly in catalysis. We suggest that pkBADH inactivation by hydrogen peroxide occurs via disulfide bond formation between vicinal catalytic cysteines (C288 and C289). Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  8. Single amino acid polymorphism in aldehyde dehydrogenase gene superfamily.

    PubMed

    Priyadharshini Christy, J; George Priya Doss, C

    2015-01-01

    The aldehyde dehydrogenase gene superfamily comprises of 19 genes and 3 pseudogenes. These superfamily genes play a vital role in the formation of molecules that are involved in life processes, and detoxification of endogenous and exogenous aldehydes. ALDH superfamily genes associated mutations are implicated in various diseases, such as pyridoxine-dependent seizures, gamma-hydroxybutyric aciduria, type II Hyperprolinemia, Sjogren-Larsson syndrome including cancer and Alzheimer's disease. Accumulation of large DNA variations data especially Single Amino acid Polymorphisms (SAPs) in public databases related to ALDH superfamily genes insisted us to conduct a survey on the disease associated mutations and predict their function impact on protein structure and function. Overall this study provides an update and highlights the importance of pathogenic mutations in associated diseases. Using KD4v and Project HOPE a computational based platform, we summarized all the deleterious properties of SAPs in ALDH superfamily genes by the providing valuable insight into structural alteration rendered due to mutation. We hope this review might provide a way to define the deleteriousness of a SAP and helps to understand the molecular basis of the associated disease and also permits precise diagnosis and treatment in the near future.

  9. Aldehyde dehydrogenase 2 in cardiac protection: a new therapeutic target?

    PubMed Central

    Budas, Grant R; Disatnik, Marie- Hélène; Mochly-Rosen, Daria

    2010-01-01

    Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is emerging as a key enzyme involved in cytoprotection in the heart. ALDH2 mediates both the detoxification of reactive aldehydes such as acetaldehyde and 4-hydroxy-2-nonenal (4-HNE) and the bioactivation of nitroglycerin (GTN) to nitric oxide (NO). In addition, chronic nitrate treatment results in ALDH2 inhibition and contributes to nitrate tolerance. Our lab recently identified ALDH2 to be a key mediator of endogenous cytoprotection. We reported that ALDH2 is phosphorylated and activated by the survival kinase protein kinase C epsilon (PKCε) and found a strong inverse correlation between ALDH2 activity and infarct size. We also identified a small molecule ALDH2 activator (Alda-1) which reduces myocardial infarct size induced by ischemia/reperfusion in vivo. In this review, we discuss evidence that ALDH2 is a key mediator of endogenous survival signaling in the heart, suggest possible cardioprotective mechanisms mediated by ALDH2, and discuss potential clinical implications of these findings. PMID:20005475

  10. Aldehyde dehydrogenase 1A1 in stem cells and cancer

    PubMed Central

    Tomita, Hiroyuki; Tanaka, Kaori; Tanaka, Takuji; Hara, Akira

    2016-01-01

    The human genome contains 19 putatively functional aldehyde dehydrogenase (ALDH) genes, which encode enzymes critical for detoxification of endogenous and exogenous aldehyde substrates through NAD(P)+-dependent oxidation. ALDH1 has three main isotypes, ALDH1A1, ALDH1A2, and ALDH1A3, and is a marker of normal tissue stem cells (SC) and cancer stem cells (CSC), where it is involved in self-renewal, differentiation and self-protection. Experiments with murine and human cells indicate that ALDH1 activity, predominantly attributed to isotype ALDH1A1, is tissue- and cancer-specific. High ALDH1 activity and ALDH1A1 overexpression are associated with poor cancer prognosis, though high ALDH1 and ALDH1A1 levels do not always correlate with highly malignant phenotypes and poor clinical outcome. In cancer therapy, ALDH1A1 provides a useful therapeutic CSC target in tissue types that normally do not express high levels of ALDH1A1, including breast, lung, esophagus, colon and stomach. Here we review the functions and mechanisms of ALDH1A1, the key ALDH isozyme linked to SC populations and an important contributor to CSC function in cancers, and we outline its potential in future anticancer strategies. PMID:26783961

  11. A specific affinity reagent to distinguish aldehyde dehydrogenases and oxidases. Enzymes catalyzing aldehyde oxidation in an adult moth

    SciTech Connect

    Tasayco, M.L.; Prestwich, G.D. )

    1990-02-25

    Aldehyde dehydrogenase (ALDH) and oxidase (AO) enzymes from the tissue extracts of male and female tobacco budworm moth (Heliothis virescens) were identified after electrophoretic protein separation. AO activity was visualized using formazan- or horseradish peroxidase-mediated staining coupled to the AO-catalyzed oxidation of benzaldehyde. A set of six soluble AO enzymes with isoelectric points from pI 4.6 to 5.3 were detected primarily in the antennal extracts. Partially purified antennal AO enzymes also oxidized both (Z)-9-tetradecenal and (Z)-11-hexadecenal, the two major pheromone components of this moth. ALDH activity was detected using a tritium-labeled affinity reagent based on a known irreversible inhibitor of this enzyme. This labeled vinyl ketone, (3H)(Z)-1,11-hexadecadien-3-one, was synthesized and used to covalently modify the soluble ALDH enzymes from tissue extracts. Molecular subunits of potential ALDH enzymes were visualized in the fluorescence autoradiograms of sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated proteins of the antenna, head, and leg tissues. Covalent modification of these protein subunits decreased specifically in the presence of excess pheromone aldehyde or benzaldehyde. Labeled vinyl ketones are thus novel tools for the identification of molecular subunits of ALDH enzymes.

  12. Biochemical properties of rat liver mitochondrial aldehyde dehydrogenase with respect to oxidation of formaldehyde.

    PubMed

    Cinti, D L; Keyes, S R; Lemelin, M A; Denk, H; Schenkman, J B

    1976-03-25

    The oxidation of formaldehyde by rat liver mitochondria in the presence of 50 mM phosphate was enhanced 2-fold by exogenous NAD+. Absolute requirement of NAD+ for formaldehyde oxidation was demonstrated by depleting the mitochondria of their NAD+ content (4.6 nmol/mg of protein), followed by reincorporation of the NAD+ into the depleted mitochondria. Aldehyde (formaldehyde) dehydrogenase activity was completely abolished in the depleted mitochondria, but the enzyme activity was restored to control levels following reincorporation of the pyridine nucleotide. Phosphate stimulation of formaldehyde oxidation could not be explained fully by the phosphate-induced swelling which enhances membrane permeability to NAD+, since stimulation of the enzyme activity by increased phosphate concentrations was still observed in the absence of exogenous NAD+. The Km for formaldehyde oxidation by the mitochondria was found to be 0.38 nM, a value similar to that obtained with varying concentrations of NAD+; both Vmax values were very similar, giving a value of 70 to 80 nmol/min/mg of protein. The pH optimum for the mitochondrial enzyme was 8.0. Inhibition of the enzyme activity by anaerobiosis was apparently due to the inability of the respiratory chain to oxidize the generated NADH. The inhibition of mitochondrial formaldehyde oxidation by succinate was found to be due to a lowering of the NAD+ level in the mitochondria. Succinate also inhibited acetaldehyde oxidation by the mitochondria. Malonate, a competitive inhibitor of succinic dehydrogenase, blocked the inhibitory effect of succinate. The respiratory chain inhibitors, rotenone, and antimycin A plus succinate, strongly inhibited formaldehyde oxidation by apparently the same mechanism, although the crude enzyme preparation (freed from the membrane) was slightly sensitive to rotenone. The mitochondria were subfractionated, and 85% of the enzyme activity was found in the inner membrane fraction (mitoplast). Furthermore, separation

  13. Inhibition of the Aldehyde Dehydrogenase 1/2 Family by Psoralen and Coumarin Derivatives.

    PubMed

    Buchman, Cameron D; Hurley, Thomas D

    2017-03-23

    Aldehyde dehydrogenase 2 (ALDH2), one of 19 ALDH superfamily members, catalyzes the NAD(+)-dependent oxidation of aldehydes to their respective carboxylic acids. In this study, we further characterized the inhibition of four psoralen and coumarin derivatives toward ALDH2 and compared them to the ALDH2 inhibitor daidzin for selectivity against five ALDH1/2 isoenzymes. Compound 2 (Ki = 19 nM) binds within the aldehyde-binding site of the free enzyme species of ALDH2. Thirty-three structural analogs were examined to develop a stronger SAR profile. Seven compounds maintained or improved upon the selectivity toward one of the five ALDH1/2 isoenzymes, including compound 36, a selective inhibitor for ALDH2 (Ki = 2.4 μM), and compound 32, which was 10-fold selective for ALDH1A1 (Ki = 1.2 μM) versus ALDH1A2. Further medicinal chemistry on the compounds' basic scaffold could enhance the potency and selectivity for ALDH1A1 or ALDH2 and generate chemical probes to examine the unique and overlapping functions of the ALDH1/2 isoenzymes.

  14. Targeting Aldehyde Dehydrogenase: a Potential Approach for Cell labeling

    PubMed Central

    Vaidyanathan, Ganesan; Song, Haijing; Affleck, Donna; McDougald, Darryl L.; Storms, Robert W.; Zalutksy, Michael R.; Chin, Bennett B.

    2009-01-01

    Introduction To advance the science and clinical application of stem cell therapy, the availability of a highly sensitive, quantitative, and translational method for tracking stem cells would be invaluable. Because hematopoetic stem cells express high levels of the cytosolic enzyme aldehyde dehydrogenase-1A1 (ALDH1), we sought to develop an agent that is specific to ALDH1 and thus to cells expressing the enzyme. Such an agent might be also helpful in identifying tumors that are resistant to cyclophosphomide chemotherapy because ALDH1 is known to be responsible for this resistance. Methods We developed schemes for the synthesis of two 3radioiodinated aldehdyes—N-formylmethyl-5-[*I]iodopyridine-3-carboxamide ([*I]FMIC) and 4-diethylamino-3-[*I]iodobenzaldehyde ([*I]DEIBA)—at no-carrier-added levels from their respective tin precursors. These agents were evaluated using pure ALDH1 and tumor cells that expressed the enzyme. Results The average radiochemical yields for the synthesis [125I]FMIC and [125I]DEIBA were 70 ± 5% and 47 ± 14%, respectively. ALDH1 converted both compounds to respective acids suggesting their suitability as ALDH1 imaging agents. Although ability of ALDH1 within the cells to oxidize one of these substrates was shown, specific uptake in ALDH-expressing tumor cells could not be demonstrated. Conclusion To pursue this approach for ALDH1 imaging, radiolabeled aldehydes need to be designed such that, in addition to being good substrates for ALDH1, the cognate products should be sufficiently polar so as to be retained within the cells. PMID:19875048

  15. Mechanism of aldehyde oxidation catalyzed by horse liver alcohol dehydrogenase.

    PubMed

    Olson, L P; Luo, J; Almarsson, O; Bruice, T C

    1996-07-30

    The mechanism of oxidation of benzaldehyde to benzoic acid catalyzed by horse liver alcohol dehydrogenase (HLADH) has been investigated using the HLADH structure at 2.1 A resolution with NAD+ and pentafluorobenzyl alcohol in the active site [Ramaswamy et al. (1994) Biochemistry 33,5230-5237]. Constructs for molecular dynamics (MD) investigations with HLADH were obtained by a best-fit superimposition of benzaldehyde or its hydrate on the pentafluorobenzyl alcohol bound to the active site Zn(II)ion. Equilibrium bond lengths, angles, and dihedral parameters for Zn(II) bonding residues His67, Cys46, and Cys174 were obtained from small-molecule X-ray crystal structures and an ab initio-derived parameterization of zinc in HLADH [Ryde, U. (1995) Proteins: Struct., Funct., Genet. 21,40-56]. Dynamic simulations in CHARMM were carried out on the following three constructs to 100 ps: (MD1) enzyme with NAD+, benzaldehyde, and zinc-ligated HO-in the active site; (MD2) enzyme with NAD+ and hydrated benzaldehyde monoanion bound to zinc via the pro-R oxygen, with a proton residing on the pro-S oxygen; and (MD3) enzyme with NAD+ and hydrated benzaldehyde monoanion bound to zinc via the pro-S oxygen, with a proton residing on the pro-R oxygen. Analyses were done of 800 sample conformations taken in the last 40 ps of dynamics. Structures from MD1 and MD3 were used to define the initial spatial arrangements of reactive functionalities for semiempirical PM3 calculations. Using PM3, model systems were calculated of ground states and some transition states for aldehyde hydration, hydride transfer, and subsequent proton shuttling. With benzaldehyde and zinc-bound hydroxide ion in the active site, the oxygen of Zn(II)-OH resided at a distance of 2.8-5.5 A from the aldehyde carbonyl carbon during the dynamics simulation. This may be compared to the PM3 transition state for attack of the Zn(II)-OH oxygen on the benzaldehyde carbonyl carbon, which has an O...C distance of 1.877 A. HLADH

  16. Aldehyde dehydrogenase 2 inhibits inflammatory response and regulates atherosclerotic plaque

    PubMed Central

    Wei, Shu-jian; Zhang, Ming-xiang; Wang, Xu-ping; Yuan, Qiu-huan; Xue, Li; Wang, Jia-li; Cui, Zhao-qiang; Zhang, Yun; Xu, Feng; Chen, Yu-guo

    2016-01-01

    Previous studies demonstrated that aldehyde dehydrogenase 2 (ALDH2) rs671 polymorphism, which eliminates ALDH2 activity down to 1%-6%, is a susceptibility gene for coronary disease. Here we investigated the underlying mechanisms based on our prior clinical and experimental studies. Male apoE−/− mice were transfected with GFP, ALDH2-overexpression and ALDH2-RNAi lentivirus respectively (n=20 each) after constrictive collars were placed around the right common carotid arteries. Consequently, ALDH2 gene silencing led to an increased en face plaque area, more unstable plaque with heavier accumulation of lipids, more macrophages, less smooth muscle cells and collagen, which were associated with aggravated inflammation. However, ALDH2 overexpression displayed opposing effects. We also found that ALDH2 activity decreased in atherosclerotic plaques of human and aged apoE−/− mice. Moreover, in vitro experiments with human umbilical vein endothelial cells further illustrated that, inhibition of ALDH2 activity resulted in elevating inflammatory molecules, an increase of nuclear translocation of NF-κB, and enhanced phosphorylation of NF-κB p65, AP-1 c-Jun, Jun-N terminal kinase and p38 MAPK, while ALDH2 activation could trigger contrary effects. These findings suggested that ALDH2 can influence plaque development and vulnerability, and inflammation via MAPK, NF-κB and AP-1 signaling pathways. PMID:27191745

  17. Aldehyde dehydrogenase inhibition as a pathogenic mechanism in Parkinson disease

    PubMed Central

    Fitzmaurice, Arthur G.; Rhodes, Shannon L.; Lulla, Aaron; Murphy, Niall P.; Lam, Hoa A.; O’Donnell, Kelley C.; Barnhill, Lisa; Casida, John E.; Cockburn, Myles; Sagasti, Alvaro; Stahl, Mark C.; Maidment, Nigel T.; Ritz, Beate; Bronstein, Jeff M.

    2013-01-01

    Parkinson disease (PD) is a neurodegenerative disorder particularly characterized by the loss of dopaminergic neurons in the substantia nigra. Pesticide exposure has been associated with PD occurrence, and we previously reported that the fungicide benomyl interferes with several cellular processes potentially relevant to PD pathogenesis. Here we propose that benomyl, via its bioactivated thiocarbamate sulfoxide metabolite, inhibits aldehyde dehydrogenase (ALDH), leading to accumulation of the reactive dopamine metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL), preferential degeneration of dopaminergic neurons, and development of PD. This hypothesis is supported by multiple lines of evidence. (i) We previously showed in mice the metabolism of benomyl to S-methyl N-butylthiocarbamate sulfoxide, which inhibits ALDH at nanomolar levels. We report here that benomyl exposure in primary mesencephalic neurons (ii) inhibits ALDH and (iii) alters dopamine homeostasis. It induces selective dopaminergic neuronal damage (iv) in vitro in primary mesencephalic cultures and (v) in vivo in a zebrafish system. (vi) In vitro cell loss was attenuated by reducing DOPAL formation. (vii) In our epidemiology study, higher exposure to benomyl was associated with increased PD risk. This ALDH model for PD etiology may help explain the selective vulnerability of dopaminergic neurons in PD and provide a potential mechanism through which environmental toxicants contribute to PD pathogenesis. PMID:23267077

  18. Aldehyde dehydrogenase inhibition as a pathogenic mechanism in Parkinson disease.

    PubMed

    Fitzmaurice, Arthur G; Rhodes, Shannon L; Lulla, Aaron; Murphy, Niall P; Lam, Hoa A; O'Donnell, Kelley C; Barnhill, Lisa; Casida, John E; Cockburn, Myles; Sagasti, Alvaro; Stahl, Mark C; Maidment, Nigel T; Ritz, Beate; Bronstein, Jeff M

    2013-01-08

    Parkinson disease (PD) is a neurodegenerative disorder particularly characterized by the loss of dopaminergic neurons in the substantia nigra. Pesticide exposure has been associated with PD occurrence, and we previously reported that the fungicide benomyl interferes with several cellular processes potentially relevant to PD pathogenesis. Here we propose that benomyl, via its bioactivated thiocarbamate sulfoxide metabolite, inhibits aldehyde dehydrogenase (ALDH), leading to accumulation of the reactive dopamine metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL), preferential degeneration of dopaminergic neurons, and development of PD. This hypothesis is supported by multiple lines of evidence. (i) We previously showed in mice the metabolism of benomyl to S-methyl N-butylthiocarbamate sulfoxide, which inhibits ALDH at nanomolar levels. We report here that benomyl exposure in primary mesencephalic neurons (ii) inhibits ALDH and (iii) alters dopamine homeostasis. It induces selective dopaminergic neuronal damage (iv) in vitro in primary mesencephalic cultures and (v) in vivo in a zebrafish system. (vi) In vitro cell loss was attenuated by reducing DOPAL formation. (vii) In our epidemiology study, higher exposure to benomyl was associated with increased PD risk. This ALDH model for PD etiology may help explain the selective vulnerability of dopaminergic neurons in PD and provide a potential mechanism through which environmental toxicants contribute to PD pathogenesis.

  19. An animal model of human aldehyde dehydrogenase deficiency

    SciTech Connect

    Chang, C.; Mann, J.; Yoshida, A.

    1994-09-01

    The genetic deficiency of ALDH2, a major mitochondrial aldehyde dehydrogenase, is intimately related to alcohol sensitivity and the degree of predisposition to alcoholic diseases in humans. The ultimate biological role of ALDH2 can be exposed by knocking out the ALDH2 gene in an animal model. As the first step for this line of studies, we cloned and characterized the ALDH2 gene from mouse C57/6J strain which is associated with a high alcohol preference. The gene spans 26 kbp and is composed of 13 exons. Embryonic stem cells were transfected with a replacement vector which contains a partially deleted exon3, a positive selection cassette (pPgk Neo), exon 4 with an artificial stop codon, exons 5, 6, 7, and a negative selection cassette (pMCI-Tk). Genomic DNAs prepared from drug resistant clones were analyzed by polymerase chain reaction and by Southern blot analysis to distinguish random integration from homologous recombination. Out of 132 clones examined, 8 had undergone homologous recombination at one of the ALDH2 alleles. The cloned transformed embryonic stem cells with a disrupted ALDH2 allele were injected into blastocysts. Transplantation of the blastocysts into surrogate mother mice yielded chimeric mice. The role of ALDH2 in alcohol preference, alcohol sensitivity and other biological and behavioral characteristics can be elucidated by examining the heterozygous and homozygous mutant strains produced by breeding of chimeric mice.

  20. Aldehyde dehydrogenases in cancer stem cells: potential as therapeutic targets

    PubMed Central

    Clark, David W.

    2016-01-01

    Resistance to current chemotherapeutic or radiation-based cancer treatment strategies is a serious concern. Cancer stem cells (CSCs) are typically able to evade treatment and establish a recurrent tumor or metastasis, and it is these that lead to the majority of cancer deaths. Therefore, a major current goal is to develop treatment strategies that eliminate the resistant CSCs as well as the bulk tumor cells in order to achieve complete disease clearance. Aldehyde dehydrogenases (ALDHs) are important for maintenance and differentiation of stem cells as well as normal development. There is expanding evidence that ALDH expression increases in response to therapy and promotes chemoresistance and survival mechanisms in CSCs. This perspective will discuss a paper by Cojoc and colleagues recently published in Cancer Research, that indicates ALDHs play a key role in resistance to radiation therapy and tumor recurrence in prostate cancer. The authors suggest that ALDHs are a potential therapeutic target for treatment prostate cancer patients to limit radiation resistance and disease recurrence. The findings are consistent with work from other cancers showing ALDHs are major contributors of CSC signaling and resistance to anti-cancer treatments. This perspective will address representative work concerning the validity of ALDH and the associated retinoic acid signaling pathway as chemotherapeutic targets for prostate as well as other cancers. PMID:28149880

  1. Detoxification of promutagenic aldehydes derived from methylpyrenes by human aldehyde dehydrogenases ALDH2 and ALDH3A1.

    PubMed

    Glatt, Hansruedi; Rost, Katharina; Frank, Heinz; Seidel, Albrecht; Kollock, Ronny

    2008-09-15

    Methylated polycyclic aromatic hydrocarbons can be metabolically activated via benzylic hydroxylation and sulpho conjugation to reactive esters, which can induce mutations and tumours. Yet, further oxidation of the alcohol may compete with this toxification. We previously demonstrated that several human alcohol dehydrogenases (ADH1C, 2, 3 and 4) oxidise various benzylic alcohols (derived from alkylated pyrenes) to their aldehydes with high catalytic efficiency. However, all these ADHs also catalysed the reverse reaction, the reduction of the aldehydes to the alcohols, with comparable or higher efficiency. Thus, final detoxification requires elimination of the aldehydes by further biotransformation. We have expressed two human aldehyde dehydrogenases (ALDH2 and 3A1) in bacteria. All pyrene aldehydes studied (1-, 2- and 4-formylpyrene, 1-formyl-6-methylpyrene and 1-formyl-8-methylpyrene) were high-affinity substrates for ALDH2 (K(m)=0.027-0.9 microM) as well as ALDH3A1 (K(m)=0.78-11 microM). Catalytic efficiencies (k(cat)/K(m)) were higher for ALDH2 than ALDH3A1 by a moderate to a very large margin depending on the substrate. Most important, they were also substantially higher than the catalytic efficiencies of the various ADHs for the reduction the aldehydes to the alcohols. These kinetic properties ensure that ALDHs, and particularly ALDH2, can complete the ADH-mediated detoxification.

  2. Rotenone decreases intracellular aldehyde dehydrogenase activity: implications for the pathogenesis of Parkinson's disease.

    PubMed

    Goldstein, David S; Sullivan, Patti; Cooney, Adele; Jinsmaa, Yunden; Kopin, Irwin J; Sharabi, Yehonatan

    2015-04-01

    Repeated systemic administration of the mitochondrial complex I inhibitor rotenone produces a rodent model of Parkinson's disease (PD). Mechanisms of relatively selective rotenone-induced damage to nigrostriatal dopaminergic neurons remain incompletely understood. According to the 'catecholaldehyde hypothesis,' buildup of the autotoxic dopamine metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL) contributes to PD pathogenesis. Vesicular uptake blockade increases DOPAL levels, and DOPAL is detoxified mainly by aldehyde dehydrogenase (ALDH). We tested whether rotenone interferes with vesicular uptake and intracellular ALDH activity. Endogenous and F-labeled catechols were measured in PC12 cells incubated with rotenone (0-1000 nM, 180 min), without or with F-dopamine (2 μM) to track vesicular uptake and catecholamine metabolism. Rotenone dose dependently increased DOPAL, F-DOPAL, and 3,4-dihydroxyphenylethanol (DOPET) levels while decreasing dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) levels and the ratio of dopamine to the sum of its deaminated metabolites. In test tubes, rotenone did not affect conversion of DOPAL to DOPAC by ALDH when NAD(+) was supplied, whereas the direct-acting ALDH inhibitor benomyl markedly increased DOPAL and decreased DOPAC concentrations in the reaction mixtures. We propose that rotenone builds up intracellular DOPAL by decreasing ALDH activity and attenuating vesicular sequestration of cytoplasmic catecholamines. The results provide a novel mechanism for selective rotenone-induced toxicity in dopaminergic neurons. We report that rotenone, a mitochondrial complex I inhibitor that produces an animal model of Parkinson's disease, increases intracellular levels of the toxic dopamine metabolite 3,4-dihydroxyphenyl-acetaldehyde (DOPAL), via decreased DOPAL metabolism by aldehyde dehydrogenase (ALDH) and decreased vesicular sequestration of cytoplasmic dopamine by the vesicular monoamine transporter (VMAT). The results provide a novel

  3. Reduced aldehyde dehydrogenase expression in preeclamptic decidual mesenchymal stem/stromal cells is restored by aldehyde dehydrogenase agonists

    PubMed Central

    Kusuma, Gina D.; Abumaree, Mohamed H.; Perkins, Anthony V.; Brennecke, Shaun P.; Kalionis, Bill

    2017-01-01

    High resistance to oxidative stress is a common feature of mesenchymal stem/stromal cells (MSC) and is associated with higher cell survival and ability to respond to oxidative damage. Aldehyde dehydrogenase (ALDH) activity is a candidate “universal” marker for stem cells. ALDH expression was significantly lower in decidual MSC (DMSC) isolated from preeclamptic (PE) patients. ALDH gene knockdown by siRNA transfection was performed to create a cell culture model of the reduced ALDH expression detected in PE-DMSC. We showed that ALDH activity in DMSC is associated with resistance to hydrogen peroxide (H2O2)-induced toxicity. Our data provide evidence that ALDH expression in DMSC is required for cellular resistance to oxidative stress. Furthermore, candidate ALDH activators were screened and two of the compounds were effective in upregulating ALDH expression. This study provides a proof-of-principle that the restoration of ALDH activity in diseased MSC is a rational basis for a therapeutic strategy to improve MSC resistance to cytotoxic damage. PMID:28205523

  4. Characterization of five fatty aldehyde dehydrogenase enzymes from Marinobacter and Acinetobacter: structural insights into the aldehyde binding pocket.

    PubMed

    Bertram, Jonathan H; Mulliner, Kalene M; Shi, Ke; Plunkett, Mary H; Nixon, Peter; Serratore, Nicholas A; Douglas, Christopher J; Aihara, Hideki; Barney, Brett M

    2017-04-07

    Enzymes involved in lipid biosynthesis and metabolism play an important role in energy conversion and storage, and in the function of structural components such as cell membranes. The fatty aldehyde dehydrogenase (FAldDH) plays a central function in the metabolism of lipid intermediates, oxidizing fatty aldehydes to the corresponding fatty acid, and competing with pathways that would further reduce the fatty aldehydes to fatty alcohols or require the fatty aldehydes to produce alkanes. In this report, the genes for four putative FAldDH enzymes from Marinobacter aquaeolei VT8 and an additional enzyme from Acinetobacter baylyi were heterologously expressed in Escherichia coli and shown to display FAldDH activity. Five enzymes (Maqu_0438, Maqu_3316, Maqu_3410, Maqu_3572 and WP_004927398) were found to act on aldehydes ranging from acetaldehyde to hexadecanal, and also acted on the unsaturated long-chain palmitoleyl and oleyl aldehydes. A comparison of the specificity of these enzymes with various aldehydes is presented. Crystallization trials yielded diffraction quality crystals of one particular FAldDH (Maqu_3316) from M. aquaeolei VT8. Crystals were independently treated with both the NAD(+) cofactor and the aldehyde substrate decanal, revealing specific details of the likely substrate binding pocket for this class of enzymes. A likely model for how the catalysis by the enzyme is accomplished is also provided.Importance: This study provides a comparison of multiple enzymes with the ability to oxidize fatty aldehydes to fatty acids, and provides a likely picture of how the fatty aldehyde and NAD(+) is bound to the enzyme to facilitate catalysis. Based on the information obtained from this structural analysis and the comparisons of specificity for the five enzymes that were characterized, correlations may be drawn to the potential roles played by specific residues within the structure.

  5. Aldehyde dehydrogenase activity in Lactococcus chungangensis: Application in cream cheese to reduce aldehyde in alcohol metabolism.

    PubMed

    Konkit, Maytiya; Choi, Woo Jin; Kim, Wonyong

    2016-03-01

    Previous studies have shown that the metabolic capability of colonic microflora may be at least as high as that of the liver or higher than that of the whole human body. Aldehyde dehydrogenase (ALDH) is an enzyme produced by these bacteria that can metabolize acetaldehyde, produce from ethanol to acetate. Lactococcus species, which is commonly used as a starter in dairy products, was recently found to possess the ALDH gene, and the activity of this enzyme was determined. In this study, the ALDH activity of Lactococcus chungangensis CAU 28(T) and 11 other type strains in the genus Lactococcus was studied. Only 5 species, 3 of dairy origin (Lactococcus lactis ssp. lactis KCTC 3769(T), Lactococcus lactis ssp. cremoris KCCM 40699(T), and Lactococcus raffinolactis DSM 20443(T)) and 2 of nondairy origin (Lactococcus fujiensis NJ317(T) and L. chungangensis CAU 28(T)), showed ALDH activity and possessed a gene encoding ALDH. All of these strains were capable of making cream cheese. Among the strains, L. chungangensis produced cream cheese that contained the highest level of ALDH and was found to reduce the level of acetaldehyde in the serum of mice. These results predict a promising role for L. chungangensis CAU28(T) to be used in cheese that can be developed as functional food.

  6. Mitochondrial aldehyde dehydrogenase prevents ROS-induced vascular contraction in angiotensin-II hypertensive mice.

    PubMed

    Choi, Hyehun; Tostes, Rita C; Webb, R Clinton

    2011-01-01

    Mitochondrial aldehyde dehydrogenase (ALDH2) is an enzyme that detoxifies aldehydes to carboxylic acids. ALDH2 deficiency is known to increase oxidative stress, which is the imbalance between reactive oxygen species (ROS) generation and antioxidant defense activity. Increased ROS contribute to vascular dysfunction and structural remodeling in hypertension. We hypothesized that ALDH2 plays a protective role to reduce vascular contraction in angiotensin-II (AngII) hypertensive mice. Endothelium-denuded aortic rings from C57BL6 mice, treated with AngII (3.6 μg/kg/min, 14 days), were used to measure isometric force development. Rings treated with daidzin (10 μmol/L), an ALDH2 inhibitor, potentiated contractile responses to phenylephrine (PE) in AngII mice. Tempol (1 mmol/L) and catalase (600 U/mL) attenuated the augmented contractile effect of daidzin. In normotensive mice, contraction to PE in the presence of the daidzin was not different from control, untreated values. AngII aortic rings transfected with ALDH2 recombinant protein decreased contractile responses to PE compared with control. These data suggest that ALDH2 reduces vascular contraction in AngII hypertensive mice. Because tempol and catalase blocked the contractile response of the ALDH2 inhibitor, ROS generation by AngII may be decreased by ALDH2, thereby preventing ROS-induced contraction.

  7. Targeting Aldehyde Dehydrogenase Cancer Stem Cells in Ovarian Cancer

    PubMed Central

    Landen, Charles N.; Goodman, Blake; Katre, Ashwini A.; Steg, Adam D.; Nick, Alpa M.; Stone, Rebecca L.; Miller, Lance D.; Mejia, Pablo Vivas; Jennings, Nicolas B.; Gershenson, David M.; Bast, Robert C.; Coleman, Robert L.; Lopez-Berestein, Gabriel; Sood, Anil K.

    2010-01-01

    Aldehyde dehydrogenase-1A1 (ALDH1A1) expression characterizes a subpopulation of cells with tumor initiating or cancer stem cell properties in several malignancies. Our goal was to characterize the phenotype of ALDH1A1-positive ovarian cancer cells and examine the biological effects of ALDH1A1 gene silencing. In our analysis of multiple ovarian cancer cell lines, we found that ALDH1A1 expression and activity was significantly higher in taxane and platinum-resistant cell lines. In patient samples, 72.9% of ovarian cancers had ALDH1A1 expression, in whom the percent of ALDH1A1-positive cells correlated negatively with progression-free survival (6.05 v 13.81 months, p<0.035). Subpopulations of A2780cp20 cells with ALDH1A1 activity were isolated for orthotopic tumor initiating studies, where tumorigenicity was approximately 50-fold higher with ALDH1A1-positive cells. Interestingly, tumors derived from ALDH1A1-positive cells gave rise to both ALDH1A1-positive and ALDH1A1-negative populations, but ALDH1A1-negative cells could not generate ALDH1A1-positive cells. In an in vivo orthotopic mouse model of ovarian cancer, ALDH1A1 silencing using nanoliposomal siRNA sensitized both taxane- and platinum-resistant cell lines to chemotherapy, significantly reducing tumor growth in mice compared to chemotherapy alone (a 74–90% reduction, p<0.015). These data demonstrate that the ALDH1A1 subpopulation is associated with chemoresistance and outcome in ovarian cancer patients, and targeting ALDH1A1 sensitizes resistant cells to chemotherapy. ALDH1A1-positive cells have enhanced, but not absolute, tumorigenicity, but do have differentiation capacity lacking in ALDH1A1-negative cells. This enzyme may be important for identification and targeting of chemoresistant cell populations in ovarian cancer. PMID:20889728

  8. Bioactivation of pentaerythrityl tetranitrate by mitochondrial aldehyde dehydrogenase.

    PubMed

    Griesberger, Martina; Kollau, Alexander; Wölkart, Gerald; Wenzl, M Verena; Beretta, Matteo; Russwurm, Michael; Koesling, Doris; Schmidt, Kurt; Gorren, Antonius C F; Mayer, Bernd

    2011-03-01

    Mitochondrial aldehyde dehydrogenase (ALDH2) contributes to vascular bioactivation of the antianginal drugs nitroglycerin (GTN) and pentaerythrityl tetranitrate (PETN), resulting in cGMP-mediated vasodilation. Although continuous treatment with GTN results in the loss of efficacy that is presumably caused by inactivation of ALDH2, PETN does not induce vascular tolerance. To clarify the mechanisms underlying the distinct pharmacological profiles of GTN and PETN, bioactivation of the nitrates was studied with aortas isolated from ALDH2-deficient and nitrate-tolerant mice, isolated mitochondria, and purified ALDH2. Pharmacological inhibition or gene deletion of ALDH2 attenuated vasodilation to both GTN and PETN to virtually the same degree as long-term treatment with GTN, whereas treatment with PETN did not cause tolerance. Purified ALDH2 catalyzed bioactivation of PETN, assayed as activation of soluble guanylate cyclase (sGC) and formation of nitric oxide (NO). The EC(50) value of PETN for sGC activation was 2.2 ± 0.5 μM. Denitration of PETN to pentaerythrityl trinitrate was catalyzed by ALDH2 with a specific activity of 9.6 ± 0.8 nmol · min(-1) · mg(-1) and a very low apparent affinity of 94.7 ± 7.4 μM. In contrast to GTN, PETN did not cause significant inactivation of ALDH2. Our data suggest that ALDH2 catalyzes bioconversion of PETN in two distinct reactions. Besides the major denitration pathway, which occurs only at high PETN concentrations, a minor high-affinity pathway may reflect vascular bioactivation of the nitrate yielding NO. The very low rate of ALDH2 inactivation, presumably as a result of low affinity of the denitration pathway, may at least partially explain why PETN does not induce vascular tolerance.

  9. Residues that influence coenzyme preference in the aldehyde dehydrogenases.

    PubMed

    González-Segura, Lilian; Riveros-Rosas, Héctor; Julián-Sánchez, Adriana; Muñoz-Clares, Rosario A

    2015-06-05

    To find out the residues that influence the coenzyme preference of aldehyde dehydrogenases (ALDHs), we reviewed, analyzed and correlated data from their known crystal structures and amino-acid sequences with their published kinetic parameters for NAD(P)(+). We found that the conformation of the Rossmann-fold loops participating in binding the adenosine ribose is very conserved among ALDHs, so that coenzyme specificity is mainly determined by the nature of the residue at position 195 (human ALDH2 numbering). Enzymes with glutamate or proline at 195 prefer NAD(+) because the side-chains of these residues electrostatically and/or sterically repel the 2'-phosphate group of NADP(+). But contrary to the conformational rigidity of proline, the conformational flexibility of glutamate may allow NADP(+)-binding in some enzymes by moving the carboxyl group away from the 2'-phosphate group, which is possible if a small neutral residue is located at position 224, and favored if the residue at position 53 interacts with Glu195 in a NADP(+)-compatible conformation. Of the residues found at position 195, only glutamate interacts with the NAD(+)-adenosine ribose; glutamine and histidine cannot since their side-chain points are opposite to the ribose, probably because the absence of the electrostatic attraction by the conserved nearby Lys192, or its electrostatic repulsion, respectively. The shorter side-chains of other residues-aspartate, serine, threonine, alanine, valine, leucine, or isoleucine-are distant from the ribose but leave room for binding the 2'-phosphate group. Generally, enzymes having a residue different from Glu bind NAD(+) with less affinity, but they can also bind NADP(+) even sometimes with higher affinity than NAD(+), as do enzymes containing Thr/Ser/Gln195. Coenzyme preference is a variable feature within many ALDH families, consistent with being mainly dependent on a single residue that apparently has no other structural or functional roles, and therefore can

  10. Rutin attenuates ethanol-induced neurotoxicity in hippocampal neuronal cells by increasing aldehyde dehydrogenase 2.

    PubMed

    Song, Kibbeum; Kim, Sokho; Na, Ji-Young; Park, Jong-Heum; Kim, Jae-Kyung; Kim, Jae-Hun; Kwon, Jungkee

    2014-10-01

    Rutin is derived from buckwheat, apples, and black tea. It has been shown to have beneficial anti-inflammatory and antioxidant effects. Ethanol is a central nervous system depressant and neurotoxin. Its metabolite, acetaldehyde, is critically toxic. Aldehyde dehydrogenase 2 (ALDH2) metabolizes acetaldehyde into nontoxic acetate. This study examined rutin's effects on ALDH2 activity in hippocampal neuronal cells (HT22 cells). Rutin's protective effects against acetaldehyde-based ethanol neurotoxicity were confirmed. Daidzin, an ALDH2 inhibitor, was used to clarify the mechanisms of rutin's protective effects. Cell viability was significantly increased after rutin treatment. Rutin significantly reversed ethanol-increased Bax, cytochrome c expression and caspase 3 activity, and decreased Bcl-2 and Bcl-xL protein expression in HT22 cells. Interestingly, rutin increased ALDH2 expression, while daidzin reversed this beneficial effect. Thus, this study demonstrates rutin protects HT22 cells against ethanol-induced neurotoxicity by increasing ALDH2 activity.

  11. Isolation of primitive human hematopoietic progenitors on the basis of aldehyde dehydrogenase activity

    PubMed Central

    Storms, Robert W.; Trujillo, Aliana P.; Springer, James B.; Shah, Lisa; Colvin, O. Michael; Ludeman, Susan M.; Smith, Clay

    1999-01-01

    Because hematopoietic stem cells are rich in aldehyde dehydrogenase (ALDH) activity, we developed a fluorescent substrate for ALDH, termed BODIPY aminoacetaldehyde (BAAA), and tested its potential for isolating primitive human hematopoietic cells. A population of cells with low orthogonal light scattering and bright fluorescence intensity (SSCloALDHbr cells) could be readily fractionated from human umbilical cord blood cells costained with BAAA and the multidrug-resistance inhibitor verapamil. The SSCloALDHbr population was depleted of lineage-committed cells, 40–90% pure for CD34+CD38lo/− cells, and enriched 50- to 100-fold for primitive hematopoietic progenitors detected in short- and long-term culture analyses. Together, these observations indicate that fractionating human hematopoietic stem cells on the basis of ALDH activity using BAAA is an effective method for isolating primitive human hematopoietic progenitors. This technique may be useful for isolating stem cells from other tissues as well. PMID:10430905

  12. Aldehyde dehydrogenase inhibition blocks mucosal fibrosis in human and mouse ocular scarring

    PubMed Central

    Ahadome, Sarah D.; Abraham, David J.; Rayapureddi, Suryanarayana; Saw, Valerie P.; Saban, Daniel R.; Calder, Virginia L.; Norman, Jill T.; Ponticos, Markella; Daniels, Julie T.; Dart, John K.

    2016-01-01

    Mucous membrane pemphigoid (MMP) is a systemic mucosal scarring disease, commonly causing blindness, for which there is no antifibrotic therapy. Aldehyde dehydrogenase family 1 (ALDH1) is upregulated in both ocular MMP (OMMP) conjunctiva and cultured fibroblasts. Application of the ALDH metabolite, retinoic acid (RA), to normal human conjunctival fibroblasts in vitro induced a diseased phenotype. Conversely, application of ALDH inhibitors, including disulfiram, to OMMP fibroblasts in vitro restored their functionality to that of normal controls. ALDH1 is also upregulated in the mucosa of the mouse model of scarring allergic eye disease (AED), used here as a surrogate for OMMP, in which topical application of disulfiram decreased fibrosis in vivo. These data suggest that progressive scarring in OMMP results from ALDH/RA fibroblast autoregulation, that the ALDH1 subfamily has a central role in immune-mediated ocular mucosal scarring, and that ALDH inhibition with disulfiram is a potential and readily translatable antifibrotic therapy. PMID:27699226

  13. The Genetics of Alcohol Metabolism: Role of Alcohol Dehydrogenase and Aldehyde Dehydrogenase Variants

    PubMed Central

    Edenberg, Howard J.

    2007-01-01

    The primary enzymes involved in alcohol metabolism are alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH). Both enzymes occur in several forms that are encoded by different genes; moreover, there are variants (i.e., alleles) of some of these genes that encode enzymes with different characteristics and which have different ethnic distributions. Which ADH or ALDH alleles a person carries influence his or her level of alcohol consumption and risk of alcoholism. Researchers to date primarily have studied coding variants in the ADH1B, ADH1C, and ALDH2 genes that are associated with altered kinetic properties of the resulting enzymes. For example, certain ADH1B and ADH1C alleles encode particularly active ADH enzymes, resulting in more rapid conversion of alcohol (i.e., ethanol) to acetaldehyde; these alleles have a protective effect on the risk of alcoholism. A variant of the ALDH2 gene encodes an essentially inactive ALDH enzyme, resulting in acetaldehyde accumulation and a protective effect. It is becoming clear that noncoding variants in both ADH and ALDH genes also may influence alcohol metabolism and, consequently, alcoholism risk; the specific nature and effects of these variants still need further study. PMID:17718394

  14. Identification of aldehyde dehydrogenase 1A1 modulators using virtual screening.

    PubMed

    Kotraiah, Vinayaka; Pallares, Diego; Toema, Deanna; Kong, Dehe; Beausoleil, Eric

    2013-06-01

    The highly similar aldehyde dehydrogenase isozymes (ALDH1A1 and ALDH2) have been implicated in the metabolism of toxic biogenic aldehydes such as 3,4-dihydroxyphenylacetaldehyde (DOPAL) and 4-hydroxy-2E-nonenal. We report the down-regulation of ALDH1A1 mRNA found in substantia nigra tissue of human Parkinson's disease (PD) samples using the Genome-Wide SpliceArray(™) (GWSA(™)) technology. Since DOPAL can rapidly inactivate ALDH1A1 in vitro, we set up a DOPAL-induced ALDH1A1 inactivation assay and used this assay to demonstrate that Alda-1, a compound originally identified as an activator of ALDH2, can also activate ALDH1A1. We carried out a virtual screening of 19,943 compounds and the top 21 hits from this screen were tested in the DOPAL inactivation assay with ALDH1A1 which led to identification of an activator as well as two inhibitors among these hits. These findings represent an attractive starting point for developing higher potency activator compounds that may have utility in restoring the metabolism of DOPAL in PD.

  15. [Alcohol dehydrogenase and aldehyde dehydrogenase as tumour markers and factors intensifying carcinogenesis in colorectal cancer].

    PubMed

    Jelski, Wojciech; Orywal, Karolina; Kedra, Bogusław; Szmitkowski, Maciej

    2008-06-01

    Numerous experiments have shown that alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are present in cells of various cancers and play role in carcinogenesis. The aim of this study was to compare the capacity for ethanol metabolism measured by ADH isoenzymes and ALDH activity, between colorectal cancer and normal colonic mucosa. We have also investigated the serum activity of these enzymes in colorectal cancer patients as potential tumour markers. The activities of ADH isoenzymes and ALDH were measured in the: cancer tissue, healthy colonic mucosa and serum of 42 patients with colorectal cancer. For the measurement of the activity of class I ADH isoenzyme and ALDH activity the fluorometric methods was employed. The total ADH activity and activity of class III and IV isoenzymes was measured by the photometric method. The activity of total alcohol dehydrogenase and class I of ADH were significantly higher in cancer cells than in healthy tissues. The other tested classes of ADH had higher activities in cancer tissue but the differences were not statistically significant. The activity of ALDH was significantly lower in the cancer cells. The activities of all tested enzymes and isoenzymes in colorectal cancer tissue were not significantly higher in drinkers than in non-drinkers. Additionally we observed statistically significant increasing activity of class I ADH isoenzymes in the sera of patients with colorectal cancer. For this reason the total ADH activity was also significantly increased. The activities of ADH III and ADH IV isoenzymes and ALDH were unchanged in the sera of patients. There were no marked differences in activities of all tested enzymes and isoenzymes between drinkers and non-drinkers (with colorectal cancer). The differences in activities of total ADH and class I ADH isoenzymes between colorectal cancer tissues and healthy mucosa might be a factor of ethanol metabolism disorders, which can intensify carcinogenesis. The increased total

  16. GRE2 from Scheffersomyces stipitis as an aldehyde reductase contributes tolerance to aldehyde inhibitors derived from lignocellulosic biomass.

    PubMed

    Wang, Xu; Ma, Menggen; Liu, Z Lewis; Xiang, Quanju; Li, Xi; Liu, Na; Zhang, Xiaoping

    2016-08-01

    Scheffersomyces (Pichia) stipitis is one of the most promising yeasts for industrial bioethanol production from lignocellulosic biomass. S. stipitis is able to in situ detoxify aldehyde inhibitors (such as furfural and 5-hydroxymethylfurfural (HMF)) to less toxic corresponding alcohols. However, the reduction enzymes involved in this reaction remain largely unknown. In this study, we reported that an uncharacterized open reading frame PICST_72153 (putative GRE2) from S. stipitis was highly induced in response to furfural and HMF stresses. Overexpression of this gene in Saccharomyces cerevisiae improved yeast tolerance to furfural and HMF. GRE2 was identified as an aldehyde reductase which can reduce furfural to FM with either NADH or NADPH as the co-factor and reduce HMF to FDM with NADPH as the co-factor. This enzyme can also reduce multiple aldehydes to their corresponding alcohols. Amino acid sequence analysis indicated that it is a member of the subclass "intermediate" of the short-chain dehydrogenase/reductase (SDR) superfamily. Although GRE2 from S. stipitis is similar to GRE2 from S. cerevisiae in a three-dimensional structure, some differences were predicted. GRE2 from S. stipitis forms loops at D133-E137 and T143-N145 locations with two α-helices at E154-K157 and E252-A254 locations, different GRE2 from S. cerevisiae with an α-helix at D133-E137 and a β-sheet at T143-N145 locations, and two loops at E154-K157 and E252-A254 locations. This research provided guidelines for the study of other SDR enzymes from S. stipitis and other yeasts on tolerant mechanisms to aldehyde inhibitors derived from lignocellulosic biomass.

  17. Enhancement of furan aldehydes conversion in Zymomonas mobilis by elevating dehydrogenase activity and cofactor regeneration.

    PubMed

    Wang, Xia; Gao, Qiuqiang; Bao, Jie

    2017-01-01

    Furfural and 5-hydroxymethylfurfural (HMF) are the two major furan aldehyde inhibitors generated from lignocellulose dilute acid pretreatment which significantly inhibit subsequent microbial cell growth and ethanol fermentation. Zymomonas mobilis is an important strain for cellulosic ethanol fermentation but can be severely inhibited by furfural and (or) HMF. Previous study showed that Z. mobilis contains its native oxidoreductases to catalyze the conversion of furfural and HMF, but the corresponding genes have not been identified. This study identified a NADPH-dependent alcohol dehydrogenase gene ZMO1771 from Z. mobilis ZM4, which is responsible for the efficient reduction of furfural and HMF. Over-expression of ZMO1771 in Z. mobilis significantly increased the conversion rate to both furfural and HMF and resulted in an accelerated cell growth and improved ethanol productivity in corn stover hydrolysate. Further, the ethanol fermentation performance was enhanced again by co-expression of the transhydrogenase gene udhA with ZMO1771 by elevating the NADPH availability. A genetically modified Z. mobilis by co-expressing alcohol dehydrogenase gene ZMO1771 with transhydrogenase gene udhA showed enhanced conversion rate of furfural and HMF and accelerated ethanol fermentability from lignocellulosic hydrolysate. The results presented in this study provide an important method on constructing robust strains for efficient ethanol fermentation from lignocellulose feedstock.

  18. Pharmacological activation of aldehyde dehydrogenase 2 by Alda-1 reverses alcohol-induced hepatic steatosis and cell death in mice.

    PubMed

    Zhong, Wei; Zhang, Wenliang; Li, Qiong; Xie, Guoxiang; Sun, Qian; Sun, Xiuhua; Tan, Xiaobing; Sun, Xinguo; Jia, Wei; Zhou, Zhanxiang

    2015-06-01

    Effective therapies for alcoholic liver disease are currently unavailable. The present study tested the efficacy of Alda-1, a specific aldehyde dehydrogenase 2 (ALDH2) activator, in treating alcoholic liver disease. Male C57BL/6J mice were exposed to alcohol for a time-course study on aldehyde metabolism. The specificity and efficacy of Alda-1 on activating hepatic ALDH2 and aldehyde clearance were determined by acute treatments. Then, mice were fed alcohol for 8 weeks with Alda-1 administration for the last 10 days to test the therapeutic potential of Alda-1. Lastly, H4IIEC3 cells were treated with ethanol, acetaldehyde, or 4-hydroxynonenal to define the link between aldehydes and hepatotoxicity. Alcohol feeding for 8 weeks induced hepatic ALDH2 dysfunction and aldehyde accumulation. One dose of Alda-1 administration elevated hepatic ALDH activity, which was blocked by the specific ALDH2 inhibitor, daidzin. Alda-1 accelerated acetaldehyde clearance after acute alcohol intoxication. Alda-1 treatment in the 8-week alcohol feeding model reversed liver damage along with reduction of hepatic aldehydes. Alda-1 re-activated transcription factors, upregulated fatty acid oxidation enzymes, and reversed steatosis. Alcohol-induced endoplasmic reticulum stress and apoptotic cell death were also attenuated by Alda-1. Acetaldehyde or 4-hydroxynonenal treatment to H4IIEC3 cells inactivated transcription factors and induced endoplasmic reticulum stress and apoptosis, while ethanol per se showed limited effects. Pharmacological activation of ALDH2 by Alda-1 reversed alcoholic steatosis and apoptosis through accelerating aldehyde clearance. This study indicates that ALDH2 is a promising molecular target and Alda-1 has therapeutic potential for treating alcoholic liver disease. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

  19. Aldehydic load and aldehyde dehydrogenase 2 profile during the progression of post-myocardial infarction cardiomyopathy: benefits of Alda-1

    PubMed Central

    Gomes, Katia M.S.; Bechara, Luiz R.G.; Lima, Vanessa M.; Ribeiro, Márcio A.C.; Campos, Juliane C.; Dourado, Paulo M.; Kowaltowski, Alicia J.; Mochly-Rosen, Daria; Ferreira, Julio C.B.

    2015-01-01

    Background/Objectives We previously demonstrated that reducing cardiac aldehydic load by aldehyde dehydrogenase 2 (ALDH2), a mitochondrial enzyme responsible for metabolizing the major lipid peroxidation product, protects against acute ischemia/reperfusion injury and chronic heart failure. However, time-dependent changes in ALDH2 profile, aldehydic load and mitochondrial bioenergetics during progression of post-myocardial infarction (post-MI) cardiomyopathy is unknown and should be established to determine the optimal time window for drug treatment. Methods Here we characterized cardiac ALDH2 activity and expression, lipid peroxidation, 4-hydroxy-2-nonenal (4-HNE) adduct formation, glutathione pool and mitochondrial energy metabolism and H2O2 release during the 4 weeks after permanent left anterior descending (LAD) coronary artery occlusion in rats. Results We observed a sustained disruption of cardiac mitochondrial function during the progression of post-MI cardiomyopathy, characterized by >50% reduced mitochondrial respiratory control ratios and up to 2 fold increase in H2O2 release. Mitochondrial dysfunction was accompanied by accumulation of cardiac and circulating lipid peroxides and 4-HNE protein adducts and down-regulation of electron transport chain complexes I and V. Moreover, increased aldehydic load was associated with a 90% reduction in cardiac ALDH2 activity and increased glutathione pool. Further supporting an ALDH2 mechanism, sustained Alda-1 treatment (starting 24hrs after permanent LAD occlusion surgery) prevented aldehydic overload, mitochondrial dysfunction and improved ventricular function in post-MI cardiomyopathy rats. Conclusion Taken together, our findings demonstrate a disrupted mitochondrial metabolism along with an insufficient cardiac ALDH2-mediated aldehyde clearance during the progression of ventricular dysfunction, suggesting a potential therapeutic value of ALDH2 activators during the progression of post-myocardial infarction

  20. Daidzin inhibits mitochondrial aldehyde dehydrogenase and suppresses ethanol intake of Syrian golden hamsters.

    PubMed

    Keung, W M; Klyosov, A A; Vallee, B L

    1997-03-04

    Daidzin is the major active principle in extracts of radix puerariae, a traditional Chinese medication that suppresses the ethanol intake of Syrian golden hamsters. It is the first isoflavone recognized to have this effect. Daidzin is also a potent and selective inhibitor of human mitochondrial aldehyde dehydrogenase (ALDH-2). To establish a link between these two activities, we have tested a series of synthetic structural analogs of daidzin. The results demonstrate a direct correlation between ALDH-2 inhibition and ethanol intake suppression and raise the possibility that daidzin may, in fact, suppress ethanol intake of golden hamsters by inhibiting ALDH-2. Hamster liver contains not only mitochondrial ALDH-2 but also high concentrations of a cytosolic form, ALDH-1, which is a very efficient catalyst of acetaldehyde oxidation. Further, the cytosolic isozyme is completely resistant to daidzin inhibition. This unusual property of the hamster ALDH-1 isozyme accounts for the fact we previously observed that daidzin can suppress ethanol intake of this species without blocking acetaldehyde metabolism. Thus, the mechanism by which daidzin suppresses ethanol intake in golden hamsters clearly differs from that proposed for the classic ALDH inhibitor disulfiram. We postulate that a physiological pathway catalyzed by ALDH-2, so far undefined, controls ethanol intake of golden hamsters and mediates the antidipsotropic effect of daidzin.

  1. Rotenone Decreases Intracellular Aldehyde Dehydrogenase Activity: Implications for the Pathogenesis of Parkinson Disease

    PubMed Central

    Goldstein, David S.; Sullivan, Patti; Cooney, Adele; Jinsmaa, Yunden; Kopin, Irwin J.; Sharabi, Yehonatan

    2015-01-01

    Repeated systemic administration of the mitochondrial complex I inhibitor rotenone produces a rodent model of Parkinson disease (PD). Mechanisms of relatively selective rotenone-induced damage to nigrostriatal dopaminergic neurons remain incompletely understood. According to the “catecholaldehyde hypothesis,” buildup of the autotoxic dopamine metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL) contributes to PD pathogenesis. Vesicular uptake blockade increases DOPAL levels, and DOPAL is detoxified mainly by aldehyde dehydrogenase (ALDH). We tested whether rotenone interferes with vesicular uptake and intracellular ALDH activity. Endogenous and F-labeled catechols were measured in PC12 cells incubated with rotenone (0-1000 nM, 180 minutes), without or with F-dopamine (2 μM) to track vesicular uptake and catecholamine metabolism. Rotenone dose-dependently increased DOPAL, F-DOPAL, and 3,4-dihydroxyphenylethanol (DOPET) levels while decreasing dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) levels and the ratio of dopamine to the sum of its deaminated metabolites. In test tubes, rotenone did not affect conversion of DOPAL to DOPAC by ALDH when NAD+ was supplied, whereas the direct-acting ALDH inhibitor benomyl markedly increased DOPAL and decreased DOPAC concentrations in the reaction mixtures. We propose that rotenone builds up intracellular DOPAL by decreasing ALDH activity and attenuating vesicular sequestration of cytoplasmic catecholamines. The results provide a novel mechanism for selective rotenone-induced toxicity in dopaminergic neurons. PMID:25645689

  2. Molecular characterization of an aldehyde/alcohol dehydrogenase gene from Clostridium acetobutylicum ATCC 824.

    PubMed Central

    Nair, R V; Bennett, G N; Papoutsakis, E T

    1994-01-01

    A gene (aad) coding for an aldehyde/alcohol dehydrogenase (AAD) was identified immediately upstream of the previously cloned ctfA (J. W. Cary, D. J. Petersen, E. T. Papoutsakis, and G. N. Bennett, Appl. Environ. Microbiol. 56:1576-1583, 1990) of Clostridium acetobutylicum ATCC 824 and sequenced. The 2,619-bp aad codes for a 96,517-Da protein. Primer extension analysis identified two transcriptional start sites 83 and 243 bp upstream of the aad start codon. The N-terminal section of AAD shows homology to aldehyde dehydrogenases of bacterial, fungal, mammalian, and plant origin, while the C-terminal section shows homology to alcohol dehydrogenases of bacterial (which includes three clostridial alcohol dehydrogenases) and yeast origin. AAD exhibits considerable amino acid homology (56% identity) over its entire sequence to the trifunctional protein encoded by adhE from Escherichia coli. Expression of aad from a plasmid in C. acetobutylicum showed that AAD, which appears as a approximately 96-kDa band in denaturing protein gels, provides elevated activities of NADH-dependent butanol dehydrogenase, NAD-dependent acetaldehyde dehydrogenase and butyraldehyde dehydrogenase, and a small increase in NADH-dependent ethanol dehydrogenase. A 957-bp open reading frame that could potentially encode a 36,704-Da protein was identified upstream of aad. Images PMID:8300540

  3. Metabolic basis of ethylene glycol monobutyl ether (2-butoxyethanol) toxicity: role of alcohol and aldehyde dehydrogenases

    SciTech Connect

    Ghanayem, B.I.; Burka, L.T.; Matthews, H.B.

    1987-07-01

    2-Butoxyethanol (BE) is a massively produced glycol ether of which more than 230 million pounds was produced in the United States in 1983. It is extensively used in aerosols and cleaning agents intended for household use. This creates a high potential for human exposure during its manufacturing and use. A single exposure of rats to BE causes severe hemolytic anemia accompanied by secondary hemoglobinuria as well as liver and kidney damage. Butoxyacetic acid (BAA) was earlier identified as a urinary metabolite of BE. In addition, we have recently identified two additional urinary metabolites of BE, namely, BE-glucuronide and BE-sulfate conjugates. The current studies were undertaken to investigate the metabolic basis of BE-induced hematotoxicity in male F344 rats. Treatment of rats with pyrazole (alcohol dehydrogenase inhibitor) protected rats against BE-induced hematotoxicity and inhibited BE metabolism to BAA. Pyrazole inhibition of BE metabolism to BAA was accompanied by increased BE metabolism to BE-glucuronide and BE-sulfate as determined by quantitative high-performance liquid chromatography analysis of BE metabolites in urine. There was approximately a 10-fold decrease in the ratio of BAA to BE-glucuronide + BE-sulfate in the urine of rats treated with pyrazole + BE compared to rats treated with BE alone. Pretreatment of rats with cyanamide (aldehyde dehydrogenase inhibitor) also significantly protected rats against BE-induced hematotoxicity and modified BE metabolism in a manner similar to that caused by pyrazole. Administration of equimolar doses of BE, the metabolic intermediate butoxyacetaldehyde, or the ultimate metabolite BAA caused similar hematotoxic effects. Cyanamide also protected rats against butoxyacetaldehyde-induced hematotoxicity.

  4. Oxidative stress and mitochondrial aldehyde dehydrogenase activity: a comparison of pentaerythritol tetranitrate with other organic nitrates.

    PubMed

    Daiber, Andreas; Oelze, Matthias; Coldewey, Meike; Bachschmid, Markus; Wenzel, Philip; Sydow, Karsten; Wendt, Maria; Kleschyov, Andrei L; Stalleicken, Dirk; Ullrich, Volker; Mülsch, Alexander; Münzel, Thomas

    2004-12-01

    Mitochondrial aldehyde dehydrogenase (ALDH-2) was recently identified to be essential for the bioactivation of glyceryl trinitrate (GTN). Here we assessed whether other organic nitrates are bioactivated by a similar mechanism. The ALDH-2 inhibitor benomyl reduced the vasodilator potency, but not the efficacy, of GTN, pentaerythritol tetranitrate (PETN), and pentaerythritol trinitrate in phenylephrine-constricted rat aorta, whereas vasodilator responses to isosorbide dinitrate, isosorbide-5-mononitrate, pentaerythritol dinitrate, pentaerythritol mononitrate, and the endothelium-dependent vasodilator acetylcholine were not affected. Likewise, benomyl decreased GTN- and PETN-elicited phosphorylation of the cGMP-activated protein kinase substrate vasodilator-stimulated phosphoprotein (VASP) but not that elicited by other nitrates. The vasodilator potency of organic nitrates correlated with their potency to inhibit ALDH-2 dehydrogenase activity in mitochondria from rat heart and increase mitochondrial superoxide formation, as detected by chemiluminescence. In contrast, mitochondrial ALDH-2 esterase activity was not affected by PETN and its metabolites, whereas it was inhibited by benomyl, GTN applied in vitro and in vivo, and some sulfhydryl oxidants. The bioactivation-related metabolism of GTN to glyceryl-1,2-dinitrate by isolated RAW macrophages was reduced by the ALDH-2 inhibitors benomyl and daidzin, as well as by GTN at concentrations >1 microM. We conclude that mitochondrial ALDH-2, specifically its esterase activity, is required for the bioactivation of the organic nitrates with high vasodilator potency, such as GTN and PETN, but not for the less potent nitrates. It is interesting that ALDH-2 esterase activity was inhibited by GTN only, not by the other nitrates tested. This difference might explain why GTN elicits mitochondrial superoxide formation and nitrate tolerance with the highest potency.

  5. Expression of a heat-stable NADPH-dependent alcohol dehydrogenase in Caldicellulosiruptor bescii results in furan aldehyde detoxification.

    PubMed

    Chung, Daehwan; Verbeke, Tobin J; Cross, Karissa L; Westpheling, Janet; Elkins, James G

    2015-01-01

    Compounds such as furfural and 5-hydroxymethylfurfural (5-HMF) are generated through the dehydration of xylose and glucose, respectively, during dilute-acid pretreatment of lignocellulosic biomass and are also potent microbial growth and fermentation inhibitors. The enzymatic reduction of these furan aldehydes to their corresponding, and less toxic, alcohols is an engineering approach that has been successfully implemented in both Saccharomyces cerevisiae and ethanologenic Escherichia coli, but has not yet been investigated in thermophiles relevant to biofuel production through consolidated bioprocessing (CBP). Developing CBP-relevant biocatalysts that are either naturally resistant to such inhibitors, or are amenable to engineered resistance, is therefore, an important component in making biofuels production from lignocellulosic biomass feasible. A butanol dehydrogenase encoding gene from Thermoanaerobacter pseudethanolicus 39E (Teth39_1597), previously shown to have furfural and 5-HMF reducing capabilities, was cloned into a suicide plasmid, pDCW171 and transformed into a lactate dehydrogenase mutant of Caldicellulosiruptor bescii. Integration of the gene into the C. bescii chromosome was verified via PCR amplification and stable expression was observed up to 75°C. Heterologous expression of the NADPH-dependent BdhA enzyme conferred increased resistance of the engineered strain to both furfural and 5-HMF relative to the wild-type and parental strains. Further, when challenged with 15 mM concentrations of either furan aldehyde, the ability to eliminate furfural or 5-HMF from the culture medium was significantly improved in the engineered strain. A genetically engineered strain of C. bescii (JWCB044) has been constructed that shows both an improved tolerance to furan aldehydes and an improved ability to eliminate furfural and 5-HMF from the culture medium. The work presented here represents the first example of engineering furan aldehyde resistance into a CBP

  6. Mitochondrial aldehyde dehydrogenase obliterates insulin resistance-induced cardiac dysfunction through deacetylation of PGC-1α

    PubMed Central

    Hu, Nan; Ren, Jun; Zhang, Yingmei

    2016-01-01

    Insulin resistance contributes to the high prevalence of type 2 diabetes mellitus, leading to cardiac anomalies. Emerging evidence depicts a pivotal role for mitochondrial injury in oxidative metabolism and insulin resistance. Mitochondrial aldehyde dehydrogenase (ALDH2) is one of metabolic enzymes detoxifying aldehydes although its role in insulin resistance remains elusive. This study was designed to evaluate the impact of ALDH2 overexpression on insulin resistance-induced myocardial damage and mechanisms involved with a focus on autophagy. Wild-type (WT) and transgenic mice overexpressing ALDH2 were fed sucrose or starch diet for 8 weeks and cardiac function and intracellular Ca2+ handling were assessed using echocardiographic and IonOptix systems. Western blot analysis was used to evaluate Akt, heme oxygenase-1 (HO-1), PGC-1α and Sirt-3. Our data revealed that sucrose intake provoked insulin resistance and compromised fractional shortening, cardiomyocyte function and intracellular Ca2+ handling (p < 0.05) along with unaltered cardiomyocyte size (p > 0.05), mitochondrial injury (elevated ROS generation, suppressed NAD+ and aconitase activity, p < 0.05 for all), the effect of which was ablated by ALDH2. In vitro incubation of the ALDH2 activator Alda-1, the Sirt3 activator oroxylin A and the histone acetyltransferase inhibitor CPTH2 rescued insulin resistance-induced changes in aconitase activity and cardiomyocyte function (p < 0.05). Inhibiting Sirt3 deacetylase using 5-amino-2-(4-aminophenyl) benzoxazole negated Alda-1-induced cardioprotective effects. Taken together, our data suggest that ALDH2 serves as an indispensable cardioprotective factor against insulin resistance-induced cardiomyopathy with a mechanism possibly associated with facilitation of the Sirt3-dependent PGC-1α deacetylation. PMID:27634872

  7. Expression of aldehyde dehydrogenase 6 reduces inhibitory effect of furan derivatives on cell growth and ethanol production in Saccharomyces cerevisiae.

    PubMed

    Park, Seong-Eon; Koo, Hyun Min; Park, Young Kyoung; Park, Sung Min; Park, Jae Chan; Lee, Oh-Kyu; Park, Yong-Cheol; Seo, Jin-Ho

    2011-05-01

    Yeast dehydrogenases and reductases were overexpressed in Saccharomyces cerevisiae D452-2 to detoxify 2-furaldehyde (furfural) and 5-hydroxymethyl furaldehyde (HMF), two potent toxic chemicals present in acid-hydrolyzed cellulosic biomass, and hence improve cell growth and ethanol production. Among those enzymes, aldehyde dehydrogenase 6 (ALD6) played the dual roles of direct oxidation of furan derivatives and supply of NADPH cofactor to their reduction reactions. Batch fermentation of S. cerevisiae D452-2/pH-ALD6 in the presence of 2g/L furfural and 0.5 g/L HMF resulted in 20-30% increases in specific growth rate, ethanol concentration and ethanol productivity, compared with those of the wild type strain. It was proposed that overexpression of ALD6 could recover the yeast cell metabolism and hence increase ethanol production from lignocellulosic biomass containing furan-derived inhibitors. Copyright © 2011 Elsevier Ltd. All rights reserved.

  8. Neurodegeneration and motor dysfunction in mice lacking cytosolic and mitochondrial aldehyde dehydrogenases: implications for Parkinson's disease.

    PubMed

    Wey, Margaret Chia-Ying; Fernandez, Elizabeth; Martinez, Paul Anthony; Sullivan, Patricia; Goldstein, David S; Strong, Randy

    2012-01-01

    Previous studies have reported elevated levels of biogenic aldehydes in the brains of patients with Parkinson's disease (PD). In the brain, aldehydes are primarily detoxified by aldehyde dehydrogenases (ALDH). Reduced ALDH1 expression in surviving midbrain dopamine neurons has been reported in brains of patients who died with PD. In addition, impaired complex I activity, which is well documented in PD, reduces the availability of the NAD(+) co-factor required by multiple ALDH isoforms to catalyze the removal of biogenic aldehydes. We hypothesized that chronically decreased function of multiple aldehyde dehydrogenases consequent to exposure to environmental toxins and/or reduced ALDH expression, plays an important role in the pathophysiology of PD. To address this hypothesis, we generated mice null for Aldh1a1 and Aldh2, the two isoforms known to be expressed in substantia nigra dopamine neurons. Aldh1a1(-/-)×Aldh2(-/-) mice exhibited age-dependent deficits in motor performance assessed by gait analysis and by performance on an accelerating rotarod. Intraperitoneal administration of L-DOPA plus benserazide alleviated the deficits in motor performance. We observed a significant loss of neurons immunoreactive for tyrosine hydroxylase (TH) in the substantia nigra and a reduction of dopamine and metabolites in the striatum of Aldh1a1(-/-)×Aldh2(-/-) mice. We also observed significant increases in biogenic aldehydes reported to be neurotoxic, including 4-hydroxynonenal (4-HNE) and the aldehyde intermediate of dopamine metabolism, 3,4-dihydroxyphenylacetaldehyde (DOPAL). These results support the hypothesis that impaired detoxification of biogenic aldehydes may be important in the pathophysiology of PD and suggest that Aldh1a1(-/-)×Aldh2(-/-) mice may be a useful animal model of PD.

  9. Crystal structure of human aldehyde dehydrogenase 1A3 complexed with NAD+ and retinoic acid

    PubMed Central

    Moretti, Andrea; Li, Jianfeng; Donini, Stefano; Sobol, Robert W.; Rizzi, Menico; Garavaglia, Silvia

    2016-01-01

    The aldehyde dehydrogenase family 1 member A3 (ALDH1A3) catalyzes the oxidation of retinal to the pleiotropic factor retinoic acid using NAD+. The level of ALDHs enzymatic activity has been used as a cancer stem cell marker and seems to correlate with tumour aggressiveness. Elevated ALDH1A3 expression in mesenchymal glioma stem cells highlights the potential of this isozyme as a prognosis marker and drug target. Here we report the first crystal structure of human ALDH1A3 complexed with NAD+ and the product all-trans retinoic acid (REA). The tetrameric ALDH1A3 folds into a three domain-based architecture highly conserved along the ALDHs family. The structural analysis revealed two different and coupled conformations for NAD+ and REA that we propose to represent two snapshots along the catalytic cycle. Indeed, the isoprenic moiety of REA points either toward the active site cysteine, or moves away adopting the product release conformation. Although ALDH1A3 shares high sequence identity with other members of the ALDH1A family, our structural analysis revealed few peculiar residues in the 1A3 isozyme active site. Our data provide information into the ALDH1As catalytic process and can be used for the structure-based design of selective inhibitors of potential medical interest. PMID:27759097

  10. Structural and functional analysis of betaine aldehyde dehydrogenase from Staphylococcus aureus.

    PubMed

    Halavaty, Andrei S; Rich, Rebecca L; Chen, Chao; Joo, Jeong Chan; Minasov, George; Dubrovska, Ievgeniia; Winsor, James R; Myszka, David G; Duban, Mark; Shuvalova, Ludmilla; Yakunin, Alexander F; Anderson, Wayne F

    2015-05-01

    When exposed to high osmolarity, methicillin-resistant Staphylococcus aureus (MRSA) restores its growth and establishes a new steady state by accumulating the osmoprotectant metabolite betaine. Effective osmoregulation has also been implicated in the acquirement of a profound antibiotic resistance by MRSA. Betaine can be obtained from the bacterial habitat or produced intracellularly from choline via the toxic betaine aldehyde (BA) employing the choline dehydrogenase and betaine aldehyde dehydrogenase (BADH) enzymes. Here, it is shown that the putative betaine aldehyde dehydrogenase SACOL2628 from the early MRSA isolate COL (SaBADH) utilizes betaine aldehyde as the primary substrate and nicotinamide adenine dinucleotide (NAD(+)) as the cofactor. Surface plasmon resonance experiments revealed that the affinity of NAD(+), NADH and BA for SaBADH is affected by temperature, pH and buffer composition. Five crystal structures of the wild type and three structures of the Gly234Ser mutant of SaBADH in the apo and holo forms provide details of the molecular mechanisms of activity and substrate specificity/inhibition of this enzyme.

  11. Structural and functional analysis of betaine aldehyde dehydrogenase from Staphylococcus aureus

    SciTech Connect

    Halavaty, Andrei S.; Rich, Rebecca L.; Chen, Chao; Joo, Jeong Chan; Minasov, George; Dubrovska, Ievgeniia; Winsor, James R.; Myszka, David G.; Duban, Mark; Shuvalova, Ludmilla; Yakunin, Alexander F.; Anderson, Wayne F.

    2015-04-25

    When exposed to high osmolarity, methicillin-resistant Staphylococcus aureus (MRSA) restores its growth and establishes a new steady state by accumulating the osmoprotectant metabolite betaine. Effective osmoregulation has also been implicated in the acquirement of a profound antibiotic resistance by MRSA. Betaine can be obtained from the bacterial habitat or produced intracellularly from choline via the toxic betaine aldehyde (BA) employing the choline dehydrogenase and betaine aldehyde dehydrogenase (BADH) enzymes. Here, it is shown that the putative betaine aldehyde dehydrogenase SACOL2628 from the early MRSA isolate COL ( SaBADH) utilizes betaine aldehyde as the primary substrate and nicotinamide adenine dinucleotide (NAD+) as the cofactor. Surface plasmon resonance experiments revealed that the affinity of NAD+, NADH and BA for SaBADH is affected by temperature, pH and buffer composition. Finally, five crystal structures of the wild type and three structures of the Gly234Ser mutant of SaBADH in the apo and holo forms provide details of the molecular mechanisms of activity and substrate specificity/inhibition of this enzyme.

  12. Structural and functional analysis of betaine aldehyde dehydrogenase from Staphylococcus aureus

    PubMed Central

    Halavaty, Andrei S.; Rich, Rebecca L.; Chen, Chao; Joo, Jeong Chan; Minasov, George; Dubrovska, Ievgeniia; Winsor, James R.; Myszka, David G.; Duban, Mark; Shuvalova, Ludmilla; Yakunin, Alexander F.; Anderson, Wayne F.

    2015-01-01

    When exposed to high osmolarity, methicillin-resistant Staphylococcus aureus (MRSA) restores its growth and establishes a new steady state by accumulating the osmoprotectant metabolite betaine. Effective osmoregulation has also been implicated in the acquirement of a profound antibiotic resistance by MRSA. Betaine can be obtained from the bacterial habitat or produced intracellularly from choline via the toxic betaine aldehyde (BA) employing the choline dehydrogenase and betaine aldehyde dehydrogenase (BADH) enzymes. Here, it is shown that the putative betaine aldehyde dehydrogenase SACOL2628 from the early MRSA isolate COL (SaBADH) utilizes betaine aldehyde as the primary substrate and nicotinamide adenine dinucleotide (NAD+) as the cofactor. Surface plasmon resonance experiments revealed that the affinity of NAD+, NADH and BA for SaBADH is affected by temperature, pH and buffer composition. Five crystal structures of the wild type and three structures of the Gly234Ser mutant of SaBADH in the apo and holo forms provide details of the molecular mechanisms of activity and substrate specificity/inhibition of this enzyme. PMID:25945581

  13. Structural and functional analysis of betaine aldehyde dehydrogenase from Staphylococcus aureus

    DOE PAGES

    Halavaty, Andrei S.; Rich, Rebecca L.; Chen, Chao; ...

    2015-04-25

    When exposed to high osmolarity, methicillin-resistant Staphylococcus aureus (MRSA) restores its growth and establishes a new steady state by accumulating the osmoprotectant metabolite betaine. Effective osmoregulation has also been implicated in the acquirement of a profound antibiotic resistance by MRSA. Betaine can be obtained from the bacterial habitat or produced intracellularly from choline via the toxic betaine aldehyde (BA) employing the choline dehydrogenase and betaine aldehyde dehydrogenase (BADH) enzymes. Here, it is shown that the putative betaine aldehyde dehydrogenase SACOL2628 from the early MRSA isolate COL ( SaBADH) utilizes betaine aldehyde as the primary substrate and nicotinamide adenine dinucleotide (NAD+)more » as the cofactor. Surface plasmon resonance experiments revealed that the affinity of NAD+, NADH and BA for SaBADH is affected by temperature, pH and buffer composition. Finally, five crystal structures of the wild type and three structures of the Gly234Ser mutant of SaBADH in the apo and holo forms provide details of the molecular mechanisms of activity and substrate specificity/inhibition of this enzyme.« less

  14. Modulation of ethanol stress tolerance by aldehyde dehydrogenase in the mycorrhizal fungus Tricholoma vaccinum.

    PubMed

    Asiimwe, Theodore; Krause, Katrin; Schlunk, Ines; Kothe, Erika

    2012-08-01

    We report the first mycorrhizal fungal aldehyde dehydrogenase gene, ald1, which was isolated from the basidiomycete Tricholoma vaccinum. The gene, encoding a protein Ald1 of 502 amino acids, is up-regulated in ectomycorrhiza. Phylogenetic analyses using 53 specific fungal aldehyde dehydrogenases from all major phyla in the kingdom of fungi including Ald1 and two partial sequences of T. vaccinum were performed to get an insight in the evolution of the aldehyde dehydrogenase family. By using competitive and real-time RT-PCR, ald1 is up-regulated in response to alcohol and aldehyde-related stress. Furthermore, heterologous expression of ald1 in Escherichia coli and subsequent in vitro enzyme activity assay demonstrated the oxidation of propionaldehyde and butyraldehyde with different kinetics using either NAD(+) or NADP(+) as cofactors. In addition, overexpression of ald1 in T. vaccinum after Agrobacterium tumefaciens-mediated transformation increased ethanol stress tolerance. These results demonstrate the ability of Ald1 to circumvent ethanol stress, a critical function in mycorrhizal habitats.

  15. Aldehyde Dehydrogenases in Arabidopsis thaliana: Biochemical Requirements, Metabolic Pathways, and Functional Analysis.

    PubMed

    Stiti, Naim; Missihoun, Tagnon D; Kotchoni, Simeon O; Kirch, Hans-Hubert; Bartels, Dorothea

    2011-01-01

    Aldehyde dehydrogenases (ALDHs) are a family of enzymes which catalyze the oxidation of reactive aldehydes to their corresponding carboxylic acids. Here we summarize molecular genetic and biochemical analyses of selected ArabidopsisALDH genes. Aldehyde molecules are very reactive and are involved in many metabolic processes but when they accumulate in excess they become toxic. Thus activity of aldehyde dehydrogenases is important in regulating the homeostasis of aldehydes. Overexpression of some ALDH genes demonstrated an improved abiotic stress tolerance. Despite the fact that several reports are available describing a role for specific ALDHs, their precise physiological roles are often still unclear. Therefore a number of genetic and biochemical tools have been generated to address the function with an emphasis on stress-related ALDHs. ALDHs exert their functions in different cellular compartments and often in a developmental and tissue specific manner. To investigate substrate specificity, catalytic efficiencies have been determined using a range of substrates varying in carbon chain length and degree of carbon oxidation. Mutational approaches identified amino acid residues critical for coenzyme usage and enzyme activities.

  16. Salivary aldehyde dehydrogenase - temporal and population variability, correlations with drinking and smoking habits and activity towards aldehydes contained in food.

    PubMed

    Giebułtowicz, Joanna; Dziadek, Marta; Wroczyński, Piotr; Woźnicka, Katarzyna; Wojno, Barbara; Pietrzak, Monika; Wierzchowski, Jacek

    2010-01-01

    Fluorimetric method based on oxidation of the fluorogenic 6-methoxy-2-naphthaldehyde was applied to evaluate temporal and population variability of the specific activity of salivary aldehyde dehydrogenase (ALDH) and the degree of its inactivation in healthy human population. Analyzed was also its dependence on drinking and smoking habits, coffee consumption, and its sensitivity to N-acetylcysteine. Both the specific activity of salivary ALDH and the degree of its inactivation were highly variable during the day, with the highest activities recorded in the morning hours. The activities were also highly variable both intra- and interpersonally, and negatively correlated with age, and this correlation was stronger for the subgroup of volunteers declaring abstinence from alcohol and tobacco. Moderately positive correlations of salivary ALDH specific activity with alcohol consumption and tobacco smoking were also recorded (r(s) ~0.27; p=0.004 and r(s) =0.30; p=0.001, respectively). Moderate coffee consumption correlated positively with the inactivation of salivary ALDH, particularly in the subgroup of non-drinking and non-smoking volunteers. It was found that mechanical stimulation of the saliva flow increases the specific activity of salivary ALDH. The specific activity of the salivary ALDH was strongly and positively correlated with that of superoxide dismutase, and somewhat less with salivary peroxidase. The antioxidant-containing drug N-acetylcysteine increased activity of salivary ALDH presumably by preventing its inactivation in the oral cavity. Some food-related aldehydes, mainly cinnamic aldehyde and anisaldehyde, were excellent substrates of the salivary ALDH3A1 enzyme, while alkenals, particularly those with short chain, were characterized by lower affinity towards this enzyme but high catalytic constants. The protective role of salivary ALDH against aldehydes in food and those found in the cigarette smoke is discussed, as well as its participation in

  17. Peptidyl aldehydes as reversible covalent inhibitors of protein tyrosine phosphatases.

    PubMed

    Fu, Hua; Park, Junguk; Pei, Dehua

    2002-08-27

    Protein tyrosine phosphatases (PTPs) are a large family of enzymes that catalyze the hydrolytic removal of the phosphoryl group from phosphotyrosyl (pY) proteins. PTP inhibitors provide potential treatment of human diseases/conditions such as diabetes and obesity as well as useful tools for studying the function of PTPs in signaling pathways. In this work, we have shown that certain aryl-substituted aldehydes act as reversible, slow-binding inhibitors of modest potency against PTP1B, SHP-1, and a dual-specificity phosphatase, VHR. Attachment of the tripeptide Gly-Glu-Glu to the para position of cinnamaldehyde resulted in an inhibitor (Cinn-GEE) of substantially increased potency against all three enzymes (e.g., K(I) = 5.4 microM against PTP1B). The mechanism of inhibition was investigated using Cinn-GEE specifically labeled with (13)C at the aldehyde carbon and (1)H-(13)C heteronuclear single-quantum coherence spectroscopy. While Cinn-GEE alone showed a single cross-peak at delta 9.64 ((1)H) and delta 201 ((13)C), the PTP1B/Cinn-GEE complex showed three distinct cross-peaks at delta 7.6-7.8 ((1)H) and 130-137 ((13)C). Mutation of the catalytic cysteine (Cys-215 in PTP1B) into alanine had no effect on the cross-peaks, whereas mutation of a conserved active-site arginine (Arg-221 in PTP1B) to alanine abolished all three cross-peaks. Similar experiments with Cinn-GEE that had been labeled with (13)C at the benzylic position revealed a change in the hybridization state (from sp(2) to sp(3)) for the benzylic carbon as a result of binding to PTP1B. These results rule out the possibility of a free aldehyde, aldehyde hydrate, or hemithioacetal as the enzyme-bound inhibitor form. Instead, the data are consistent with the formation of an enamine between the aldehyde group of the inhibitor and the guanidine group of Arg-221 in the PTP1B active site. These aldehydes may provide a general core structure that can be further developed into highly potent and specific PTP

  18. Peptide aldehyde inhibitors of hepatitis A virus 3C proteinase.

    PubMed

    Malcolm, B A; Lowe, C; Shechosky, S; McKay, R T; Yang, C C; Shah, V J; Simon, R J; Vederas, J C; Santi, D V

    1995-06-27

    Picornaviral 3C proteinases are a group of closely related thiol proteinases responsible for processing of the viral polyprotein into its component proteins. These proteinases adopt a chymotrypsin-like fold [Allaire et al. (1994) Nature 369, 72-77; Matthews et al. (1994) Cell 77, 761-771] and a display an active-site configuration like those of the serine proteinases. Peptide-aldehydes based on the preferred peptide substrates for hepatitis A virus (HAV) 3C proteinase were synthesized by reduction of a thioester precursor. Acetyl-Leu-Ala-Ala-(N,N'-dimethylglutaminal) was found to be a reversible, slow-binding inhibitor for HAV 3C with a Ki* of (4.2 +/- 0.8) x 10(-8) M. This inhibitor showed 50-fold less activity against the highly homologous human rhinovirus (strain 14) 3C proteinase, whose peptide substrate specificity is slightly different, suggesting a high degree of selectivity. NMR spectrometry of the adduct of the 13C-labeled inhibitor with the HAV-3C proteinase indicate that a thiohemiacetal is formed between the enzyme and the aldehyde carbon as previously noted for peptide-aldehyde inhibitors of papain [Lewis & Wolfenden (1977) Biochemistry 16,4890-4894; Gamcsik et al. (1983) J. Am. Chem. Soc. 105, 6324-6325]. The adduct can also be observed by electrospray mass spectrometry.

  19. Aldehyde dehydrogenase 3A1 associates with prostate tumorigenesis

    PubMed Central

    Yan, J; De Melo, J; Cutz, J-C; Aziz, T; Tang, D

    2014-01-01

    Background: Accumulating evidence demonstrates high levels of aldehyde dehydrogense (ALDH) activity in human cancer types, in part, because of its association with cancer stem cells. Whereas ALDH1A1 and ALDH7A1 isoforms were reported to associate with prostate tumorigenesis, whether other ALDH isoforms are associated with prostate cancer (PC) remains unclear. Methods: ALDH3A1 expression was analysed in various PC cell lines. Xenograft tumours and 54 primary and metastatic PC tumours were stained using immunohistochemistry for ALDH3A1 expression. Results: In comparison with the non-stem counterparts, a robust upregulation of ALDH3A1 was observed in DU145-derived PC stem cells (PCSCs). As DU145 PCSCs produced xenograft tumours with more advanced features compared with those derived from DU145 cells, higher levels of ALDH3A1 were detected in the former; a dramatic elevation of ALDH3A1 occurred in DU145 cell-derived lung metastasis compared with local xenograft tumours. Furthermore, while ALDH3A1 was not observed in prostate glands, ALDH3A1 was clearly present in PIN, and further increased in carcinomas. In comparison with the paired local carcinomas, ALDH3A1 was upregulated in lymph node metastatic tumours; the presence of ALDH3A1 in bone metastatic PC was also demonstrated. Conclusions: We report here the association of ALDH3A1 with PC progression. PMID:24762960

  20. Expression and activity of alcohol and aldehyde dehydrogenases in melanoma cells and in melanocytes.

    PubMed

    Amann, Philipp M; Hofmann, Claudia; Freudenberger, Muriel; Holland-Cunz, Stefan; Eichmüller, Stefan B; Bazhin, Alexandr V

    2012-03-01

    Disturbances in vitamin A metabolism are an important attribute of some cancer cells. Most evidence point that these disturbances lead to decreasing of the retinoic acid concentration in tumor cells. Up to now, in benign and malignant skin cells the features of vitamin A metabolism with its participating enzymes are not entirely understood. Alcohol and aldehyde dehydrogenases (ALDH) are involved in the retinol metabolism, oxidizing retinol, and retinal in retinoic acid or reducing retinal in retinol. In this work we investigated the expression and enzymatic activity of alcohol and ALDH in melanoma cells compared to their benign counterparts. We demonstrated that melanoma cell lines and melanocytes despite similar pattern of the enzyme expression, show different general ALDH activity. Retinal, the substrate of ALDH, could stimulate the ALDH activity through up-regulation of retinaldehyde dehydrogenase 1 and aldehyde dehydrogenase 6. Furthermore, we found that retinoids regulate alcohol dehydrogenase activity, probably via effects on alcohol dehydrogenase expression at the post-transcriptional level. We suggest that melanoma cells in contrast to melanocytes should favor the retinal reduction over its oxidation. The decreasing cellular amount of the precursor molecules of retinoic acid could result in a changed gene regulation in melanoma cells. Copyright © 2011 Wiley Periodicals, Inc.

  1. Effects of zinc deficiency on ethanol metabolism and alcohol and aldehyde dehydrogenase activities.

    PubMed

    Das, I; Burch, R E; Hahn, H K

    1984-10-01

    Alcohol dehydrogenase, low Km and high Km mitochondrial and microsomal aldehyde dehydrogenase, and in vivo ethanol elimination rates were determined in five groups of male Sprague-Dawley rats given liquid diets, as follows: control (C), control plus 5% ethanol (CE), pair-fed control and zinc-deficient (PC-ZnD), zinc-deficient (ZnD), and zinc-deficient plus 5% ethanol (ZnDE). Rats fed CE had decreased liver and serum zinc content. The animals given ZnD diets had an even more dramatic decrease in their tissue zinc concentrations and displayed marked growth retardation. The in vivo blood ethanol elimination rate was increased in animals fed ethanol, and this increase was accompanied by increased alcohol and aldehyde dehydrogenase activities. There was a significant decrease in the ethanol elimination rate in rats given ZnD and ZnDE diets. Alcohol dehydrogenase activities in rats with deficient zinc levels also were decreased, and there were no changes in acetaldehyde dehydrogenase activities. Our results suggest that the metabolism of ethanol to acetaldehyde is impaired in zinc deficiency, but acetaldehyde to acetate conversion appears normal.

  2. Oxidation of alcohols and reduction of aldehydes derived from methyl- and dimethylpyrenes by cDNA-expressed human alcohol dehydrogenases.

    PubMed

    Kollock, Ronny; Frank, Heinz; Seidel, Albrecht; Meinl, Walter; Glatt, Hansruedi

    2008-03-12

    Some methylated pyrenes can form DNA adducts in rat tissues after benzylic hydroxylation and sulpho conjugation. However, oxidation of the intermediate alcohols to carboxylic acids is an important competing pathway leading to detoxification. We previously showed that co-administration of ethanol or 4-methylpyrazole strongly enhances DNA adduct formation by 1-hydroxymethylpyrene, indicating an involvement of alcohol dehydrogenases (ADHs) in the detoxification. This mechanism may be involved in the observed synergism of smoking and alcohol consumption in certain human cancers. In a preceding study, cDNA-expressed human ADH2 efficiently oxidised 1-, 2- and 4-hydroxymethylpyrene; these reactions were inhibited in the presence of ethanol or 4-methylpyrazole. Here we report that ADH1C, ADH3 and ADH4 also show substantial activity towards these substrates and two further congeners, 1-hydroxymethyl-6-methylpyrene and 1-hydroxymethyl-8-methylpyrene. All four ADH forms also catalysed the reverse reaction, implying that the aldehydes have to be sequestered by other enzymes, such as aldehyde dehydrogenases, for final detoxification. ADH1C and ADH4 activities towards hydroxymethylpyrenes were more strongly inhibited in the presence of ethanol and 4-methylpyrazole than those of ADH2. ADH3 was only inhibited at very high concentrations of the modulators. In conclusions, several human ADHs are capable of detoxifying benzylic alcohols of alkylated polycyclic aromatic hydrocarbons. However, some competing substrates and inhibitors may affect all these redundant detoxification systems, although to various extents.

  3. Aldehyde dehydrogenase polymorphism in North American, South American, and Mexican Indian populations.

    PubMed Central

    Goedde, H W; Agarwal, D P; Harada, S; Rothhammer, F; Whittaker, J O; Lisker, R

    1986-01-01

    While about 40% of the South American Indian populations (Atacameños, Mapuche, Shuara) were found to be deficient in aldehyde dehydrogenase isozyme I (ALDH2 or E2), preliminary investigations showed very low incidence of isozyme deficiency among North American natives (Sioux, Navajo) and Mexican Indians (mestizo). Possible implications of such trait differences on cross-cultural behavioral response to alcohol drinking are discussed. PMID:3953578

  4. Aldehyde dehydrogenase polymorphism in North American, South American, and Mexican Indian populations.

    PubMed

    Goedde, H W; Agarwal, D P; Harada, S; Rothhammer, F; Whittaker, J O; Lisker, R

    1986-03-01

    While about 40% of the South American Indian populations (Atacameños, Mapuche, Shuara) were found to be deficient in aldehyde dehydrogenase isozyme I (ALDH2 or E2), preliminary investigations showed very low incidence of isozyme deficiency among North American natives (Sioux, Navajo) and Mexican Indians (mestizo). Possible implications of such trait differences on cross-cultural behavioral response to alcohol drinking are discussed.

  5. Fatty aldehyde dehydrogenases in Acinetobacter sp. strain HO1-N: role in hexadecanol metabolism.

    PubMed Central

    Singer, M E; Finnerty, W R

    1985-01-01

    The role of fatty aldehyde dehydrogenases (FALDHs) in hexadecane and hexadecanol metabolism was studied in Acinetobacter sp. strain HO1-N. Two distinct FALDHs were demonstrated in Acinetobacter sp. strain HO1-N: a membrane-bound, NADP-dependent FALDH activity induced 5-, 15-, and 9-fold by growth on hexadecanol, dodecyl aldehyde, and hexadecane, respectively, and a constitutive, NAD-dependent, membrane-localized FALDH. The NADP-dependent FALDH exhibited apparent Km and Vmax values for decyl aldehyde of 5.0, 13.0, 18.0, and 18.3 microM and 537.0, 500.0, 25.0, and 38.0 nmol/min in hexadecane-, hexadecanol-, ethanol-, palmitate-grown cells, respectively. FALDH isozymes ald-a, ald-b, and ald-c were demonstrated by gel electrophoresis in extracts of hexadecane- and hexadecanol-grown cells. ald-a, ald-b, and ald-d were present in dodecyl aldehyde-grown cells, while palmitate-grown control cells contained ald-b and ald-d. Dodecyl aldehyde-negative mutants were isolated and grouped into two phenotypic classes based on growth: class 1 mutants were hexadecane and hexadecanol negative and class 2 mutants were hexadecane and hexadecanol positive. Specific activity of NADP-dependent FALDH in Ald21 (class 1 mutant) was 85% lower than that of wild-type FALDH, while the specific activity of Ald24 (class 2 mutant) was 55% greater than that of wild-type FALDH. Ald21R, a dodecyl aldehyde-positive revertant able to grow on hexadecane, hexadecanol, and dodecyl aldehyde, exhibited a 100% increase in the specific activity of the NADP-dependent FALDH. The oxidation of [3H]hexadecane byAld21 yielded the accumulation of 61% more fatty aldehyde than the wild type, while Ald24 accumulated 27% more fatty aldehyde, 95% more fatty alcohol, and 65% more wax ester than the wild type. This study provides genetic and physiological evidence for the role of fatty aldehyde as an essential metabolic intermediate and NADP-dependent FALDH as a key enzyme in the dissimilation of hexadecane, hexadecanol

  6. Fluorescence lifetime analysis and effect of magnesium ions on binding of NADH to human aldehyde dehydrogenase 1

    USDA-ARS?s Scientific Manuscript database

    Aldehyde dehydrogenase 1 (ALDH1) catalyzes oxidation of toxic aldehydes to carboxylic acids. Physiologic levels of Mg2+ ions influence ALDH1 activity in part by increasing NADH binding affinity to the enzyme thus reducing activity. By using time-resolved fluorescence spectroscopy, we have resolved t...

  7. Characterisation of recombinant human fatty aldehyde dehydrogenase: implications for Sjögren-Larsson syndrome.

    PubMed

    Lloyd, Matthew D; Boardman, Kieren D E; Smith, Andrew; van den Brink, Daan M; Wanders, Ronald J A; Threadgill, Michael D

    2007-10-01

    Fatty aldehyde dehydrogenase (FALDH) is an NAD+-dependent oxidoreductase involved in the metabolism of fatty alcohols. Enzyme activity has been implicated in the pathology of diabetes and cancer. Mutations in the human gene inactivate the enzyme and cause accumulation of fatty alcohols in Sjögren-Larsson syndrome, a neurological disorder resulting in physical and mental handicaps. Microsomal FALDH was expressed in E. coli and purified. Using an in vitro activity assay an optimum pH of approximately 9.5 and temperature of approximately 35 degrees C were determined. Medium- and long-chain fatty aldehydes were converted to the corresponding acids and kinetic parameters determined. The enzyme showed high activity with heptanal, tetradecanal, hexadecanal and octadecanal with lower activities for the other tested substrates. The enzyme was also able to convert some fatty alcohol substrates to their corresponding aldehydes and acids, at 25-30% the rate of aldehyde oxidation. A structural model of FALDH has been constructed, and catalytically important residues have been proposed to be involved in alcohol and aldehyde oxidation: Gln-120, Glu-207, Cys-241, Phe-333, Tyr-410 and His-411. These results place FALDH in a central role in the fatty alcohol/acid interconversion cycle, and provide a direct link between enzyme inactivation and disease pathology caused by accumulation of alcohols.

  8. The effect of inhibition of aldehyde dehydrogenase on nasal uptake of inspired acetaldehyde.

    PubMed

    Stanek, J J; Morris, J B

    1999-06-01

    At exposure concentrations of 750 ppm or more, acetaldehyde is a rodent inhalation carcinogen that induces nasal tumors. Aldehyde dehydrogenase (ALDH) is thought to be an important detoxifying enzyme for aldehydes. Although nasal tissues express ALDH, the importance of this enzyme relative to delivered dosage rates at high-inspired concentrations is not well defined. To provide such information, uptake of inspired acetaldehyde was measured at an inspiratory flow rate that approximated the minute ventilation rate in the surgically isolated nasal cavity of F 344 rats pretreated with either saline (control) or the ALDH inhibitor, cyanamide (10 mg/kg s.c.). ALDH activities (substrate concentration 3 times the K(m)) in anterior (respiratory mucosa) and posterior (olfactory mucosa) nasal tissues averaged 160 and 210 nmol/min), respectively, in control animals (total activity 370 nmol/min), compared to 60 and 80 nmol/min, respectively, in cyanamide-pretreated rats (p < 0.05), indicating that approximately 60% inhibition was obtained. Nasal uptake was measured at 3 inspired concentrations: 10, 300, and 1500 ppm. At these concentrations, acetaldehyde uptake efficiency averaged 54, 37, and 34% in saline-pretreated rats, respectively (p < 0.05). In absolute terms, the delivered dosage rates at these exposure concentrations averaged 21, 420, and 1990 nmol/min. The concentration dependence on uptake suggests a saturable process was involved. At inspired concentrations of 300 ppm or more, the delivered dosage rates exceeded the measured specific activity for nasal ALDH of 370 nmol/min. Cyanamide pretreatment abolished the concentration dependence. Specifically, uptake efficiencies in cyanamide-pretreated rats averaged 30, 27, and 31% at inspired concentrations of 10, 300, and 1500 ppm, respectively (p > 0.05); delivered dosage rates were 12, 310, and 1780 nmol/min. Thus, cyanamide pretreatment reduced nasal-delivered dosage rates at inspired concentrations of 10, 300, and 1500

  9. Aldehyde dehydrogenase activity selects for lung adenocarcinoma stem cells dependent on notch signaling.

    PubMed

    Sullivan, James P; Spinola, Monica; Dodge, Michael; Raso, Maria G; Behrens, Carmen; Gao, Boning; Schuster, Katja; Shao, Chunli; Larsen, Jill E; Sullivan, Laura A; Honorio, Sofia; Xie, Yang; Scaglioni, Pier P; DiMaio, J Michael; Gazdar, Adi F; Shay, Jerry W; Wistuba, Ignacio I; Minna, John D

    2010-12-01

    Aldehyde dehydrogenase (ALDH) is a candidate marker for lung cancer cells with stem cell-like properties. Immunohistochemical staining of a large panel of primary non-small cell lung cancer (NSCLC) samples for ALDH1A1, ALDH3A1, and CD133 revealed a significant correlation between ALDH1A1 (but not ALDH3A1 or CD133) expression and poor prognosis in patients including those with stage I and N0 disease. Flow cytometric analysis of a panel of lung cancer cell lines and patient tumors revealed that most NSCLCs contain a subpopulation of cells with elevated ALDH activity, and that this activity is associated with ALDH1A1 expression. Isolated ALDH(+) lung cancer cells were observed to be highly tumorigenic and clonogenic as well as capable of self-renewal compared with their ALDH(-) counterparts. Expression analysis of sorted cells revealed elevated Notch pathway transcript expression in ALDH(+) cells. Suppression of the Notch pathway by treatment with either a γ-secretase inhibitor or stable expression of shRNA against NOTCH3 resulted in a significant decrease in ALDH(+) lung cancer cells, commensurate with a reduction in tumor cell proliferation and clonogenicity. Taken together, these findings indicate that ALDH selects for a subpopulation of self-renewing NSCLC stem-like cells with increased tumorigenic potential, that NSCLCs harboring tumor cells with ALDH1A1 expression have inferior prognosis, and that ALDH1A1 and CD133 identify different tumor subpopulations. Therapeutic targeting of the Notch pathway reduces this ALDH(+) component, implicating Notch signaling in lung cancer stem cell maintenance.

  10. Role of aldehyde dehydrogenase in hypoxic vasodilator effects of nitrite in rats and humans.

    PubMed

    Arif, Sayqa; Borgognone, Alessandra; Lin, Erica Lai-Sze; O'Sullivan, Aine G; Sharma, Vishal; Drury, Nigel E; Menon, Ashvini; Nightingale, Peter; Mascaro, Jorge; Bonser, Robert S; Horowitz, John D; Feelisch, Martin; Frenneaux, Michael P; Madhani, Melanie

    2015-07-01

    Hypoxic conditions favour the reduction of nitrite to nitric oxide (NO) to elicit vasodilatation, but the mechanism(s) responsible for bioconversion remains ill defined. In the present study, we assess the role of aldehyde dehydrogenase 2 (ALDH2) in nitrite bioactivation under normoxia and hypoxia in the rat and human vasculature. The role of ALDH2 in vascular responses to nitrite was studied using rat thoracic aorta and gluteal subcutaneous fat resistance vessels from patients with heart failure (HF; 16 patients) in vitro and by measurement of changes in forearm blood flow (FBF) during intra-arterial nitrite infusion (21 patients) in vivo. Specifically, we investigated the effects of (i) ALDH2 inhibition by cyanamide or propionaldehyde and the (ii) tolerance-independent inactivation of ALDH2 by glyceryl trinitrate (GTN) on the vasodilator activity of nitrite. In each setting, nitrite effects were measured via evaluation of the concentration-response relationship under normoxic and hypoxic conditions in the absence or presence of ALDH2 inhibitors. Both in rat aorta and human resistance vessels, dilatation to nitrite was diminished following ALDH2 inhibition, in particular under hypoxia. In humans there was a non-significant trend towards attenuation of nitrite-mediated increases in FBF. In human and rat vascular tissue in vitro, hypoxic nitrite-mediated vasodilatation involves ALDH2. In patients with HF in vivo, the role of this enzyme in nitrite bioactivation is at the most, modest, suggesting the involvement of other more important mechanisms. © 2015 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of The British Pharmacological Society.

  11. Aldehyde dehydrogenase activity selects for lung adenocarcinoma stem cells dependent on Notch signaling

    PubMed Central

    Sullivan, James P.; Spinola, Monica; Dodge, Michael; Raso, Maria G.; Behrens, Carmen; Gao, Boning; Schuster, Katja; Shao, Chunli; Larsen, Jill E.; Sullivan, Laura A.; Honorio, Sofia; Xie, Yang; Scaglioni, Pier P.; DiMaio, J. Michael; Gazdar, Adi F.; Shay, Jerry W.; Wistuba, Ignacio I.; Minna, John D.

    2010-01-01

    Aldehyde dehydrogenase (ALDH) is a candidate marker for lung cancer cells with stem cell-like properties. Immunohistochemical staining of a large panel of primary non-small cell lung cancer (NSCLC) samples for the ALDH1A1, ALDH3A1 and CD133 revealed a significant correlation between ALDH1A1 (but not ALDH3A1 or CD133) expression and poor prognosis in patients including those with stage I and N0 disease. Flow cytometric analysis of a panel of lung cancer cell lines and patient tumors revealed most NSCLCs contain a subpopulation of cells with elevated ALDH activity, and that this activity is associated with ALDH1A1 expression. Isolated ALDH+ lung cancer cells were observed to be highly tumorigenic and clonogenic as well as capable of self-renewal compared to their ALDH− counterparts. Expression analysis of sorted cells revealed elevated Notch pathway transcript expression in ALDH+ cells. Suppression of the Notch pathway by treatment with either a gamma-secretase inhibitor or stable expression of shRNA against NOTCH3 resulted in a significant decrease in ALDH+ lung cancer cells, commensurate with a reduction in tumor cell proliferation and clonogenicity. Taken together, these findings indicate that ALDH selects for a subpopulation of self-renewing NSCLC stem-like cells with increased tumorigenic potential, that NSCLCs harboring tumor cells with ALDH1A1 expression have inferior prognosis, and that ALDH1A1 and CD133 identify different tumor subpopulations. Therapeutic targeting of the Notch pathway reduces this ALDH+ component, implicating Notch signaling in lung cancer stem cell maintenance. PMID:21118965

  12. Inhibition of human alcohol and aldehyde dehydrogenases by cimetidine and assessment of its effects on ethanol metabolism.

    PubMed

    Lai, Ching-Long; Li, Yeung-Pin; Liu, Chiu-Ming; Hsieh, Hsiu-Shan; Yin, Shih-Jiun

    2013-02-25

    Previous studies have reported that cimetidine, an H2-receptor antagonist used to treat gastric and duodenal ulcers, can inhibit alcohol dehydrogenases (ADHs) and ethanol metabolism. Human alcohol dehydrogenases and aldehyde dehydrogenases (ALDHs), the principal enzymes responsible for metabolism of ethanol, are complex enzyme families that exhibit functional polymorphisms among ethnic groups and distinct tissue distributions. We investigated the inhibition by cimetidine of alcohol oxidation by recombinant human ADH1A, ADH1B1, ADH1B2, ADH1B3, ADH1C1, ADH1C2, ADH2, and ADH4, and aldehyde oxidation by ALDH1A1 and ALDH2 at pH 7.5 and a cytosolic NAD(+) concentration. Cimetidine acted as competitive or noncompetitive inhibitors for the ADH and ALDH isozymes/allozymes with near mM inhibition constants. The metabolic interactions between cimetidine and ethanol/acetaldehyde were assessed by computer simulation using the inhibition equations and the determined kinetic constants. At therapeutic drug levels (0.015 mM) and physiologically relevant concentrations of ethanol (10 mM) and acetaldehyde (10 μM) in target tissues, cimetidine could weakly inhibit (<5%) the activities of ADH1B2 and ADH1B3 in liver, ADH2 in liver and small intestine, ADH4 in stomach, and ALDH1A1 in the three tissues, but not significantly affect ADH1A, ADH1B1, ADH1C1/2, or ALDH2. At higher drug levels, which may accumulate in cells (0.2 mM), the activities of the weakly-inhibited enzymes may be decreased more significantly. The quantitative effects of cimetidine on metabolism of ethanol and other physiological substrates of ADHs need further investigation. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  13. Enrichment of chemical libraries docked to protein conformational ensembles and application to aldehyde dehydrogenase 2.

    PubMed

    Wang, Bo; Buchman, Cameron D; Li, Liwei; Hurley, Thomas D; Meroueh, Samy O

    2014-07-28

    Molecular recognition is a complex process that involves a large ensemble of structures of the receptor and ligand. Yet, most structure-based virtual screening is carried out on a single structure typically from X-ray crystallography. Explicit-solvent molecular dynamics (MD) simulations offer an opportunity to sample multiple conformational states of a protein. Here we evaluate our recently developed scoring method SVMSP in its ability to enrich chemical libraries docked to MD structures of seven proteins from the Directory of Useful Decoys (DUD). SVMSP is a target-specific rescoring method that combines machine learning with statistical potentials. We find that enrichment power as measured by the area under the ROC curve (ROC-AUC) is not affected by increasing the number of MD structures. Among individual MD snapshots, many exhibited enrichment that was significantly better than the crystal structure, but no correlation between enrichment and structural deviation from crystal structure was found. We followed an innovative approach by training SVMSP scoring models using MD structures (SVMSPMD). The resulting models were applied to two difficult cases (p38 and CDK2) for which enrichment was not better than random. We found remarkable increase in enrichment power, particularly for p38, where the ROC-AUC increased by 0.30 to 0.85. Finally, we explored approaches for a priori identification of MD snapshots with high enrichment power from an MD simulation in the absence of active compounds. We found that the use of randomly selected compounds docked to the target of interest using SVMSP led to notable enrichment for EGFR and Src MD snapshots. SVMSP rescoring of protein-compound MD structures was applied for the search of small-molecule inhibitors of the mitochondrial enzyme aldehyde dehydrogenase 2 (ALDH2). Rank-ordering of a commercial library of 50 000 compounds docked to MD structures of ALDH2 led to five small-molecule inhibitors. Four compounds had IC50s below 5

  14. Enrichment of Chemical Libraries Docked to Protein Conformational Ensembles and Application to Aldehyde Dehydrogenase 2

    PubMed Central

    2015-01-01

    Molecular recognition is a complex process that involves a large ensemble of structures of the receptor and ligand. Yet, most structure-based virtual screening is carried out on a single structure typically from X-ray crystallography. Explicit-solvent molecular dynamics (MD) simulations offer an opportunity to sample multiple conformational states of a protein. Here we evaluate our recently developed scoring method SVMSP in its ability to enrich chemical libraries docked to MD structures of seven proteins from the Directory of Useful Decoys (DUD). SVMSP is a target-specific rescoring method that combines machine learning with statistical potentials. We find that enrichment power as measured by the area under the ROC curve (ROC-AUC) is not affected by increasing the number of MD structures. Among individual MD snapshots, many exhibited enrichment that was significantly better than the crystal structure, but no correlation between enrichment and structural deviation from crystal structure was found. We followed an innovative approach by training SVMSP scoring models using MD structures (SVMSPMD). The resulting models were applied to two difficult cases (p38 and CDK2) for which enrichment was not better than random. We found remarkable increase in enrichment power, particularly for p38, where the ROC-AUC increased by 0.30 to 0.85. Finally, we explored approaches for a priori identification of MD snapshots with high enrichment power from an MD simulation in the absence of active compounds. We found that the use of randomly selected compounds docked to the target of interest using SVMSP led to notable enrichment for EGFR and Src MD snapshots. SVMSP rescoring of protein–compound MD structures was applied for the search of small-molecule inhibitors of the mitochondrial enzyme aldehyde dehydrogenase 2 (ALDH2). Rank-ordering of a commercial library of 50 000 compounds docked to MD structures of ALDH2 led to five small-molecule inhibitors. Four compounds had IC50s below

  15. Evaluation of alcohol dehydrogenase and aldehyde dehydrogenase enzymes as bi-enzymatic anodes in a membraneless ethanol microfluidic fuel cell

    NASA Astrophysics Data System (ADS)

    Galindo-de-la-Rosa, J.; Arjona, N.; Arriaga, L. G.; Ledesma-García, J.; Guerra-Balcázar, M.

    2015-12-01

    Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (AldH) enzymes were immobilized by covalent binding and used as the anode in a bi-enzymatic membraneless ethanol hybrid microfluidic fuel cell. The purpose of using both enzymes was to optimize the ethanol electro-oxidation reaction (EOR) by using ADH toward its direct oxidation and AldH for the oxidation of aldehydes as by-products of the EOR. For this reason, three enzymatic bioanode configurations were evaluated according with the location of enzymes: combined, vertical and horizontally separated. In the combined configuration, a current density of 16.3 mA cm-2, a voltage of 1.14 V and a power density of 7.02 mW cm-2 were obtained. When enzymes were separately placed in a horizontal and vertical position the ocp drops to 0.94 V and to 0.68 V, respectively. The current density also falls to values of 13.63 and 5.05 mA cm-2. The decrease of cell performance of bioanodes with separated enzymes compared with the combined bioanode was of 31.7% and 86.87% for the horizontal and the vertical array.

  16. The action of progesterone and diethylstilboestrol on the dehydrogenase and esterase activities of a purified aldehyde dehydrogenase from rabbit liver.

    PubMed Central

    Julian, R; Duncan, S

    1977-01-01

    A steroid-sensitive aldehyde dehydrogenase (EC 1.2.1.3) was purified from rabbit liver and is homogeneous by the criterion of electrophoresis in polyacrylamide gels with or without sodium dodecyl sulphate. The enzyme is tetrameric, of subunit mo.wt. 48 300, and contains no tightly bound zinc. The fluorescence of the protein is decreased in the presence of progesterone, which is inhibitory to the reactions catalysed by the enzyme. When NADH is bound to the enzyme, the fluorescence of the coenzyme is augmented to an extent independent of the presence of steroids or acetaldehyde. The purified enzyme catalyses the oxidation of acetaldehyde and glucuronolactone, and the hydrolysis of 4-nitrophenyl acetate. Each of these reactions is inhibited by progesterone in such a manner as to suggest the formation of a catalytically active enzyme-hormone complex. Diethylstilboestrol inhibits the hydrolysis of esters by this enzyme, but stimulates the oxidation of aldehydes, except at low aldehyde concentrations; the ligand is then inhibitory. NADH inhibits the hydrolysis of 4-nitrophenyl acetate by the enzyme in a partially competitive fashion. PMID:192192

  17. Identification and overexpression of a bifunctional aldehyde/alcohol dehydrogenase responsible for ethanol production in Thermoanaerobacter mathranii.

    PubMed

    Yao, Shuo; Mikkelsen, Marie Just

    2010-01-01

    Thermoanaerobacter mathranii contains four genes, adhA, adhB, bdhA and adhE, predicted to code for alcohol dehydrogenases involved in ethanol metabolism. These alcohol dehydrogenases were characterized as NADP(H)-dependent primary alcohol dehydrogenase (AdhA), secondary alcohol dehydrogenase (AdhB), butanol dehydrogenase (BdhA) and NAD(H)-dependent bifunctional aldehyde/alcohol dehydrogenase (AdhE), respectively. Here we observed that AdhE is an important enzyme responsible for ethanol production in T. mathranii based on the constructed adh knockout strains. An adhE knockout strain fails to produce ethanol as a fermentation product, while other adh knockout strains showed no significant difference from the wild type. Further analysis revealed that the ΔadhE strain was defective in aldehyde dehydrogenase activity, but still maintained alcohol dehydrogenase activity. This showed that AdhE is the major aldehyde dehydrogenase in the cell and functions predominantly in the acetyl-CoA reduction to acetaldehyde in the ethanol formation pathway. Finally, AdhE was conditionally expressed from a xylose-induced promoter in a recombinant strain (BG1E1) with a concomitant deletion of a lactate dehydrogenase. Overexpressions of AdhE in strain BG1E1 with xylose as a substrate facilitate the production of ethanol at an increased yield. Copyright © 2010 S. Karger AG, Basel.

  18. The bifunctional aldehyde-alcohol dehydrogenase controls ethanol and acetate production in Entamoeba histolytica under aerobic conditions.

    PubMed

    Pineda, Erika; Encalada, Rusely; Olivos-García, Alfonso; Néquiz, Mario; Moreno-Sánchez, Rafael; Saavedra, Emma

    2013-01-16

    By applying metabolic control analysis and inhibitor titration we determined the degree of control (flux control coefficient) of pyruvate:ferredoxin oxidoreductase (PFOR) and bifunctional aldehyde-alcohol dehydrogenase (ADHE) over the fluxes of fermentative glycolysis of Entamoeba histolytica subjected to aerobic conditions. The flux-control coefficients towards ethanol and acetate formation determined for PFOR titrated with diphenyleneiodonium were 0.07 and 0.09, whereas for ADHE titrated with disulfiram were 0.33 and -0.19, respectively. ADHE inhibition induced significant accumulation of glycolytic intermediates and lower ATP content. These results indicate that ADHE exerts significant flux-control on the carbon end-product formation of amoebas subjected to aerobic conditions. Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  19. Molecular cloning, genomic organization, and chromosomal localization of an additional human aldehyde dehydrogenase gene, ALDH6

    SciTech Connect

    Hsu, L.C.; Wen-Chung, Chang; Hiraoka, L.

    1994-11-15

    Aldehyde dehydrogenase isozymes have been suggested to play a major role in the detoxification of aldehydes generated by alcohol metabolism and lipid peroxidation. The authors previously cloned and characterized four human nonallelic ALDH genes encoding different isozymes. The existence of a unique ALDH isozyme in human saliva and its polymorphism has been demonstrated previously. In this paper, they describe the cloning, characterization, and chromosomal mapping of an aldehyde dehydrogenase gene (ALDH6) expressed in the human salivary gland. The cloned ALDH6 cDNA is 3457 bp in length and contains an open reading frame encoding 512 amino acid residues. The deduced amino acid sequence showed that ALDH6 is larger than the human liver ALDH1 by 11 amino acid residues at the N-terminal, and the degree of identity between the two isozymes is 70% with an alignment of 500 amino acid residues. The human ALDH6 gene spans about 37 kb and consists of 13 exons. The putative TATA and CCAAT boxes and Sp1 binding sites are found in the 5{prime} upstream region of the gene. Northern blot analysis demonstrated that the ALDH6 gene is expressed at low levels in many tissues and at higher levels in salivary gland, stomach, and kidney. The ALDH6 gene was assigned to chromosome 15q26 using fluorescence in situ hybridization. 52 refs., 6 figs., 1 tab.

  20. Metabolic engineering of glycine betaine synthesis: plant betaine aldehyde dehydrogenases lacking typical transit peptides are targeted to tobacco chloroplasts where they confer betaine aldehyde resistance.

    PubMed

    Rathinasabapathi, B; McCue, K F; Gage, D A; Hanson, A D

    1994-01-01

    Certain higher plants synthesize and accumulate glycine betaine, a compound with osmoprotectant properties. Biosynthesis of glycine betaine proceeds via the pathway choline-->betaine aldehyde-->glycine betaine. Plants such as tobacco (Nicotiana tabacum L.) which do not accumulate glycine betaine lack the enzymes catalyzing both reactions. As a step towards engineering glycine betaine accumulation into a non-accumulator, spinach and sugar beet complementary-DNA sequences encoding the second enzyme of glycine-betaine synthesis (betaine aldehyde dehydrogenase, BADH, EC 1.2.1.8) were expressed in tobacco. Despite the absence of a typical transit peptide, BADH was targeted to the chloroplast in leaves of transgenic plants. Levels of extractable BADH were comparable to those in spinach and sugar beet, and the molecular weight, isoenzyme profile and Km for betaine aldehyde of the BADH enzymes from transgenic plants were the same as for native spinach or sugar beet BADH. Transgenic plants converted supplied betaine aldehyde to glycine betaine at high rates, demonstrating that they were able to transport betaine aldehyde across both the plasma membrane and the chloroplast envelope. The glycine betaine produced in this way was not further metabolized and reached concentrations similar to those in plants which accumulate glycine betaine naturally. Betaine aldehyde was toxic to non-transformed tobacco tissues whereas transgenic tissues were resistant due to detoxification of betaine aldehyde to glycine betaine. Betaine aldehyded ehydrogenase is therefore of interest as a potential selectable marker, as well as in the metabolic engineering of osmoprotectant biosynthesis.

  1. Aldehyde dehydrogenase-2 regulates nociception in rodent models of acute inflammatory pain

    PubMed Central

    Zambelli, Vanessa O.; Gross, Eric R.; Chen, Che-Hong; Gutierrez, Vanessa P.; Cury, Yara; Mochly-Rosen, Daria

    2014-01-01

    Exogenous aldehydes can cause pain in animal models, suggesting that aldehyde dehydrogenase 2 (ALDH2), which metabolizes many aldehydes, may regulate nociception. To test this hypothesis, we generated a knock-in mouse with an inactivating point mutation in ALDH2 (ALDH2*2), which is also present in human ALDH2 of ~540 million East Asians. The ALDH2*1/*2 heterozygotic mice exhibited a larger response to painful stimuli than their wild-type littermates, and this heightened nociception was inhibited by an ALDH2-selective activator (Alda-1). No effect on inflammation per se was observed. Using a rat model, we then showed that nociception tightly correlated with ALDH activity (R2=0.90) and that reduced nociception was associated with less early growth response protein 1 (EGR1) in the spinal cord and less reactive aldehyde accumulation at the insult site (including acetaldehyde and 4-hydroxynonenal). Further, acetaldehyde and formalin-induced nociceptive behavior was greater in the ALDH2*1/*2 mice than wild-type mice. Finally, Alda-1 treatment was also beneficial when given even after the inflammatory agent was administered. Our data in rodent models suggest that the mitochondrial enzyme ALDH2 regulates nociception and could serve as a molecular target for pain control, with ALDH2 activators, such as Alda-1, as potential non-narcotic cardiac-safe analgesics. Furthermore, our results suggest a possible genetic basis for East Asians’ apparent lower pain tolerance. PMID:25163478

  2. Aldehyde dehydrogenase-2 regulates nociception in rodent models of acute inflammatory pain.

    PubMed

    Zambelli, Vanessa O; Gross, Eric R; Chen, Che-Hong; Gutierrez, Vanessa P; Cury, Yara; Mochly-Rosen, Daria

    2014-08-27

    Exogenous aldehydes can cause pain in animal models, suggesting that aldehyde dehydrogenase-2 (ALDH2), which metabolizes many aldehydes, may regulate nociception. To test this hypothesis, we generated a knock-in mouse with an inactivating point mutation in ALDH2 (ALDH2*2), which is also present in human ALDH2 of ~540 million East Asians. The ALDH2*1/*2 heterozygotic mice exhibited a larger response to painful stimuli than their wild-type littermates, and this heightened nociception was inhibited by an ALDH2-selective activator (Alda-1). No effect on inflammation per se was observed. Using a rat model, we then showed that nociception tightly correlated with ALDH activity (R(2) = 0.90) and that reduced nociception was associated with less early growth response protein 1 (EGR1) in the spinal cord and less reactive aldehyde accumulation at the insult site (including acetaldehyde and 4-hydroxynonenal). Further, acetaldehyde- and formalin-induced nociceptive behavior was greater in the ALDH2*1/*2 mice than in the wild-type mice. Finally, Alda-1 treatment was even beneficial when given after the inflammatory agent was administered. Our data in rodent models suggest that the mitochondrial enzyme ALDH2 regulates nociception and could serve as a molecular target for pain control, with ALDH2 activators, such as Alda-1, as potential non-narcotic, cardiac-safe analgesics. Furthermore, our results suggest a possible genetic basis for East Asians' apparent lower pain tolerance.

  3. Rice aldehyde dehydrogenase7 is needed for seed maturation and viability.

    PubMed

    Shin, Jun-Hye; Kim, Sung-Ryul; An, Gynheung

    2009-02-01

    Aldehyde dehydrogenases (ALDHs) catalyze the irreversible oxidation of a wide range of reactive aldehydes to their corresponding carboxylic acids. Although the proteins have been studied from various organisms and at different growth stages, their roles in seed development have not been well elucidated. We obtained T-DNA insertional mutants in OsALDH7, which is remarkably inducible by oxidative and abiotic stresses. Interestingly, endosperms from the osaldh7 null mutants accumulated brown pigments during desiccation and storage. Extracts from the mutant seeds showed a maximum absorbance peak at 360 nm, the wavelength that melanoidin absorbs. Under UV light, those extracts also exhibited much stronger fluorescence than the wild type, suggesting that the pigments are melanoidin. These pigments started to accumulate in the late seed developmental stage, the time when OsALDH7 expression began to increase significantly. Purified OsALDH7 protein showed enzyme activities to malondialdehyde, acetaldehyde, and glyceraldehyde. These results suggest that OsALDH7 is involved in removing various aldehydes formed by oxidative stress during seed desiccation. The mutant seeds were more sensitive to our accelerated aging treatment and accumulated more malondialdehyde than the wild type. These data imply that OsALDH7 plays an important role in maintaining seed viability by detoxifying the aldehydes generated by lipid peroxidation.

  4. NADP-Dependent Aldehyde Dehydrogenase from Archaeon Pyrobaculum sp.1860: Structural and Functional Features

    PubMed Central

    Petrova, Tatiana E.; Artemova, Natalia V.; Boyko, Konstantin M.; Shabalin, Ivan G.; Rakitina, Tatiana V.; Polyakov, Konstantin M.; Popov, Vladimir O.

    2016-01-01

    We present the functional and structural characterization of the first archaeal thermostable NADP-dependent aldehyde dehydrogenase AlDHPyr1147. In vitro, AlDHPyr1147 catalyzes the irreversible oxidation of short aliphatic aldehydes at 60–85°С, and the affinity of AlDHPyr1147 to the NADP+ at 60°С is comparable to that for mesophilic analogues at 25°С. We determined the structures of the apo form of AlDHPyr1147 (3.04 Å resolution), three binary complexes with the coenzyme (1.90, 2.06, and 2.19 Å), and the ternary complex with the coenzyme and isobutyraldehyde as a substrate (2.66 Å). The nicotinamide moiety of the coenzyme is disordered in two binary complexes, while it is ordered in the ternary complex, as well as in the binary complex obtained after additional soaking with the substrate. AlDHPyr1147 structures demonstrate the strengthening of the dimeric contact (as compared with the analogues) and the concerted conformational flexibility of catalytic Cys287 and Glu253, as well as Leu254 and the nicotinamide moiety of the coenzyme. A comparison of the active sites of AlDHPyr1147 and dehydrogenases characterized earlier suggests that proton relay systems, which were previously proposed for dehydrogenases of this family, are blocked in AlDHPyr1147, and the proton release in the latter can occur through the substrate channel. PMID:27956891

  5. An Activator of Mutant and Wildtype Aldehyde Dehydrogenase Reduces Ischemic Damage to the Heart

    PubMed Central

    Chen, Che-Hong; Budas, Grant R.; Churchill, Eric N.; Disatnik, Marie-Helene; Hurley, Thomas D.; Mochly-Rosen, Daria

    2009-01-01

    There is substantial interest in the development of drugs that limit the extent of ischemia-induced cardiac damage caused by myocardial infarction or by certain surgical procedures. Here an unbiased proteomic search identified mitochondrial aldehyde dehydrogenase 2 (ALDH2) as an enzyme whose activation correlates with reduced ischemic heart damage in rodent models. A high-throughput screen yielded a small-molecule activator of ALDH2 (Alda-1) that, when administered to rats prior to an ischemic event, reduced infarct size by 60%, most likely through its inhibitory effect on the formation of cytotoxic aldehydes. In vitro, Alda-1 was a particularly effective activator of ALDH2*2, an inactive mutant form of the enzyme that is found in 40% of East Asian populations. Thus, pharmacologic enhancement of ALDH2 activity may be useful for patients with wildtype or mutant ALDH2 subjected to cardiac ischemia, such as during coronary bypass surgery. (140/140 words) PMID:18787169

  6. Aldehyde dehydrogenase-2 inhibition blocks remote preconditioning in experimental and human models.

    PubMed

    Contractor, Hussain; Støttrup, Nicolaj B; Cunnington, Colin; Manlhiot, Cedric; Diesch, Jonathan; Ormerod, Julian O M; Jensen, Rebekka; Bøtker, Hans Erik; Redington, Andrew; Schmidt, Michael R; Ashrafian, Houman; Kharbanda, Rajesh K

    2013-05-01

    Mitochondrial aldehyde dehydrogenase-2 (ALDH-2) is involved in preconditioning pathways, but its role in remote ischaemic preconditioning (rIPC) is unknown. We investigated its role in animal and human models of rIPC. (i) In a rabbit model of myocardial infarction, rIPC alone reduced infarct size [69 ± 5.8 % (n = 11) to 40 ± 6.5 % (n = 12), P = 0.019]. However, rIPC protection was lost after pre-treatment with the ALDH-2 inhibitor cyanamide (62 ± 7.6 % controls, n = 10, versus 61 ± 6.9 % rIPC after cyanamide, n = 10, P > 0.05). (ii) In a forearm plethysmography model of endothelial ischaemia-reperfusion injury, 24 individuals of Asian ethnic origin underwent combined rIPC and ischaemia-reperfusion (IR). 11 had wild-type (WT) enzyme and 13 carried the Glu504Lys (ALDH2*2) polymorphism (rendering ALDH-2 functionally inactive). In WT individuals, rIPC protected against impairment of response to acetylcholine (P = 0.9), but rIPC failed to protect carriers of Glu504Lys polymorphism (P = 0.004). (iii) In a second model of endothelial IR injury, 12 individuals participated in a double-blind placebo-controlled crossover study, receiving the ALDH-2 inhibitor disulfiram 600 mg od or placebo for 48 h prior to assessment of flow-mediated dilation (FMD) before and after combined rIPC and IR. With placebo, rIPC was effective with no difference in FMD before and after IR (6.18 ± 1.03 % and 4.76 ± 0.93 % P = 0.1), but disulfiram inhibited rIPC with a reduction in FMD after IR (7.87 ± 1.27 % and 3.05 ± 0.53 %, P = 0.001). This study demonstrates that ALDH-2 is involved in the rIPC pathway in three distinct rabbit and human models. This has potential implications for future clinical studies of remote conditioning.

  7. Catalysis of dehydrogenation of 4-trans-(N,N-dimethylamino)cinnamaldehyde by aldehyde dehydrogenase.

    PubMed

    Pietruszko, R; Lehmann, T; Dryjanski, M; Abriola, D P; Ambroziak, W

    2001-01-30

    4-trans-(N,N-dimethylamino)cinnamaldehyde (DACA) is a chromophoric and fluorogenic substrate of aldehyde dehydrogenase. Fluorescence of DACA is enhanced by binding to aldehyde dehydrogenase in the absence of catalysis both in the presence and absence of the coenzyme analogue 5'AMP. DACA binds to aldehyde dehydrogenase with a dissociation constant of 1-3 microM and stoichiometry of 2 mol mol(-1) enzyme. Incorporation of DACA during catalysis was also investigated and found to be 2 mol DACA mol(-1) enzyme. Effect of pH on the stoichiometry of DACA incorporation during catalysis has shown that DACA incorporation remained constant at 2 mol DACA mol(-1) enzyme, despite a 74-fold velocity enhancement between pH 5.0 and 9.0. Increase of pH increased decomposition of enzyme-acyl intermediate without affecting the rate-limiting step of the reaction. At pH 7.0 the pH stimulated velocity enhancement was 10-fold over that at pH 5.0; further velocity enhancement (11.5-fold that of pH 7.0) was achieved by 150 microM Mg(2+) ions. The velocity at pH 7.0 with Mg(2+) exceeded that of pH 9.0, and that at maximal pH stimulation at pH 9.5. It was observed that level of intermediate decreased to about 1 mol mol(-1) enzyme, indicating that Mg(2+) ions increased the rate of decomposition of the enzyme-acyl intermediate and shifted the rate-limiting step of the reaction to another step in the reaction sequence.

  8. Gene expression profiles of prostate cancer stem cells isolated by aldehyde dehydrogenase activity assay.

    PubMed

    Nishida, Sachiyo; Hirohashi, Yoshihiko; Torigoe, Toshihiko; Kitamura, Hiroshi; Takahashi, Akari; Masumori, Naoya; Tsukamoto, Taiji; Sato, Noriyuki

    2012-07-01

    Prostate cancer cells include a small population of cancer stem-like/cancer initiating cells, which have roles in cancer initiation and progression. Recently aldehyde dehydrogenase activity was used to isolate stem cells of various cancer and normal cells. We evaluated the aldehyde dehydrogenase activity of the human prostate cancer cell line 22Rv1 (ATCC®) with the ALDEFLUOR® assay and determined its potency as prostate cancer stem-like/cancer initiating cells. The human prostate cancer cell line 22Rv1 was labeled with ALDEFLUOR reagent and analyzed by flow cytometry. ALDH1(high) and ALDH1(low) cells were isolated and tumorigenicity was evaluated by xenograft transplantation into NOD/SCID mice. Tumor sphere forming ability was evaluated by culturing in a floating condition. Invasion capability was evaluated by the Matrigel™ invasion assay. Gene expression profiling was assessed by microarrays and reverse transcriptase-polymerase chain reaction. ALDH1(high) cells were detected in 6.8% of 22Rv1 cells, which showed significantly higher tumorigenicity than ALDH1(low) cells in NOD/SCID mice (p < 0.05). Gene expression profiling revealed higher expression of the stem cell related genes PROM1 and NKX3-1 in ALDH1(high) cells than in ALDH1(low) cells. ALDH1(high) cells also showed higher invasive capability and sphere forming capability than ALDH1(low) cells. Results indicate that cancer stem-like/cancer initiating cells are enriched in the ALDH1(high) population of the prostate cancer cell line 22Rv1. This approach may provide a breakthrough to further clarify prostate cancer stem-like/cancer initiating cells. To our knowledge this is the first report of cancer stem-like/cancer initiating cells of 22Rv1 using the aldehyde dehydrogenase activity assay. Copyright © 2012 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  9. Benomyl, aldehyde dehydrogenase, DOPAL, and the catecholaldehyde hypothesis for the pathogenesis of Parkinson's disease.

    PubMed

    Casida, John E; Ford, Breanna; Jinsmaa, Yunden; Sullivan, Patti; Cooney, Adele; Goldstein, David S

    2014-08-18

    The dopamine metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL) is detoxified mainly by aldehyde dehydrogenase (ALDH). We find that the fungicide benomyl potently and rapidly inhibits ALDH and builds up DOPAL in vivo in mouse striatum and in vitro in PC12 cells and human cultured fibroblasts and glial cells. The in vivo results resemble those noted previously with knockouts of the genes encoding ALDH1A1 and 2, a mouse model of aging-related Parkinson's disease (PD). Exposure to pesticides that inhibit ALDH may therefore increase PD risk via DOPAL buildup. This study lends support to the "catecholaldehyde hypothesis" that the autotoxic dopamine metabolite DOPAL plays a pathogenic role in PD.

  10. Structure of daidzin, a naturally occurring anti-alcohol-addiction agent, in complex with human mitochondrial aldehyde dehydrogenase.

    PubMed

    Lowe, Edward D; Gao, Guang-Yao; Johnson, Louise N; Keung, Wing Ming

    2008-08-14

    The ALDH2*2 gene encoding the inactive variant form of mitochondrial aldehyde dehydrogenase (ALDH2) protects nearly all carriers of this gene from alcoholism. Inhibition of ALDH2 has hence become a possible strategy to treat alcoholism. The natural product 7-O-glucosyl-4'-hydroxyisoflavone (daidzin), isolated from the kudzu vine ( Peruraria lobata), is a specific inhibitor of ALDH2 and suppresses ethanol consumption. Daidzin is the active principle in a herbal remedy for "alcohol addiction" and provides a lead for the design of improved ALDH2. The structure of daidzin/ALDH2 in complex at 2.4 A resolution shows the isoflavone moiety of daidzin binding close to the aldehyde substrate-binding site in a hydrophobic cleft and the glucosyl function binding to a hydrophobic patch immediately outside the isoflavone-binding pocket. These observations provide an explanation for both the specificity and affinity of daidzin (IC50 =80 nM) and the affinity of analogues with different substituents at the glucosyl position.

  11. Aldehyde dehydrogenase (ALDH) superfamily in plants: gene nomenclature and comparative genomics.

    PubMed

    Brocker, Chad; Vasiliou, Melpomene; Carpenter, Sarah; Carpenter, Christopher; Zhang, Yucheng; Wang, Xiping; Kotchoni, Simeon O; Wood, Andrew J; Kirch, Hans-Hubert; Kopečný, David; Nebert, Daniel W; Vasiliou, Vasilis

    2013-01-01

    In recent years, there has been a significant increase in the number of completely sequenced plant genomes. The comparison of fully sequenced genomes allows for identification of new gene family members, as well as comprehensive analysis of gene family evolution. The aldehyde dehydrogenase (ALDH) gene superfamily comprises a group of enzymes involved in the NAD(+)- or NADP(+)-dependent conversion of various aldehydes to their corresponding carboxylic acids. ALDH enzymes are involved in processing many aldehydes that serve as biogenic intermediates in a wide range of metabolic pathways. In addition, many of these enzymes function as 'aldehyde scavengers' by removing reactive aldehydes generated during the oxidative degradation of lipid membranes, also known as lipid peroxidation. Plants and animals share many ALDH families, and many genes are highly conserved between these two evolutionarily distinct groups. Conversely, both plants and animals also contain unique ALDH genes and families. Herein we carried out genome-wide identification of ALDH genes in a number of plant species-including Arabidopsis thaliana (thale crest), Chlamydomonas reinhardtii (unicellular algae), Oryza sativa (rice), Physcomitrella patens (moss), Vitis vinifera (grapevine) and Zea mays (maize). These data were then combined with previous analysis of Populus trichocarpa (poplar tree), Selaginella moellindorffii (gemmiferous spikemoss), Sorghum bicolor (sorghum) and Volvox carteri (colonial algae) for a comprehensive evolutionary comparison of the plant ALDH superfamily. As a result, newly identified genes can be more easily analyzed and gene names can be assigned according to current nomenclature guidelines; our goal is to clarify previously confusing and conflicting names and classifications that might confound results and prevent accurate comparisons between studies.

  12. Aldehyde dehydrogenase (ALDH) superfamily in plants: gene nomenclature and comparative genomics

    PubMed Central

    Brocker, Chad; Vasiliou, Melpomene; Carpenter, Sarah; Carpenter, Christopher; Zhang, Yucheng; Wang, Xiping; Kotchoni, Simeon O.; Wood, Andrew J.; Kirch, Hans-Hubert; Kopečný, David; Nebert, Daniel W.

    2012-01-01

    In recent years, there has been a significant increase in the number of completely sequenced plant genomes. The comparison of fully sequenced genomes allows for identification of new gene family members, as well as comprehensive analysis of gene family evolution. The aldehyde dehydrogenase (ALDH) gene superfamily comprises a group of enzymes involved in the NAD+- or NADP+-dependent conversion of various aldehydes to their corresponding carboxylic acids. ALDH enzymes are involved in processing many aldehydes that serve as biogenic intermediates in a wide range of metabolic pathways. In addition, many of these enzymes function as ‘aldehyde scavengers’ by removing reactive aldehydes generated during the oxidative degradation of lipid membranes, also known as lipid peroxidation. Plants and animals share many ALDH families, and many genes are highly conserved between these two evolutionarily distinct groups. Conversely, both plants and animals also contain unique ALDH genes and families. Herein we carried outgenome-wide identification of ALDH genes in a number of plant species—including Arabidopsis thaliana (thale crest), Chlamydomonas reinhardtii (unicellular algae), Oryza sativa (rice), Physcomitrella patens (moss), Vitis vinifera (grapevine) and Zea mays (maize). These data were then combined with previous analysis of Populus trichocarpa (poplar tree), Selaginella moellindorffii (gemmiferous spikemoss), Sorghum bicolor (sorghum) and Volvox carteri (colonial algae) for a comprehensive evolutionary comparison of the plant ALDH superfamily. As a result, newly identified genes can be more easily analyzed and gene names can be assigned according to current nomenclature guidelines; our goal is to clarify previously confusing and conflicting names and classifications that might confound results and prevent accurate comparisons between studies. PMID:23007552

  13. Structural insights into the production of 3-hydroxypropionic acid by aldehyde dehydrogenase from Azospirillum brasilense

    PubMed Central

    Son, Hyeoncheol Francis; Park, Sunghoon; Yoo, Tae Hyeon; Jung, Gyoo Yeol; Kim, Kyung-Jin

    2017-01-01

    3-Hydroxypropionic acid (3-HP) is an important platform chemical to be converted to acrylic acid and acrylamide. Aldehyde dehydrogenase (ALDH), an enzyme that catalyzes the reaction of 3-hydroxypropionaldehyde (3-HPA) to 3-HP, determines 3-HP production rate during the conversion of glycerol to 3-HP. To elucidate molecular mechanism of 3-HP production, we determined the first crystal structure of a 3-HP producing ALDH, α-ketoglutarate-semialdehyde dehydrogenase from Azospirillum basilensis (AbKGSADH), in its apo-form and in complex with NAD+. Although showing an overall structure similar to other ALDHs, the AbKGSADH enzyme had an optimal substrate binding site for accepting 3-HPA as a substrate. Molecular docking simulation of 3-HPA into the AbKGSADH structure revealed that the residues Asn159, Gln160 and Arg163 stabilize the aldehyde- and the hydroxyl-groups of 3-HPA through hydrogen bonds, and several hydrophobic residues, such as Phe156, Val286, Ile288, and Phe450, provide the optimal size and shape for 3-HPA binding. We also compared AbKGSADH with other reported 3-HP producing ALDHs for the crucial amino acid residues for enzyme catalysis and substrate binding, which provides structural implications on how these enzymes utilize 3-HPA as a substrate. PMID:28393833

  14. Genomic organization and expression of the human fatty aldehyde dehydrogenase gene (FALDH)

    SciTech Connect

    Rogers, G.R.; Markova, N.G.; Compton, J.G.

    1997-01-15

    Mutations in the fatty aldehyde dehydrogenase (FALDH) gene cause Sjoegren-Larsson syndrome (SLS) - a disease characterized by mental retardation, spasticity, and congenital ichthyosis. To facilitate mutation analysis in SLS and to study the pathogenesis of FALDH deficiency, we have determined the structural organization and characterized expression of the FALDH (proposed designation ALDH10) gene. The gene consists of 10 exons spanning about 30.5 kb. A TATA-less promoter is associated with the major transcription initiation site found to be 258 hp upstream of the ATG codon. The G4C-rich sequences surrounding the transcription initiation site encompassed regulatory elements that interacted with proteins in HeLa nuclear extracts and were able to promote transcription in vitro. FALDH is widely expressed as three transcripts of 2, 3.8, and 4.0 kb, which originate from multiple polyadenylation signals in the 3{prime} UTR. An alternatively spliced mRNA was detected that contains an extra exon and encodes an enzyme that is likely to have altered membrane-binding properties. The FALDH gene lies only 50-85 kb from ALDH3, an aldehyde dehydrogenase gene that has homologous sequence and intron/exon structure. 25 refs., 4 figs., 1 tab.

  15. The effect of disulfiram on the aldehyde dehydrogenases of sheep liver.

    PubMed Central

    Kitson, T M

    1975-01-01

    1. The effect of disulfiram on the activity of the cytoplasmic and mitochondrial aldehyde dehydrogenases of sheep liver was studied. 2. Disulfiram causes an immediate inhibition of the enzyme reaction. The effect on the cytoplasmic enzyme is much greater than on the mitochondrial enzyme. 3. In both cases, the initial partial inhibition is followed by a gradual irreversible loss of activity. 4. The pH-rate profile of the inactivation of the mitochondrial enzyme by disulfiram and the pH-dependence of the maximum velocity of the enzyme-catalysed reaction are both consistent with the involvement of a thiol group. 5. Excess of 2-mercaptoethanol or GSH abolishes the effect of disulfiram. However, equimolar amounts of either of these reagents and disulfiram cause an effect greater than does disulfiram alone. It was shown that the mixed disulphide, Et2N-CS-SS-CH2-CH2OH, strongly inhibits aldehyde dehydrogenase. 6. The inhibitory effect of diethyldithiocarbamate in vitro is due mainly to contamination by disulfiram. PMID:3167

  16. High Aldehyde Dehydrogenase Activity Identifies a Subset of Human Mesenchymal Stromal Cells with Vascular Regenerative Potential.

    PubMed

    Sherman, Stephen E; Kuljanin, Miljan; Cooper, Tyler T; Putman, David M; Lajoie, Gilles A; Hess, David A

    2017-03-15

    During culture expansion, multipotent mesenchymal stromal cells (MSCs) differentially express aldehyde dehydrogenase (ALDH), an intracellular detoxification enzyme that protects long-lived cells against oxidative stress. Thus, MSC selection based on ALDH-activity may be used to reduce heterogeneity and distinguish MSC subsets with improved regenerative potency. After expansion of human bone marrow-derived MSCs, cell progeny was purified based on low versus high ALDH-activity (ALDH(hi) ) by fluorescence-activated cell sorting, and each subset was compared for multipotent stromal and provascular regenerative functions. Both ALDH(l) ° and ALDH(hi) MSC subsets demonstrated similar expression of stromal cell (>95% CD73(+) , CD90(+) , CD105(+) ) and pericyte (>95% CD146(+) ) surface markers and showed multipotent differentiation into bone, cartilage, and adipose cells in vitro. Conditioned media (CDM) generated by ALDH(hi) MSCs demonstrated a potent proliferative and prosurvival effect on human microvascular endothelial cells (HMVECs) under serum-free conditions and augmented HMVEC tube-forming capacity in growth factor-reduced matrices. After subcutaneous transplantation within directed in vivo angiogenesis assay implants into immunodeficient mice, ALDH(hi) MSC or CDM produced by ALDH(hi) MSC significantly augmented murine vascular cell recruitment and perfused vessel infiltration compared with ALDH(l) ° MSC. Although both subsets demonstrated strikingly similar mRNA expression patterns, quantitative proteomic analyses performed on subset-specific CDM revealed the ALDH(hi) MSC subset uniquely secreted multiple proangiogenic cytokines (vascular endothelial growth factor beta, platelet derived growth factor alpha, and angiogenin) and actively produced multiple factors with chemoattractant (transforming growth factor-β, C-X-C motif chemokine ligand 1, 2, and 3 (GRO), C-C motif chemokine ligand 5 (RANTES), monocyte chemotactic protein 1 (MCP-1), interleukin [IL]-6, IL-8

  17. Betaine Aldehyde Dehydrogenase expression during physiological cardiac hypertrophy induced by pregnancy.

    PubMed

    Rosas-Rodríguez, Jesús Alfredo; Soñanez-Organis, José Guadalupe; Godoy-Lugo, José Arquimides; Espinoza-Salazar, Juan Alberto; López-Jacobo, Cesar Jeravy; Stephens-Camacho, Norma Aurora; González-Ochoa, Guadalupe

    2017-08-26

    Betaine Aldehyde Dehydrogenase (betaine aldehyde: NAD(P)(+) oxidoreductase, (E.C. 1.2.1.8; BADH) catalyze the irreversible oxidation of betaine aldehyde (BA) to glycine betaine (GB) and is essential for polyamine catabolism, γ-aminobutyric acid synthesis, and carnitine biosynthesis. GB is an important osmolyte that regulates the homocysteine levels, contributing to a vascular risk factor reduction. In this sense, distinct investigations describe the physiological roles of GB, but there is a lack of information about the GB novo synthesis process and regulation during cardiac hypertrophy induced by pregnancy. In this work, the BADH mRNA expression, protein level, and activity were quantified in the left ventricle before, during, and after pregnancy. The mRNA expression, protein content and enzyme activity along with GB content of BADH increased 2.41, 1.95 and 1.65-fold respectively during late pregnancy compared to not pregnancy, and returned to basal levels at postpartum. Besides, the GB levels increased 1.53-fold during pregnancy and remain at postpartum. Our results demonstrate that physiological cardiac hypertrophy induced BADH mRNA expression and activity along with GB production, suggesting that BADH participates in the adaptation process of physiological cardiac hypertrophy during pregnancy, according to the described GB role in cellular osmoregulation, osmoprotection and reduction of vascular risk. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Identification and characterization of aldehyde dehydrogenase 9 from Lampetra japonica and its protective role against cytotoxicity.

    PubMed

    Zhao, Chunhui; Wang, Dan; Feng, Bin; Gou, Meng; Liu, Xin; Li, Qingwei

    2015-09-01

    Aldehyde dehydrogenases (ALDHs), which oxidize aldehyde to corresponding acids, play a major role in the detoxification of various endogenous and exogenous aldehydes. In this study, we cloned and characterized ALDH9 (designated LjALDH9) from Arctic lamprey Lampetra japonica. The open reading frame of LjALDH9 was 1566 bp, encoding 521 amino acids with a predicted molecular mass of 55.68 kDa. LjALDH9 protein had a signal peptide and Aldedh domain with the active site Cys315. In addition, LjALDH9 shares high sequence homology with ALDH9 of jawed vertebrates. Real-time quantitative PCR revealed that LjALDH9 was highly expressed in the buccal gland. A reactive LjALDH9 protein was obtained by prokaryotic expression, two-step-denaturing and refolding and affinity purification. During enzyme activity analysis of recombinant LjALDH9, we found that the most suitable reaction conditions were pH7.0, 16-23 °C and Mn(2+) as the activator. Our study provides theoretical proof that LjALDH9 plays an important role in the parasitic life phase of lamprey.

  19. Acute and chronic ethanol exposure differentially alters alcohol dehydrogenase and aldehyde dehydrogenase activity in the zebrafish liver.

    PubMed

    Tran, Steven; Nowicki, Magda; Chatterjee, Diptendu; Gerlai, Robert

    2015-01-02

    Chronic ethanol exposure paradigms have been successfully used in the past to induce behavioral and central nervous system related changes in zebrafish. However, it is currently unknown whether chronic ethanol exposure alters ethanol metabolism in adult zebrafish. In the current study we examine the effect of acute ethanol exposure on adult zebrafish behavioral responses, as well as alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) activity in the liver. We then examine how two different chronic ethanol exposure paradigms (continuous and repeated ethanol exposure) alter behavioral responses and liver enzyme activity during a subsequent acute ethanol challenge. Acute ethanol exposure increased locomotor activity in a dose-dependent manner. ADH activity was shown to exhibit an inverted U-shaped curve and ALDH activity was decreased by ethanol exposure at all doses. During the acute ethanol challenge, animals that were continuously housed in ethanol exhibited a significantly reduced locomotor response and increased ADH activity, however, ALDH activity did not change. Zebrafish that were repeatedly exposed to ethanol demonstrated a small but significant attenuation of the locomotor response during the acute ethanol challenge but ADH and ALDH activity was similar to controls. Overall, we identified two different chronic ethanol exposure paradigms that differentially alter behavioral and physiological responses in zebrafish. We speculate that these two paradigms may allow dissociation of central nervous system-related and liver enzyme-dependent ethanol induced changes in zebrafish.

  20. The diagnostic value of alcohol dehydrogenase (ADH) isoenzymes and aldehyde dehydrogenase (ALDH) measurement in the sera of colorectal cancer patients.

    PubMed

    Jelski, Wojciech; Mroczko, Barbara; Szmitkowski, Maciej

    2010-10-01

    The activity of total alcohol dehydrogenase (ADH) and class I isoenzymes is significantly higher in colorectal cancer tissue than in healthy mucosa. The activity of these enzymes in cancer cells is probably reflected in the sera and could thus be helpful for diagnosing colorectal cancer. The aim of this study was to investigate a potential role of ADH and aldehyde dehydrogenase (ALDH) as tumor markers for colorectal cancer. We defined diagnostic sensitivity, specificity, positive and negative predictive values, and receiver-operating characteristics (ROC) curve for tested enzymes. Serum samples were taken from 182 patients with colorectal cancer before treatment and from 160 control subjects. Total ADH activity and class III and IV isoenzymes were measured by photometric, but ALDH activity and ADH I and II by the fluorometric method, with class-specific fluorogenic substrates. There was significant increase in the activity of ADH I isoenzyme and ADH total in the sera of colorectal cancer patients compared to the control. The diagnostic sensitivity for ADH I was 76%, specificity 82%, AND positive and negative predictive values were 85 and 74%, respectively. The sensitivity and specificity of ADH I increased with the stage of the carcinoma. The area under ROC curve for ADH I was 0.72. The results suggest a potential role for ADH I as marker for colorectal cancer.

  1. The activity of class I, II, III, and IV alcohol dehydrogenase isoenzymes and aldehyde dehydrogenase in endometrial cancer.

    PubMed

    Orywal, Karolina; Jelski, Wojciech; Zdrodowski, Michał; Szmitkowski, Maciej

    2010-01-01

    The metabolism of cancerous cells is in many ways different than in healthy cells. In endometrial cancer, cells exhibit activity of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), which participate in the metabolism of many biological substances. The aim of this study was to compare the metabolism of endometrial cancer cells and normal endometrial cells by measurement of ADH isoenzymes and ALDH activities in these tissues. The study material consists of cancerous endometrial tissues obtained from 34 patients. Total ADH activity was measured using the photometric method and ALDH activity using the fluorometric method. For the measurement of class I and II ADH isoenzyme activity, we employed the fluorometric method, with class-specific fluorogenic substrates. The activity of class III and IV ADH was measured using the photometric method. The activity of the class I ADH isoenzyme was significantly higher in the endometrial cancer tissues when compared with normal endometrial tissues. The other classes of ADH tested did not show significant differences between activity of cancerous cells and healthy endometrium. The activity of total ADH was also significantly higher in endometrial cancer. The increased activity of total ADH in endometrial cancer, especially the class I isoenzyme and normal activity of ALDH, may be the cause of disorders in metabolic pathways that use these isoenzymes and could increase the concentration of acetaldehyde, which is cancerogenic substance. J. Clin. Lab. Anal. 24:334-339, 2010. © 2010 Wiley-Liss, Inc.

  2. The diagnostic value of alcohol dehydrogenase (ADH) isoenzymes and aldehyde dehydrogenase (ALDH) measurement in the sera of gastric cancer patients.

    PubMed

    Jelski, Wojciech; Orywal, Karolina; Laniewska, Magdalena; Szmitkowski, Maciej

    2010-12-01

    Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are present in gastric cancer cells (GC). Moreover, the activity of total ADH and class IV isoenzymes is significantly higher in cancer tissue than in healthy mucosa. The activity of these enzymes in cancer cells is probably reflected in the sera and could thus be helpful for diagnostics of gastric cancer. The aim of this study was to investigate a potential role of ADH and ALDH as tumor markers for gastric cancer. We defined diagnostic sensitivity, specificity, predictive value for positive and negative results, and receiver-operating characteristics (ROC) curve for tested enzymes. Serum samples were taken from 168 patients with gastric cancer before treatment and from 168 control subjects. Total ADH activity and class III and IV isoenzymes were measured by photometric but ALDH activity and ADH I and II by the fluorometric method, with class-specific fluorogenic substrates. There was significant increase in the activity of ADH IV isoenzyme and ADH total in the sera of gastric cancer patients compared to the control. The diagnostic sensitivity for ADH IV was 73%, specificity 79%, positive and negative predictive values were 81 and 72% respectively. Area under ROC curve for ADH IV was 0.67. The results suggest a potential role for ADH IV as marker of gastric cancer.

  3. Ethanol utilization regulatory protein: profile alignments give no evidence of origin through aldehyde and alcohol dehydrogenase gene fusion.

    PubMed Central

    Nicholas, H. B.; Persson, B.; Jörnvall, H.; Hempel, J.

    1995-01-01

    The suggestion that the ethanol regulatory protein from Aspergillus has its evolutionary origin in a gene fusion between aldehyde and alcohol dehydrogenase genes (Hawkins AR, Lamb HK, Radford A, Moore JD, 1994, Gene 146:145-158) has been tested by profile analysis with aldehyde and alcohol dehydrogenase family profiles. We show that the degree and kind of similarity observed between these profiles and the ethanol regulatory protein sequence is that expected from random sequences of the same composition. This level of similarity fails to support the suggested gene fusion. PMID:8580855

  4. Aldehyde dehydrogenase 2 activation in heart failure restores mitochondrial function and improves ventricular function and remodelling

    PubMed Central

    Gomes, Katia M.S.; Campos, Juliane C.; Bechara, Luiz R.G.; Queliconi, Bruno; Lima, Vanessa M.; Disatnik, Marie-Helene; Magno, Paulo; Chen, Che-Hong; Brum, Patricia C.; Kowaltowski, Alicia J.; Mochly-Rosen, Daria; Ferreira, Julio C.B.

    2014-01-01

    Aims We previously demonstrated that pharmacological activation of mitochondrial aldehyde dehydrogenase 2 (ALDH2) protects the heart against acute ischaemia/reperfusion injury. Here, we determined the benefits of chronic activation of ALDH2 on the progression of heart failure (HF) using a post-myocardial infarction model. Methods and results We showed that a 6-week treatment of myocardial infarction-induced HF rats with a selective ALDH2 activator (Alda-1), starting 4 weeks after myocardial infarction at a time when ventricular remodelling and cardiac dysfunction were present, improved cardiomyocyte shortening, cardiac function, left ventricular compliance and diastolic function under basal conditions, and after isoproterenol stimulation. Importantly, sustained Alda-1 treatment showed no toxicity and promoted a cardiac anti-remodelling effect by suppressing myocardial hypertrophy and fibrosis. Moreover, accumulation of 4-hydroxynonenal (4-HNE)-protein adducts and protein carbonyls seen in HF was not observed in Alda-1-treated rats, suggesting that increasing the activity of ALDH2 contributes to the reduction of aldehydic load in failing hearts. ALDH2 activation was associated with improved mitochondrial function, including elevated mitochondrial respiratory control ratios and reduced H2O2 release. Importantly, selective ALDH2 activation decreased mitochondrial Ca2+-induced permeability transition and cytochrome c release in failing hearts. Further supporting a mitochondrial mechanism for ALDH2, Alda-1 treatment preserved mitochondrial function upon in vitro aldehydic load. Conclusions Selective activation of mitochondrial ALDH2 is sufficient to improve the HF outcome by reducing the toxic effects of aldehydic overload on mitochondrial bioenergetics and reactive oxygen species generation, suggesting that ALDH2 activators, such as Alda-1, have a potential therapeutic value for treating HF patients. PMID:24817685

  5. Fatty aldehyde dehydrogenase multigene family involved in the assimilation of n-alkanes in Yarrowia lipolytica.

    PubMed

    Iwama, Ryo; Kobayashi, Satoshi; Ohta, Akinori; Horiuchi, Hiroyuki; Fukuda, Ryouichi

    2014-11-28

    In the n-alkane assimilating yeast Yarrowia lipolytica, n-alkanes are oxidized to fatty acids via fatty alcohols and fatty aldehydes, after which they are utilized as carbon sources. Here, we show that four genes (HFD1-HFD4) encoding fatty aldehyde dehydrogenases (FALDHs) are involved in the metabolism of n-alkanes in Y. lipolytica. A mutant, in which all of four HFD genes are deleted (Δhfd1-4 strain), could not grow on n-alkanes of 12-18 carbons; however, the expression of one of those HFD genes restored its growth on n-alkanes. Production of Hfd2Ap or Hfd2Bp, translation products of transcript variants generated from HFD2 by the absence or presence of splicing, also supported the growth of the Δhfd1-4 strain on n-alkanes. The FALDH activity in the extract of the wild-type strain was increased when cells were incubated in the presence of n-decane, whereas this elevation in FALDH activity by n-decane was not observed in Δhfd1-4 strain extract. Substantial FALDH activities were detected in the extracts of Escherichia coli cells expressing the HFD genes. Fluorescent microscopic observation suggests that Hfd3p and Hfd2Bp are localized predominantly in the peroxisome, whereas Hfd1p and Hfd2Ap are localized in both the endoplasmic reticulum and the peroxisome. These results suggest that the HFD multigene family is responsible for the oxidation of fatty aldehydes to fatty acids in the metabolism of n-alkanes, and raise the possibility that Hfd proteins have diversified by gene multiplication and RNA splicing to efficiently assimilate or detoxify fatty aldehydes in Y. lipolytica. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Coupled hydroperoxide lyase and alcohol dehydrogenase for selective synthesis of aldehyde or alcohol.

    PubMed

    Gargouri, Mohamed; Akacha, Najla Ben; Legoy, Marie-Dominique

    2004-11-01

    The main objective of this work was to improve the selective synthesis of a volatile compound: aldehyde or alcohol using a coupled-enzyme system. A novel method of synthesis of C6-aldehyde or alcohol was carried out in the presence of hydroperoxide lyase (HPLS) activity coupled to alcohol dehydrogenase (ADH) activity. After cleavage of the initial substrate, hydroperoxy fatty acid catalyzed by HPLS, the second enzyme, ADH, can catalyze the reduction of the aldehyde to the corresponding alcohol, or the oxidation of contaminating alcohol into aldehyde, depending on the cofactor present in the medium (oxidized or reduced form). We succeeded in improving the synthesis of one of the products. When coupling HPLS to NADP, the selectivity of hexanal production from 13-hydroperoxy linoleic acid was improved, and hexanol production was reduced 5 to 10 times after 15 min of reaction at 15 degrees C and pH 7.0. In another experiment, HPLS was coupled to ADH in the presence of NADH. The production of alcohol (hexenols) was then favored especially when using 13-hydroperoxy linolenic acid as substrate at concentrations >15 mM, reaching 95% of the products. Coupling of the enzymatic reactions (cleavage reduction) not only reduced the number of steps but also allowed us to increase the conversion rate of the initial substrate (hydroperoxy fatty acid). Structures of the compounds produced in this work were confirmed using gas chromatography-mass spectroscopy analysis. Each of these products has its own delicately different fresh odor that can be used in various applications.

  7. A novel NADPH-dependent aldehyde reductase gene from Saccharomyces cerevisiae NRRL Y-12632 involved in the detoxification of aldehyde inhibitors derived from lignocellulosic biomass conversion.

    PubMed

    Liu, Z Lewis; Moon, Jaewoong

    2009-10-01

    Aldehyde inhibitors such as furfural, 5-hydroxymethylfurfural, anisaldehyde, benzaldehyde, cinnamaldehyde, and phenylaldehyde are commonly generated during lignocellulosic biomass conversion process for low-cost cellulosic ethanol production that interferes with subsequent microbial growth and fermentation. In situ detoxification of the aldehyde inhibitors is possible by the tolerant ethanologenic yeast that involves multiple genes including numerous functional reductases. In this study, we report a novel aldehyde reductase gene clone Y63 from ethanologenic yeast Saccharomyces cerevisiae NRRL Y12632, representing the uncharacterized ORF YGL157W, which demonstrated NADPH-dependent reduction activities toward at least 14 aldehyde substrates. The identity of gene clone Y63 is the same with YGL157W of SGD since a variation of only 35 nucleotides in genomic sequence and three amino acid residues were observed between the two that share the same length of 347 residues in size. As one among the highly induced genes, YGL157W of Y-12632 showed significantly high levels of transcript abundance in response to furfural and HMF challenges. Based on the deduced amino acid sequence and the most conserved functional motif analyses including closely related reductases from five other yeast species to this date, YGL157W was identified as a member of the subclass 'intermediate' of the SDR (short-chain dehydrogenase/reductase) superfamily with the following typical characteristics: the most conserved catalytic site to lie at Tyr(169)-X-X-X-Lys(173); an indispensable reduction catalytic triad at Ser(131), Tyr(169), and Lys(173), and an approved cofactor-binding motif at Gly(11)-X-X-Gly(14)-X-X-Ala(17) near the N-terminus. YGL039W, YDR541C, and YOL151W (GRE2) appeared to be the similar type of enzymes falling into the same category of the intermediate subfamily.

  8. Aldehyde dehydrogenase 2 activation and coevolution of its εPKC-mediated phosphorylation sites.

    PubMed

    Nene, Aishwarya; Chen, Che-Hong; Disatnik, Marie-Hélène; Cruz, Leslie; Mochly-Rosen, Daria

    2017-01-05

    Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is a key enzyme for the metabolism of many toxic aldehydes such as acetaldehyde, derived from alcohol drinking, and 4HNE, an oxidative stress-derived lipid peroxidation aldehyde. Post-translational enhancement of ALDH2 activity can be achieved by serine/threonine phosphorylation by epsilon protein kinase C (εPKC). Elevated ALDH2 is beneficial in reducing injury following myocardial infarction, stroke and other oxidative stress and aldehyde toxicity-related diseases. We have previously identified three εPKC phosphorylation sites, threonine 185 (T185), serine 279 (S279) and threonine 412 (T412), on ALDH2. Here we further characterized the role and contribution of each phosphorylation site to the enhancement of enzymatic activity by εPKC. Each individual phosphorylation site was mutated to a negatively charged amino acid, glutamate, to mimic a phosphorylation, or to a non-phosphorylatable amino acid, alanine. ALDH2 enzyme activities and protection against 4HNE inactivation were measured in the presence or absence of εPKC phosphorylation in vitro. Coevolution of ALDH2 and its εPKC phosphorylation sites was delineated by multiple sequence alignments among a diverse range of species and within the ALDH multigene family. We identified S279 as a critical εPKC phosphorylation site in the activation of ALDH2. The critical catalytic site, cysteine 302 (C302) of ALDH2 is susceptible to adduct formation by reactive aldehyde, 4HNE, which readily renders the enzyme inactive. We show that phosphomimetic mutations of T185E, S279E and T412E confer protection of ALDH2 against 4HNE-induced inactivation, indicating that phosphorylation on these three sites by εPKC likely also protects the enzyme against reactive aldehydes. Finally, we demonstrate that the three ALDH2 phosphorylation sites co-evolved with εPKC over a wide range of species. Alignment of 18 human ALDH isozymes, indicates that T185 and S279 are unique ALDH2, ε

  9. Class 2 aldehyde dehydrogenase. Characterization of the hamster enzyme, sensitive to daidzin and conserved within the family of multiple forms.

    PubMed

    Hjelmqvist, L; Lundgren, R; Norin, A; Jörnvall, H; Vallee, B; Klyosov, A; Keung, W M

    1997-10-13

    Mitochondrial (class 2) hamster aldehyde dehydrogenase has been purified and characterized. Its primary structure has been determined and correlated with the tertiary structure recently established for this class from another species. The protein is found to represent a constant class within a complex family of multiple forms. Variable segments that occur in different species correlate with non-functional segments, in the same manner as in the case of the constant class of alcohol dehydrogenases (class III type) of another protein family, but distinct from the pattern of the corresponding variable enzymes. Hence, in both these protein families, overall variability and segment architectures behave similarly, with at least one 'constant' form in each case, class III in the case of alcohol dehydrogenases, and at least class 2 in the case of aldehyde dehydrogenases.

  10. Sodium borohydride removes aldehyde inhibitors for enhancing biohydrogen fermentation.

    PubMed

    Lin, Richen; Cheng, Jun; Ding, Lingkan; Song, Wenlu; Zhou, Junhu; Cen, Kefa

    2015-12-01

    To enhance biohydrogen production from glucose and xylose in the presence of aldehyde inhibitors, reducing agent (i.e., sodium borohydride) was in situ added for effective detoxification. The detoxification efficiencies of furfural (96.7%) and 5-hydroxymethylfurfural (5-HMF, 91.7%) with 30mM NaBH4 were much higher than those of vanillin (77.3%) and syringaldehyde (69.3%). Biohydrogen fermentation was completely inhibited without detoxification, probably because of the consumption of nicotinamide adenine dinucleotide (NADH) by inhibitors reduction (R-CHO+2NADH→R-CH2OH+2NAD(+)). Addition of 30mM NaBH4 provided the reducing power necessary for inhibitors reduction (4R-CHO+NaBH4+2H2O→4R-CH2OH+NaBO2). The recovered reducing power in fermentation resulted in 99.3% recovery of the hydrogen yield and 64.6% recovery of peak production rate. Metabolite production and carbon conversion after detoxification significantly increased to 63.7mM and 81.9%, respectively. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Herbicidal Activity of an Isopropylmalate Dehydrogenase Inhibitor.

    PubMed Central

    Wittenbach, V. A.; Teaney, P. W.; Hanna, W. S.; Rayner, D. R.; Schloss, J. V.

    1994-01-01

    Isopropylmalate dehydrogenase (IPMDH) is the third enzyme specific to leucine biosynthesis. It catalyzes the oxidative decarboxylation of 3-isopropylmalate (3-IPM) to 2-ketoisocaproic acid. The partially purified enzyme from pea (Pisum sativum L.) shows a broad pH optimum of 7.8 to 9.1 and has Km values for 3-IPM and NAD of 18 and 40 [mu]M, respectively. O-Isobutenyl oxalylhydroxamate (O-IbOHA) has been discovered to be an excellent inhibitor of the pea IPMDH, with an apparent inhibitor constant of 5 nM. As an herbicide, O-IbOHA showed only moderate activity on a variety of broadleaf and grass species. We characterized the herbicidal activity of O-IbOHA on corn (Zea mays L.), a sensitive species; giant foxtail (Setaria faberi) and morning glory (Ipomoea purpurea [L.] Roth), moderately tolerant species; and soybean [Glycine max L. Merr.), a tolerant species. Differences in tolerance among the species were not due to differences in the sensitivity of IPMDH. Studies with [14C]O-IbOHA suggested that uptake and translocation were not major limitations for herbicidal activity, nor were they determinants of tolerance. Moreover, metabolism could not account for the difference in tolerance of corn, foxtail, and morning glory, although it might account for the tolerance of soybean. Herbicidal activity on all four species was correlated with the accumulation of 3-IPM in the plants. PMID:12232331

  12. GRE2 from Scheffersomyces stipitis as an aldehyde reductase contributes tolerance to aldehyde inhibitors derived from lignocellulosic biomass

    USDA-ARS?s Scientific Manuscript database

    Scheffersomyces (Pichia) stipitis is one of the most promising yeasts for industrial bioethanol production from lignocellulosic biomass. S. stipitis is able to in situ detoxify aldehyde inhibitors [such as furfural and 5-hydroxymethylfurfural (HMF)] to less toxic corresponding alcohols. However, the...

  13. Alcohol and aldehyde dehydrogenases: structures of the human liver enzymes, functional properties and evolutionary aspects.

    PubMed

    Jörnvall, H; Hempel, J; von Bahr-Lindström, H; Höög, J O; Vallee, B L

    1987-01-01

    polyol dehydrogenases are encountered. The two isozymes of human aldehyde dehydrogenase also exhibit considerable differences, with only 68% structural identity. The results show an early divergence into isozymes before the man/horse species radiation. Cys-302 is a functionally important residue and is located in one of the regions with conserved hydrophobic properties. Other regions with large differences in hydropathic properties may explain the absence of cross-hybridizing isozyme forms of human liver aldehyde dehydrogenase.

  14. Cloning and molecular evolution of the aldehyde dehydrogenase 2 gene (Aldh2) in bats (Chiroptera).

    PubMed

    Chen, Yao; Shen, Bin; Zhang, Junpeng; Jones, Gareth; He, Guimei

    2013-02-01

    Old World fruit bats (Pteropodidae) and New World fruit bats (Phyllostomidae) ingest significant quantities of ethanol while foraging. Mitochondrial aldehyde dehydrogenase (ALDH2, encoded by the Aldh2 gene) plays an important role in ethanol metabolism. To test whether the Aldh2 gene has undergone adaptive evolution in frugivorous and nectarivorous bats in relation to ethanol elimination, we sequenced part of the coding region of the gene (1,143 bp, ~73 % coverage) in 14 bat species, including three Old World fruit bats and two New World fruit bats. Our results showed that the Aldh2 coding sequences are highly conserved across all bat species we examined, and no evidence of positive selection was detected in the ancestral branches leading to Old World fruit bats and New World fruit bats. Further research is needed to determine whether other genes involved in ethanol metabolism have been the targets of positive selection in frugivorous and nectarivorous bats.

  15. Dual coenzyme activities of high-Km aldehyde dehydrogenase from rat liver mitochondria.

    PubMed

    Tsai, C S; Senior, D J

    1990-04-01

    Various kinetic approaches were carried out to investigate kinetic attributes for the dual coenzyme activities of mitochondrial aldehyde dehydrogenase from rat liver. The enzyme catalyses NAD(+)- and NADP(+)-dependent oxidations of ethanal by an ordered bi-bi mechanism with NAD(P)+ as the first reactant bound and NAD(P)H as the last product released. The two coenzymes presumably interact with the kinetically identical site. NAD+ forms the dynamic binary complex with the enzyme, while the enzyme-NAD(P)H complex formation is associated with conformation change(s). A stopped-flow burst of NAD(P)H formation, followed by a slower steady-state turnover, suggests that either the deacylation or the release of NAD(P)H is rate limiting. Although NADP+ is reduced by a faster burst rate, NAD+ is slightly favored as the coenzyme by virtue of its marginally faster turnover rate.

  16. Azotobacter vinelandii Aldehyde Dehydrogenase Regulated by ς54: Role in Alcohol Catabolism and Encystment

    PubMed Central

    Gama-Castro, Socorro; Núñez, Cinthia; Segura, Daniel; Moreno, Soledad; Guzmán, Josefina; Espín, Guadalupe

    2001-01-01

    Encystment in Azotobacter vinelandii is induced by n-butanol or β-hydroxybutyrate (BHB). We identified a gene, encoding an aldehyde dehydrogenase, that was named aldA. An aldA mutation impaired bacterial growth on n-butanol, ethanol, or hexanol as the sole carbon source. Expression of aldA increased in cells shifted from sucrose to n-butanol and was shown to be dependent on the alternative ς54 factor. A mutation in rpoN encoding the ς54 factor also impaired growth on alcohols. Encystment on n-butanol, but not on BHB, was impaired in aldA or rpoN mutants, indicating that n-butanol is not an inducer of encystment by itself but must be catabolized in order to induce encystment. PMID:11591659

  17. Purification of a heterodimeric betaine aldehyde dehydrogenase from wild amaranth plants subjected to water deficit.

    PubMed

    Figueroa-Soto, C G; Valenzuela-Soto, E M

    2001-07-27

    Betaine aldehyde dehydrogenase was purified to homogeneity from wild-type amaranth plants subjected to water deficit. The enzyme has a native molecular mass of 125 kDa; it is formed by two subunits, one of the subunits with a molecular mass of 63 kDa and the second one of 70 kDa as determined by SDS-PAGE and double dimension electrophoresis. IEF studies showed two bands with pI values of 4.93 and 4.85, respectively. Possible glycosilation of the 63- and 70-kDa subunits were tested with negative results. Both subunits cross-reacted strongly with polyclonal antibody raised against porcine kidney BADH. Also antiserum rose against HSP70 cross-reacted strongly with the wild amaranth BADH 70-kDa subunit. The enzyme was stable to extreme pH's and temperatures, and high KCl concentrations. Product inhibition of BADH was not observed.

  18. Aldehyde Dehydrogenase 2 Has Cardioprotective Effects on Myocardial Ischaemia/Reperfusion Injury via Suppressing Mitophagy

    PubMed Central

    Ji, Wenqing; Wei, Shujian; Hao, Panpan; Xing, Junhui; Yuan, Qiuhuan; Wang, Jiali; Xu, Feng; Chen, Yuguo

    2016-01-01

    Mitophagy, a selective form of autophagy, is excessively activated in myocardial ischemia/reperfusion (I/R). The study investigated whether aldehyde dehydrogenase 2 (ALDH2) exerted its cardioprotective effect by regulating mitophagy. Myocardial infarct size and apoptosis after I/R in rats were ameliorated by Alda-1, an ALDH2 activator, and aggravated by ALDH2 inhibition. Both in I/R rats and hypoxia/reoxygenation H9C2 cells, ALDH2 activation suppressed phosphatase and tensin homolog-induced putative kinase 1 (PINK1)/Parkin expression, regulating mitophagy, by preventing 4-hydroxynonenal, reactive oxygen species and mitochondrial superoxide accumulation. Furthermore, the effect was enhanced by ALDH2 inhibition. Thus, ALDH2 may protect hearts against I/R injury by suppressing PINK1/Parkin–dependent mitophagy. PMID:27148058

  19. A Personalized Medicine Approach for Asian Americans with the Aldehyde Dehydrogenase 2*2 Variant

    PubMed Central

    Gross, Eric R.; Zambelli, Vanessa O.; Small, Bryce A.; Ferreira, Julio C.B.; Chen, Che-Hong; Mochly-Rosen, Daria

    2015-01-01

    Asian Americans are one of the fastest-growing populations in the United States. A relatively large subset of this population carries a unique loss-of-function point mutation in aldehyde dehydrogenase 2 (ALDH2), ALDH2*2. Found in approximately 560 million people of East Asian descent, ALDH2*2 reduces enzymatic activity by approximately 60% to 80% in heterozygotes. Furthermore, this variant is associated with a higher risk for several diseases affecting many organ systems, including a particularly high incidence relative to the general population of esophageal cancer, myocardial infarction, and osteoporosis. In this review, we discuss the pathophysiology associated with the ALDH2*2 variant, describe why this variant needs to be considered when selecting drug treatments, and suggest a personalized medicine approach for Asian American carriers of this variant. We also discuss future clinical and translational perspectives regarding ALDH2*2 research. PMID:25292432

  20. The effects of molybate, tungstate and lxd on aldehyde oxidase and xanthine dehydrogenase in Drosophila melanogaster.

    PubMed

    Bentley, M M; Williamson, J H; Oliver, M J

    1981-01-01

    The effects of dietary sodium molybdate and sodium tungstate on eye color and aldehyde oxidase and xanthine dehydrogenase activities have been determined in Drosophila melanogaster. Dietary sodium tungstate administration has been used as a screening procedure to identify two new lxd alleles. Tungstate administration results in increased frequencies of "brown-eyed" flies in lxd stocks and a coordinate decrease in AO and XDH activities in all genotypes tested. The two new lxd alleles affect AO and XDH in a qualitatively but not quantitatively similar fashion to the original lxd allele. AO and XDH activity and AO-CRM levels appear much more sensitive to mutational perturbations of this gene-enzyme than do XDH-CRM levels in the genotypes tested.

  1. Aldehyde dehydrogenase activity in cancer stem cells from canine mammary carcinoma cell lines.

    PubMed

    Michishita, M; Akiyoshi, R; Suemizu, H; Nakagawa, T; Sasaki, N; Takemitsu, H; Arai, T; Takahashi, K

    2012-08-01

    Increasing evidence suggests that diverse solid tumours arise from a small population of cells known as cancer stem cells or tumour-initiating cells. Cancer stem cells in several solid tumours are enriched for aldehyde dehydrogenase (ALDH) activity. High levels of ALDH activity (ALDH(high)) were detected in four cell lines derived from canine mammary carcinomas. ALDH(high) cells were enriched in a CD44(+)CD24(-) population having self-renewal capacity. Xenotransplantation into immunodeficient mice demonstrated that 1×10(4) ALDH(high) cells were sufficient for tumour formation in all injected mice, whereas 1×10(4) ALDH(low) cells failed to initiate any tumours. ALDH(high)-derived tumours contained both ALDH(+) and ALDH(-) cells, indicating that these cells had cancer stem cell-like properties.

  2. Influence of fermentation conditions on specific activity of the enzymes alcohol and aldehyde dehydrogenase from yeasts.

    PubMed

    Mauricio, J C; Ortega, J M

    1993-01-01

    The effects of anaerobic, semi-aerobic and short aeration fermentation conditions and the addition of ergosterol and oleic acid to musts on the specific activity of alcohol and aldehyde dehydrogenase (ADH and ALDH) from two yeast species, Saccharomyces cerevisiae and Torulaspora delbrueckii, were studied. ADH I biosynthesis only occurred during the first few hours of fermentation. ADH II from S. cerevisiae and ALDH-NADP+ from the two yeast species behaved as constitutive enzymes under all fermentation conditions. ADH II from T. delbrueckii was only synthesized in small amounts, and its activity was always lower than in S. cerevisiae, where it was responsible for the termination of alcoholic fermentation during the steady growth phase.

  3. Structure and mechanism of benzaldehyde dehydrogenase from Pseudomonas putida ATCC 12633, a member of the Class 3 aldehyde dehydrogenase superfamily.

    PubMed

    Zahniser, Megan P D; Prasad, Shreenath; Kneen, Malea M; Kreinbring, Cheryl A; Petsko, Gregory A; Ringe, Dagmar; McLeish, Michael J

    2017-03-01

    Benzaldehyde dehydrogenase from Pseudomonas putida (PpBADH) belongs to the Class 3 aldehyde dehydrogenase (ALDH) family. The Class 3 ALDHs are unusual in that they are generally dimeric (rather than tetrameric), relatively non-specific and utilize both NAD+ and NADP+. To date, X-ray structures of three Class 3 ALDHs have been determined, of which only two have cofactor bound, both in the NAD+ form. Here we report the crystal structure of PpBADH in complex with NADP+ and a thioacyl intermediate adduct. The overall architecture of PpBADH resembles that of most other members of the ALDH superfamily, and the cofactor binding residues are well conserved. Conversely, the pattern of cofactor binding for the rat Class 3 ALDH differs from that of PpBADH and other ALDHs. This has been interpreted in terms of a different mechanism for the rat enzyme. Comparison with the PpBADH structure, as well as multiple sequence alignments, suggest that one of two conserved glutamates, at positions 215 (209 in rat) and 337 (333 in rat), would act as the general base necessary to hydrolyze the thioacyl intermediate. While the latter is the general base in the rat Class 3 ALDH, site-specific mutagenesis indicates that Glu215 is the likely candidate for PpBADH, a result more typical of the Class 1 and 2 ALDH families. Finally, this study shows that hydride transfer is not rate limiting, lending further credence to the suggestion that PpBADH is more similar to the Class 1 and 2 ALDHs than it is to other Class 3 ALDHs. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  4. Inhibition of telomerase activity preferentially targets aldehyde dehydrogenase-positive cancer stem-like cells in lung cancer

    PubMed Central

    2011-01-01

    Background Mortality rates for advanced lung cancer have not declined for decades, even with the implementation of novel chemotherapeutic regimens or the use of tyrosine kinase inhibitors. Cancer Stem Cells (CSCs) are thought to be responsible for resistance to chemo/radiotherapy. Therefore, targeting CSCs with novel compounds may be an effective approach to reduce lung tumor growth and metastasis. We have isolated and characterized CSCs from non-small cell lung cancer (NSCLC) cell lines and measured their telomerase activity, telomere length, and sensitivity to the novel telomerase inhibitor MST312. Results The aldehyde dehydrogenase (ALDH) positive lung cancer cell fraction is enriched in markers of stemness and endowed with stem cell properties. ALDH+ CSCs display longer telomeres than the non-CSC population. Interestingly, MST312 has a strong antiproliferative effect on lung CSCs and induces p21, p27 and apoptosis in the whole tumor population. MST312 acts through activation of the ATM/pH2AX DNA damage pathway (short-term effect) and through decrease in telomere length (long-term effect). Administration of this telomerase inhibitor (40 mg/kg) in the H460 xenograft model results in significant tumor shrinkage (70% reduction, compared to controls). Combination therapy consisting of irradiation (10Gy) plus administration of MST312 did not improve the therapeutic efficacy of the telomerase inhibitor alone. Treatment with MST312 reduces significantly the number of ALDH+ CSCs and their telomeric length in vivo. Conclusions We conclude that antitelomeric therapy using MST312 mainly targets lung CSCs and may represent a novel approach for effective treatment of lung cancer. PMID:21827695

  5. Evidence for two-step binding of reduced nicotinamide-adenine dinucleotide to aldehyde dehydrogenase.

    PubMed Central

    MacGibbon, A K; Buckley, P D; Blackwell, L F

    1977-01-01

    The displacement of NADH from cytoplasmic aldehyde dehydrogenase (EC 1.2.1.3) from sheep liver was studied by using NAD+, 1,10-phenanthroline, ADP-ribose, deamino-NAD+ and pyridine-3-aldehyde-adenine dinucleotide as displacing agents, by following the decrease in fluorescence as a function of time. The data obtained could be fitted by assuming two first-order processes were occurring, a faster process with an apparent rate constant of 0.85 +/- 0.20 s-1 and a relative amplitude of 60 +/- 10% and a slower process with an apparent rate constant of 0.20 +/- 0.05 s-1 and a relative amplitude of 40 +/- 10% (except for pyridine-3-aldehyde-adenine dinucleotide, where the apparent rate constant for the slow process was 0.05 s-1). The displacement rates did not change significantly when the pH was varied from 6.0 to 9.0. Kinetic data are also reported for the dependence of the rate of binding of NADH to the enzyme on the total concentration of NADH. Detailed arguments are presented based on the isolation and purification procedures, the equilibrium coenzyme-binding studies and the kinetic data, which lead to the following model for the release of NADH from the enzyme: (formula: see article). The parameters that best fit the data are: k + 1 = 0.2 s-1; k - 1 = 0.05 s-1; k + 2 = 0.8 s-1 and k - 2 = 5 X 10(5)litre-mol-1-s-1. The slow phase of the NADH release is similar to the steady-state turnover number for substrates such as acetaldehyde and propionaldehyde and appears to contribute significantly to the limitation of the steady-state rate. Images Fig. 1. Fig. 2. Fig. 3. Fig. 5. PMID:21657

  6. Alda-1 is an agonist and chemical chaperone for the common human aldehyde dehydrogenase 2 variant

    SciTech Connect

    Perez-Miller, Samantha; Younus, Hina; Vanam, Ram; Chen, Che-Hong; Mochly-Rosen, Daria; Hurley, Thomas D.

    2010-04-19

    In approximately one billion people, a point mutation inactivates a key detoxifying enzyme, aldehyde dehydrogenase (ALDH2). This mitochondrial enzyme metabolizes toxic biogenic and environmental aldehydes, including the endogenously produced 4-hydroxynonenal (4HNE) and the environmental pollutant acrolein, and also bioactivates nitroglycerin. ALDH2 is best known, however, for its role in ethanol metabolism. The accumulation of acetaldehyde following the consumption of even a single alcoholic beverage leads to the Asian alcohol-induced flushing syndrome in ALDH2*2 homozygotes. The ALDH2*2 allele is semidominant, and heterozygotic individuals show a similar but less severe phenotype. We recently identified a small molecule, Alda-1, that activates wild-type ALDH2 and restores near-wild-type activity to ALDH2*2. The structures of Alda-1 bound to ALDH2 and ALDH2*2 reveal how Alda-1 activates the wild-type enzyme and how it restores the activity of ALDH2*2 by acting as a structural chaperone.

  7. Genome-Wide Identification and Functional Classification of Tomato (Solanum lycopersicum) Aldehyde Dehydrogenase (ALDH) Gene Superfamily

    PubMed Central

    Lopez-Valverde, Francisco J.; Robles-Bolivar, Paula; Lima-Cabello, Elena; Gachomo, Emma W.; Kotchoni, Simeon O.

    2016-01-01

    Aldehyde dehydrogenases (ALDHs) is a protein superfamily that catalyzes the oxidation of aldehyde molecules into their corresponding non-toxic carboxylic acids, and responding to different environmental stresses, offering promising genetic approaches for improving plant adaptation. The aim of the current study is the functional analysis for systematic identification of S. lycopersicum ALDH gene superfamily. We performed genome-based ALDH genes identification and functional classification, phylogenetic relationship, structure and catalytic domains analysis, and microarray based gene expression. Twenty nine unique tomato ALDH sequences encoding 11 ALDH families were identified, including a unique member of the family 19 ALDH. Phylogenetic analysis revealed 13 groups, with a conserved relationship among ALDH families. Functional structure analysis of ALDH2 showed a catalytic mechanism involving Cys-Glu couple. However, the analysis of ALDH3 showed no functional gene duplication or potential neo-functionalities. Gene expression analysis reveals that particular ALDH genes might respond to wounding stress increasing the expression as ALDH2B7. Overall, this study reveals the complexity of S. lycopersicum ALDH gene superfamily and offers new insights into the structure-functional features and evolution of ALDH gene families in vascular plants. The functional characterization of ALDHs is valuable and promoting molecular breeding in tomato for the improvement of stress tolerance and signaling. PMID:27755582

  8. Genome-Wide Identification and Functional Classification of Tomato (Solanum lycopersicum) Aldehyde Dehydrogenase (ALDH) Gene Superfamily.

    PubMed

    Jimenez-Lopez, Jose C; Lopez-Valverde, Francisco J; Robles-Bolivar, Paula; Lima-Cabello, Elena; Gachomo, Emma W; Kotchoni, Simeon O

    2016-01-01

    Aldehyde dehydrogenases (ALDHs) is a protein superfamily that catalyzes the oxidation of aldehyde molecules into their corresponding non-toxic carboxylic acids, and responding to different environmental stresses, offering promising genetic approaches for improving plant adaptation. The aim of the current study is the functional analysis for systematic identification of S. lycopersicum ALDH gene superfamily. We performed genome-based ALDH genes identification and functional classification, phylogenetic relationship, structure and catalytic domains analysis, and microarray based gene expression. Twenty nine unique tomato ALDH sequences encoding 11 ALDH families were identified, including a unique member of the family 19 ALDH. Phylogenetic analysis revealed 13 groups, with a conserved relationship among ALDH families. Functional structure analysis of ALDH2 showed a catalytic mechanism involving Cys-Glu couple. However, the analysis of ALDH3 showed no functional gene duplication or potential neo-functionalities. Gene expression analysis reveals that particular ALDH genes might respond to wounding stress increasing the expression as ALDH2B7. Overall, this study reveals the complexity of S. lycopersicum ALDH gene superfamily and offers new insights into the structure-functional features and evolution of ALDH gene families in vascular plants. The functional characterization of ALDHs is valuable and promoting molecular breeding in tomato for the improvement of stress tolerance and signaling.

  9. Complex, unusual conformational changes in kidney betaine aldehyde dehydrogenase suggested by chemical modification with disulfiram.

    PubMed

    Ayala-Castro, Hector G; Valenzuela-Soto, Elisa M; Figueroa-Soto, Ciria G; Muñoz-Clares, Rosario A

    2007-12-15

    The NAD+-dependent animal betaine aldehyde dehydrogenases participate in the biosynthesis of glycine betaine and carnitine, as well as in polyamines catabolism. We studied the kinetics of inactivation of the porcine kidney enzyme (pkBADH) by the drug disulfiram, a thiol-reagent, with the double aim of exploring the enzyme dynamics and investigating whether it could be an in vivo target of disulfiram. Both inactivation by disulfiram and reactivation by reductants were biphasic processes with equal limiting amplitudes. Under certain conditions half of the enzyme activity became resistant to disulfiram inactivation. NAD+ protected almost 100% at 10 microM but only 50% at 5mM, and vice versa if the enzyme was pre-incubated with NAD+ before the chemical modification. NADH, betaine aldehyde, and glycine betaine also afforded greater protection after pre-incubation with the enzyme than without pre-incubation. Together, these findings suggest two kinds of active sites in this seemingly homotetrameric enzyme, and complex, unusual ligand-induced conformational changes. In addition, they indicate that, in vivo, pkBADH is most likely protected against disulfiram inactivation.

  10. Cloning and characterization of a novel betaine aldehyde dehydrogenase gene from Suaeda corniculata.

    PubMed

    Wang, F W; Wang, M L; Guo, C; Wang, N; Li, X W; Chen, H; Dong, Y Y; Chen, X F; Wang, Z M; Li, H Y

    2016-06-20

    Glycine betaine is an important quaternary ammonium compound that is produced in response to several abiotic stresses in many organisms. The synthesis of glycine betaine requires the catalysis of betaine aldehyde dehydrogenase (BADH), which can convert betaine aldehyde into glycine betaine in plants, especially in halotolerant plants. In this study, we isolated the full-length cDNA of BADH from Suaeda corniculata (ScBADH) using reverse transcriptase-polymerase chain reaction and rapid amplification of cDNA ends. Next, we analyzed the expression profile of ScBADH using real-time PCR. The results showed that ScBADH expression was induced in the roots, stems, and leaves of S. corniculata seedlings under salt and drought stress. Next, ScBADH was overexpressed in Arabidopsis, resulting in the transgenic plants exhibiting enhanced tolerance over wild-type plants under salt and drought stress. We then analyzed the levels of glycine betaine and proline, as well as superoxide dismutase (SOD) activity, during salt stress in WT and transgenic Arabidopsis. The results indicated that overexpression of ScBADH produced more glycine betaine and proline, and increased SOD activity under NaCl treatment. Our results suggest that ScBADH might be a positive regulator in plants during the response to NaCl.

  11. Structure-based mutational studies of substrate inhibition of betaine aldehyde dehydrogenase BetB from Staphylococcus aureus.

    PubMed

    Chen, Chao; Joo, Jeong Chan; Brown, Greg; Stolnikova, Ekaterina; Halavaty, Andrei S; Savchenko, Alexei; Anderson, Wayne F; Yakunin, Alexander F

    2014-07-01

    Inhibition of enzyme activity by high concentrations of substrate and/or cofactor is a general phenomenon demonstrated in many enzymes, including aldehyde dehydrogenases. Here we show that the uncharacterized protein BetB (SA2613) from Staphylococcus aureus is a highly specific betaine aldehyde dehydrogenase, which exhibits substrate inhibition at concentrations of betaine aldehyde as low as 0.15 mM. In contrast, the aldehyde dehydrogenase YdcW from Escherichia coli, which is also active against betaine aldehyde, shows no inhibition by this substrate. Using the crystal structures of BetB and YdcW, we performed a structure-based mutational analysis of BetB and introduced the YdcW residues into the BetB active site. From a total of 32 mutations, those in five residues located in the substrate binding pocket (Val288, Ser290, His448, Tyr450, and Trp456) greatly reduced the substrate inhibition of BetB, whereas the double mutant protein H448F/Y450L demonstrated a complete loss of substrate inhibition. Substrate inhibition was also reduced by mutations of the semiconserved Gly234 (to Ser, Thr, or Ala) located in the BetB NAD(+) binding site, suggesting some cooperativity between the cofactor and substrate binding sites. Substrate docking analysis of the BetB and YdcW active sites revealed that the wild-type BetB can bind betaine aldehyde in both productive and nonproductive conformations, whereas only the productive binding mode can be modeled in the active sites of YdcW and the BetB mutant proteins with reduced substrate inhibition. Thus, our results suggest that the molecular mechanism of substrate inhibition of BetB is associated with the nonproductive binding of betaine aldehyde.

  12. A bacterial aromatic aldehyde dehydrogenase critical for the efficient catabolism of syringaldehyde

    PubMed Central

    Kamimura, Naofumi; Goto, Takayuki; Takahashi, Kenji; Kasai, Daisuke; Otsuka, Yuichiro; Nakamura, Masaya; Katayama, Yoshihiro; Fukuda, Masao; Masai, Eiji

    2017-01-01

    Vanillin and syringaldehyde obtained from lignin are essential intermediates for the production of basic chemicals using microbial cell factories. However, in contrast to vanillin, the microbial conversion of syringaldehyde is poorly understood. Here, we identified an aromatic aldehyde dehydrogenase (ALDH) gene responsible for syringaldehyde catabolism from 20 putative ALDH genes of Sphingobium sp. strain SYK-6. All these genes were expressed in Escherichia coli, and nine gene products, including previously characterized BzaA, BzaB, and vanillin dehydrogenase (LigV), exhibited oxidation activities for syringaldehyde to produce syringate. Among these genes, SLG_28320 (desV) and ligV were most highly and constitutively transcribed in the SYK-6 cells. Disruption of desV in SYK-6 resulted in a significant reduction in growth on syringaldehyde and in syringaldehyde oxidation activity. Furthermore, a desV ligV double mutant almost completely lost its ability to grow on syringaldehyde. Purified DesV showed similar kcat/Km values for syringaldehyde (2100 s−1·mM−1) and vanillin (1700 s−1·mM−1), whereas LigV substantially preferred vanillin (8800 s−1·mM−1) over syringaldehyde (1.4 s−1·mM−1). These results clearly demonstrate that desV plays a major role in syringaldehyde catabolism. Phylogenetic analyses showed that DesV-like ALDHs formed a distinct phylogenetic cluster separated from the vanillin dehydrogenase cluster. PMID:28294121

  13. Aldehyde dehydrogenase inhibition combined with phenformin treatment reversed NSCLC through ATP depletion

    PubMed Central

    Lee, Jae-Seon; Nam, Boas; Seong, Tae Wha; Son, Jaekyoung; Jang, Hyonchol; Hong, Kyeong Man; Lee, Cheolju; Kim, Soo-Youl

    2016-01-01

    Among ALDH isoforms, ALDH1L1 in the folate pathway showed highly increased expression in non-small-cell lung cancer cells (NSCLC). Based on the basic mechanism of ALDH converting aldehyde to carboxylic acid with by-product NADH, we suggested that ALDH1L1 may contribute to ATP production using NADH through oxidative phosphorylation. ALDH1L1 knockdown reduced ATP production by up to 60% concomitantly with decrease of NADH in NSCLC. ALDH inhibitor, gossypol, also reduced ATP production in a dose dependent manner together with decrease of NADH level in NSCLC. A combination treatment of gossypol with phenformin, mitochondrial complex I inhibitor, synergized ATP depletion, which efficiently induced cell death. Pre-clinical xenograft model using human NSCLC demonstrated a remarkable therapeutic response to the combined treatment of gossypol and phenformin. PMID:27384481

  14. Salivary aldehyde dehydrogenase--reversible oxidation of the enzyme and its inhibition by caffeine, investigated using fluorimetric method.

    PubMed

    Wierzchowski, Jacek; Pietrzak, Monika; Szelag, Malgorzata; Wroczyński, Piotr

    2008-05-01

    We have applied fluorimetric method to monitor aldehyde dehydrogenase (ALDH*) activity in human saliva samples to study inactivation, reactivation and inhibition of the enzyme. Saliva samples were collected to buffer stock solution, containing various thiols, and assayed in the presence of the fluorogenic substrate 6-dimethylamino-2-naphthaldehyde and NAD(+). Fluorescence of the produced 6-dimethylamino-2-naphthalene carboxylate was used to measure the reaction rate. Kinetic parameters for the highly fluorogenic substrate, 6-dimethylamino-2-naphthaldehyde were measured, with apparent K(m) of 7.9 microM at pH 7.3. The apparent K(m) for NAD(+) was 1.2 microM. The observed ALDH activity is unstable in the absence of thiols, but can be stabilized by 1mM glutathione, and inactivated enzyme can be re-activated within 10 min by treatment of 0.5 mM DTT. Two-assay procedure was applied to measure degree of inactivation of ALDH in saliva samples. It was found that degree of ALDH inactivation in fresh samples, stabilized by glutathione, is between 0% and 90%, with average value ca. 40%. Caffeine and theophylline were shown to be moderate inhibitors of salivary ALDH. Oxidation of the salivary ALDH in fresh saliva may be reliably measured using fluorimetric two-assay procedure. Preliminary statistics indicate that in most individuals this enzyme is partially inactive. Inhibition of the salivary ALDH by caffeine may have consequences for nutrition safety.

  15. Inhibition of aldehyde dehydrogenase 2 activity enhances antimycin-induced rat cardiomyocytes apoptosis through activation of MAPK signaling pathway.

    PubMed

    Zhang, Peng; Xu, Danling; Wang, Shijun; Fu, Han; Wang, Keqiang; Zou, Yunzeng; Sun, Aijun; Ge, Junbo

    2011-12-01

    Aldehyde dehydrogenase 2 (ALDH2), a mitochondrial-specific enzyme, has been proved to be involved in oxidative stress-induced cell apoptosis, while little is known in cardiomyocytes. This study was aimed at investigating the role of ALDH2 in antimycin A-induced cardiomyocytes apoptosis by suppressing ALDH2 activity with a specific ALDH2 inhibitor Daidzin. Antimycin A (40μg/ml) was used to induce neonatal cardiomyocytes apoptosis. Daidzin (60μM) effectively inhibited ALDH2 activity by 50% without own effect on cell apoptosis, and significantly enhanced antimycin A-induced cardiomyocytes apoptosis from 33.5±4.4 to 56.5±6.4% (Hochest method, p<0.05), and from 57.9±1.9 to 74.0±11.9% (FACS, p<0.05). Phosphorylation of activated MAPK signaling pathway, including extracellular signal-regulated kinase (ERK1/2), c-Jun NH2-terminal kinase (JNK) and p38 was also increased in antimycin A and daidzin treated cardiomyocytes compared to the cells treated with antimycin A alone. These findings indicated that modifying mitochondrial ALDH2 activity/expression might be a potential therapeutic option on reducing oxidative insults induced cardiomyocytes apoptosis.

  16. The Arabidopsis thaliana REDUCED EPIDERMAL FLUORESCENCE1 Gene Encodes an Aldehyde Dehydrogenase Involved in Ferulic Acid and Sinapic Acid Biosynthesis

    PubMed Central

    Nair, Ramesh B.; Bastress, Kristen L.; Ruegger, Max O.; Denault, Jeff W.; Chapple, Clint

    2004-01-01

    Recent research has significantly advanced our understanding of the phenylpropanoid pathway but has left in doubt the pathway by which sinapic acid is synthesized in plants. The reduced epidermal fluorescence1 (ref1) mutant of Arabidopsis thaliana accumulates only 10 to 30% of the sinapate esters found in wild-type plants. Positional cloning of the REF1 gene revealed that it encodes an aldehyde dehydrogenase, a member of a large class of NADP+-dependent enzymes that catalyze the oxidation of aldehydes to their corresponding carboxylic acids. Consistent with this finding, extracts of ref1 leaves exhibit low sinapaldehyde dehydrogenase activity. These data indicate that REF1 encodes a sinapaldehyde dehydrogenase required for sinapic acid and sinapate ester biosynthesis. When expressed in Escherichia coli, REF1 was found to exhibit both sinapaldehyde and coniferaldehyde dehydrogenase activity, and further phenotypic analysis of ref1 mutant plants showed that they contain less cell wall–esterified ferulic acid. These findings suggest that both ferulic acid and sinapic acid are derived, at least in part, through oxidation of coniferaldehyde and sinapaldehyde. This route is directly opposite to the traditional representation of phenylpropanoid metabolism in which hydroxycinnamic acids are instead precursors of their corresponding aldehydes. PMID:14729911

  17. Macrocytosis, macrocytic anemia, and genetic polymorphisms of alcohol dehydrogenase-1B and aldehyde dehydrogenase-2 in Japanese alcoholic men.

    PubMed

    Yokoyama, Akira; Yokoyama, Tetsuji; Brooks, Philip J; Mizukami, Takeshi; Matsui, Toshifumi; Kimura, Mitsuru; Matsushita, Sachio; Higuchi, Susumu; Maruyama, Katsuya

    2014-05-01

    Oxidation of ethanol by alcohol dehydrogenase (ADH) generates acetaldehyde (AcH), which is converted to acetate by aldehyde dehydrogenase-2 (ALDH2). Roughly 40% of East Asians are ALDH2-deficient due to an inactive enzyme encoded by the ALDH2*2 allele. ALDH2-deficient individuals have a dramatically elevated risk of esophageal cancer from alcohol consumption. We investigated the relationship between ALDH2*2, ADH1B*2 (encoding a highly active ADH) and erythrocyte abnormalities, in a population of Japanese alcoholic men (N = 1,238). Macrocytosis (mean corpuscular volume [MCV] ≥100 fl) and macrocytic anemia (MCV ≥100 fl and hemoglobin <13.5 g/dl) were found in 62.4 and 24.1% of the subjects, respectively. Age-adjusted daily alcohol consumption did not differ according to ADH1B and ALDH2 genotypes. However, macrocytosis and macrocytic anemia were strongly associated with the ALDH2*1/*2 genotype multivariate odds ratios (ORs; 95% confidence interval [CI] = 2.85 [1.95 to 4.18] and 3.68 [2.64 to 5.15], respectively, versus ALDH2*1/*1). In comparison with the ADH1B*1/*1 and ALDH2*1/*1 genotype combination, the ADH1B*1/*1 and ALDH2*1/*2 genotype combination and the ADH1B*2 allele and ALDH2*1/*2 genotype combination increased stepwise the ORs (95% CI) for macrocytosis (1.65 [0.92 to 2.94] and 4.07 [2.33 to 7.11], respectively, p for difference in OR = 0.015) and macrocytic anemia (2.80 [1.52 to 5.15] and 5.32 [3.29 to 8.62], respectively, p for difference in OR = 0.045). Genotype effects were more prominent on the risks of the more advanced erythrocyte abnormalities. Older age, cigarette smoking, and low body mass index independently increased the risks of the erythrocyte abnormalities. Consumption of beer, which contains folate, decreased the risks, whereas consumption of alcoholic beverages lacking folate did not. These results suggest that the erythrocyte abnormalities in alcoholics are attributable to high AcH exposure as well as to nutritional deficiencies and may

  18. Human aldehyde dehydrogenase 3A1 (ALDH3A1) exhibits chaperone-like function.

    PubMed

    Voulgaridou, Georgia-Persephoni; Tsochantaridis, Ilias; Mantso, Theodora; Franco, Rodrigo; Panayiotidis, Mihalis I; Pappa, Aglaia

    2017-08-01

    Aldehyde dehydrogenase 3A1 (ALDH3A1) is a metabolic enzyme that catalyzes the oxidation of various aldehydes. Certain types of epithelial tissues in mammals, especially those continually exposed to environmental stress (e.g., corneal epithelium), express ALDH3A1 at high levels and its abundance in such tissues is perceived to help to maintain cellular homeostasis under conditions of oxidative stress. Metabolic as well as non-metabolic roles for ALDH3A1 have been associated with its mediated resistance to cellular oxidative stress. In this study, we provide evidence that ALDH3A1 exhibits molecular chaperone-like activity further supporting its multifunctional role. Specifically, we expressed and purified the human ALDH3A1 in E. coli and used the recombinant protein to investigate its in vitro ability to protect SmaI and citrate synthase (from precipitation and/or deactivation) under thermal stress conditions. Our results indicate that recombinant ALDH3A1 exhibits significant chaperone function in vitro. Furthermore, over-expression of the fused histidine-tagged ALDH3A1 confers host E. coli cells with enhanced resistance to thermal shock, while ALDH3A1 over-expression in the human corneal cell line HCE-2 was sufficient for protecting them from the cytotoxic effects of both hydrogen peroxide and tert-butyl hydroperoxide. These results further support the chaperone-like function of human ALDH3A1. Taken together, ALDH3A1, in addition to its primary metabolic role in fundamental cellular detoxification processes, appears to play an essential role in protecting cellular proteins against aggregation under stress conditions. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Pre-steady-state kinetic studies on cytoplasmic sheep liver aldehyde dehydrogenase.

    PubMed

    MacGibbon, A K; Blackwell, L F; Buckley, P D

    1977-11-01

    Stopped-flow experiments in which sheep liver cytoplasmic aldehyde dehydrogenase (EC 1.2.1.3) was rapidly mixed with NAD(+) and aldehyde showed a burst of NADH formation, followed by a slower steady-state turnover. The kinetic data obtained when the relative concentrations and orders of mixing of NAD(+) and propionaldehyde with the enzyme were varied were fitted to the following mechanism: [Formula: see text] where the release of NADH is slow. By monitoring the quenching of protein fluorescence on the binding of NAD(+), estimates of 2x10(5) litre.mol(-1).s(-1) and 2s(-1) were obtained for k(+1) and k(-1) respectively. Although k(+3) could be determined from the dependence of the burst rate constant on the concentration of propionaldehyde to be 11s(-1), k(+2) and k(-2) could not be determined uniquely, but could be related by the equation: (k(-2)+k(+3))/k(+2) =50x10(-6)mol.litre(-1). No significant isotope effect was observed when [1-(2)H]propionaldehyde was used as substrate. The burst rate constant was pH-dependent, with the greatest rate constants occurring at high pH. Similar data were obtained by using acetaldehyde, where for this substrate (k(-2)+k(+3))/k(+2)=2.3x10 (-3)mol.litre(-1) and k(+3) is 23s(-1). When [1,2,2,2-(2)H]acetaldehyde was used, no isotope effect was observed on k(+3), but there was a significant effect on k(+2) and k(-2). A burst of NADH production has also been observed with furfuraldehyde, trans-4-(NN-dimethylamino)cinnamaldehyde, formaldehyde, benzaldehyde, 4-(imidazol-2-ylazo)benzaldehyde, p-methoxybenzaldehyde and p-methylbenzaldehyde as substrates, but not with p-nitrobenzaldehyde.

  20. Effects of Alda-1, an Aldehyde Dehydrogenase-2 Agonist, on Hypoglycemic Neuronal Death

    PubMed Central

    Ikeda, Tetsuhiko; Takahashi, Tetsuya; Tsujita, Mika; Kanazawa, Masato; Toriyabe, Masafumi; Koyama, Misaki; Itoh, Kosuke; Nakada, Tsutomu; Nishizawa, Masatoyo; Shimohata, Takayoshi

    2015-01-01

    Hypoglycemic encephalopathy (HE) is caused by a lack of glucose availability to neuronal cells, and no neuroprotective drugs have been developed as yet. Studies on the pathogenesis of HE and the development of new neuroprotective drugs have been conducted using animal models such as the hypoglycemic coma model and non-coma hypoglycemia model. However, both models have inherent problems, and establishment of animal models that mimic clinical situations is desirable. In this study, we first developed a short-term hypoglycemic coma model in which rats could be maintained in an isoelectric electroencephalogram (EEG) state for 2 min and subsequent hyperglycemia without requiring anti-seizure drugs and an artificial ventilation. This condition caused the production of 4-hydroxy-2-nonenal (4-HNE), a cytotoxic aldehyde, in neurons of the hippocampus and cerebral cortex, and a marked increase in neuronal death as evaluated by Fluoro-Jade B (FJB) staining. We also investigated whether N-(1,3-benzodioxole-5-ylmethyl)-2,6-dichlorobenzamide (Alda-1), a small-molecule agonist of aldehyde dehydrogenase-2, could attenuate 4-HNE levels and reduce hypoglycemic neuronal death. After confirming that EEG recordings remained isoelectric for 2 min, Alda-1 (8.5 mg/kg) or vehicle (dimethyl sulfoxide; DMSO) was administered intravenously with glucose to maintain a blood glucose level of 250 to 270 mg/dL. Fewer 4-HNE and FJB-positive cells were observed in the cerebral cortex of Alda-1-treated rats than in DMSO-treated rats 24 h after glucose administration (P = 0.002 and P = 0.020). Thus, activation of the ALDH2 pathway could be a molecular target for HE treatment, and Alda-1 is a potentially neuroprotective agent that exerts a beneficial effect on neurons when intravenously administered simultaneously with glucose. PMID:26083658

  1. Aldehyde Dehydrogenase Gene Superfamily in Populus: Organization and Expression Divergence between Paralogous Gene Pairs.

    PubMed

    Tian, Feng-Xia; Zang, Jian-Lei; Wang, Tan; Xie, Yu-Li; Zhang, Jin; Hu, Jian-Jun

    2015-01-01

    Aldehyde dehydrogenases (ALDHs) constitute a superfamily of NAD(P)+-dependent enzymes that catalyze the irreversible oxidation of a wide range of reactive aldehydes to their corresponding nontoxic carboxylic acids. ALDHs have been studied in many organisms from bacteria to mammals; however, no systematic analyses incorporating genome organization, gene structure, expression profiles, and cis-acting elements have been conducted in the model tree species Populus trichocarpa thus far. In this study, a comprehensive analysis of the Populus ALDH gene superfamily was performed. A total of 26 Populus ALDH genes were found to be distributed across 12 chromosomes. Genomic organization analysis indicated that purifying selection may have played a pivotal role in the retention and maintenance of PtALDH gene families. The exon-intron organizations of PtALDHs were highly conserved within the same family, suggesting that the members of the same family also may have conserved functionalities. Microarray data and qRT-PCR analysis indicated that most PtALDHs had distinct tissue-specific expression patterns. The specificity of cis-acting elements in the promoter regions of the PtALDHs and the divergence of expression patterns between nine paralogous PtALDH gene pairs suggested that gene duplications may have freed the duplicate genes from the functional constraints. The expression levels of some ALDHs were up- or down-regulated by various abiotic stresses, implying that the products of these genes may be involved in the adaptation of Populus to abiotic stresses. Overall, the data obtained from our investigation contribute to a better understanding of the complexity of the Populus ALDH gene superfamily and provide insights into the function and evolution of ALDH gene families in vascular plants.

  2. Engineering the nucleotide coenzyme specificity and sulfhydryl redox sensitivity of two stress-responsive aldehyde dehydrogenase isoenzymes of Arabidopsis thaliana.

    PubMed

    Stiti, Naim; Adewale, Isaac O; Petersen, Jan; Bartels, Dorothea; Kirch, Hans-Hubert

    2011-03-15

    Lipid peroxidation is one of the consequences of environmental stress in plants and leads to the accumulation of highly toxic, reactive aldehydes. One of the processes to detoxify these aldehydes is their oxidation into carboxylic acids catalyzed by NAD(P)+-dependent ALDHs (aldehyde dehydrogenases). We investigated kinetic parameters of two Arabidopsis thaliana family 3 ALDHs, the cytosolic ALDH3H1 and the chloroplastic isoform ALDH3I1. Both enzymes had similar substrate specificity and oxidized saturated aliphatic aldehydes. Catalytic efficiencies improved with the increase of carbon chain length. Both enzymes were also able to oxidize α,β-unsaturated aldehydes, but not aromatic aldehydes. Activity of ALDH3H1 was NAD+-dependent, whereas ALDH3I1 was able to use NAD+ and NADP+. An unusual isoleucine residue within the coenzyme-binding cleft was responsible for the NAD+-dependence of ALDH3H1. Engineering the coenzyme-binding environment of ALDH3I1 elucidated the influence of the surrounding amino acids. Enzyme activities of both ALDHs were redox-sensitive. Inhibition was correlated with oxidation of both catalytic and non-catalytic cysteine residues in addition to homodimer formation. Dimerization and inactivation could be reversed by reducing agents. Mutant analysis showed that cysteine residues mediating homodimerization are located in the N-terminal region. Modelling of the protein structures revealed that the redox-sensitive cysteine residues are located at the surfaces of the subunits.

  3. The mitochondrial monoamine oxidase-aldehyde dehydrogenase pathway: a potential site of action of daidzin.

    PubMed

    Rooke, N; Li, D J; Li, J; Keung, W M

    2000-11-02

    Recent studies showed that daidzin suppresses ethanol intake in ethanol-preferring laboratory animals. In vitro, it potently and selectively inhibits the mitochondrial aldehyde dehydrogenase (ALDH-2). Further, it inhibits the conversion of monoamines such as serotonin (5-HT) and dopamine (DA) into their respective acid metabolites, 5-hydroxyindole-3-acetic acid (5-HIAA) and 3,4-dihydroxyphenylacetic acid (DOPAC) in isolated hamster or rat liver mitochondria. Studies on the suppression of ethanol intake and inhibition of 5-HIAA (or DOPAC) formation by six structural analogues of daidzin suggested a potential link between these two activities. This, together with the finding that daidzin does not affect the rates of mitochondria-catalyzed oxidative deamination of these monoamines, raised the possibility that the ethanol intake-suppressive (antidipsotropic) action of daidzin is not mediated by the monoamines but rather by their reactive biogenic aldehyde intermediates such as 5-hydroxyindole-3-acetaldehyde (5-HIAL) and/or 3,4-dihydroxyphenylacetaldehyde (DOPAL) which accumulate in the presence of daidzin. To further evaluate this possibility, we synthesized more structural analogues of daidzin and tested and compared their antidipsotropic activities in Syrian golden hamsters with their effects on monoamine metabolism in isolated hamster liver mitochondria using 5-HT as the substrate. Effects of daidzin and its structural analogues on the activities of monoamine oxidase (MAO) and ALDH-2, the key enzymes involved in 5-HT metabolism in the mitochondria, were also examined. Results from these studies reveal a positive correlation between the antidipsotropic activities of these analogues and their abilities to increase 5-HIAL accumulation during 5-HT metabolism in isolated hamster liver mitochondria. Daidzin analogues that potently inhibit ALDH-2 but have no or little effect on MAO are most antidipsotropic, whereas those that also potently inhibit MAO exhibit little, if

  4. Alternative Biotransformation of Retinal to Retinoic Acid or Retinol by an Aldehyde Dehydrogenase from Bacillus cereus

    PubMed Central

    Hong, Seung-Hye; Ngo, Ho-Phuong-Thuy; Nam, Hyun-Koo; Kim, Kyoung-Rok

    2016-01-01

    ABSTRACT A novel bacterial aldehyde dehydrogenase (ALDH) that converts retinal to retinoic acid was first identified in Bacillus cereus. The amino acid sequence of ALDH from B. cereus (BcALDH) was more closely related to mammalian ALDHs than to bacterial ALDHs. This enzyme converted not only small aldehydes to carboxylic acids but also the large aldehyde all-trans-retinal to all-trans-retinoic acid with NAD(P)+. We newly found that BcALDH and human ALDH (ALDH1A1) could reduce all-trans-retinal to all-trans-retinol with NADPH. The catalytic residues in BcALDH were Glu266 and Cys300, and the cofactor-binding residues were Glu194 and Glu457. The E266A and C300A variants showed no oxidation activity. The E194S and E457V variants showed 15- and 7.5-fold higher catalytic efficiency (kcat/Km) for the reduction of all-trans-retinal than the wild-type enzyme, respectively. The wild-type, E194S variant, and E457V variant enzymes with NAD+ converted 400 μM all-trans-retinal to 210 μM all-trans-retinoic acid at the same amount for 240 min, while with NADPH, they converted 400 μM all-trans-retinal to 20, 90, and 40 μM all-trans-retinol, respectively. These results indicate that BcALDH and its variants are efficient biocatalysts not only in the conversion of retinal to retinoic acid but also in its conversion to retinol with a cofactor switch and that retinol production can be increased by the variant enzymes. Therefore, BcALDH is a novel bacterial enzyme for the alternative production of retinoic acid and retinol. IMPORTANCE Although mammalian ALDHs have catalyzed the conversion of retinal to retinoic acid with NAD(P)+ as a cofactor, a bacterial ALDH involved in the conversion is first characterized. The biotransformation of all-trans-retinal to all-trans-retinoic acid by BcALDH and human ALDH was altered to the biotransformation to all-trans-retinol by a cofactor switch using NADPH. Moreover, the production of all-trans-retinal to all-trans-retinol was changed by mutations

  5. Expression of a gene encoding mitochondrial aldehyde dehydrogenase in rice increases under submerged conditions.

    PubMed

    Nakazono, M; Tsuji, H; Li, Y; Saisho, D; Arimura, S; Tsutsumi, N; Hirai, A

    2000-10-01

    It is known that alcoholic fermentation is important for survival of plants under anaerobic conditions. Acetaldehyde, one of the intermediates of alcoholic fermentation, is not only reduced by alcohol dehydrogenase but also can be oxidized by aldehyde dehydrogenase (ALDH). To determine whether ALDH plays a role in anaerobic metabolism in rice (Oryza sativa L. cv Nipponbare), we characterized a cDNA clone encoding mitochondrial ALDH from rice (Aldh2a). Analysis of sub-cellular localization of ALDH2a protein using green fluorescent protein and an in vitro ALDH assay using protein extracts from Escherichia coli cells that overexpressed ALDH2a indicated that ALDH2a functions in the oxidation of acetaldehyde in mitochondria. A Southern-blot analysis indicated that mitochondrial ALDH is encoded by at least two genes in rice. We found that the Aldh2a mRNA was present at high levels in leaves of dark-grown seedlings, mature leaf sheaths, and panicles. It is interesting that expression of the rice Aldh2a gene, unlike the expression of the tobacco (Nicotiana tabacum) Aldh2a gene, was induced in rice seedlings by submergence. Experiments with ruthenium red, which is a blocker of Ca(2+) fluxes in rice as well as maize (Zea mays), suggest that the induction of expression of Adh1 and Pdc1 by low oxygen stress is regulated by elevation of the cytosolic Ca(2+) level. However, the induction of Aldh2a gene expression may not be controlled by the cytosolic Ca(2+) level elevation. A possible involvement of ALDH2a in the submergence tolerance of rice is discussed.

  6. RNA interference against aldehyde dehydrogenase-2: development of tools for alcohol research.

    PubMed

    Cortínez, Gabriel; Sapag, Amalia; Israel, Yedy

    2009-03-01

    Liver alcohol dehydrogenase oxidizes ethanol to acetaldehyde, which is further oxidized to acetate by aldehyde dehydrogenase-2 (ALDH2*1). Individuals who carry a low-activity ALDH2 (ALDH2*2) display high blood acetaldehyde levels after ethanol consumption, which leads to dysphoric effects, such as facial flushing, nausea, dizziness, and headache ("Asian alcohol phenotype"), which result in an aversion to alcohol and protection against alcohol abuse and alcoholism. Mimicking this phenotype may reduce alcohol consumption in alcoholics. RNA interference (RNAi) is a cell process in which a short interfering RNA (siRNA) of 21-25 bp guides the degradation of a complementary target mRNA. Thus, siRNAs may be useful in mimicking the Asian phenotype by inhibiting ALDH2 gene expression. We determined the inhibitory effect of three chemically synthesized siRNAs targeted against rat ALDH2 mRNA in human embryonic kidney cells (HEK-293 cell lines) transfected with a plasmid carrying the rat ALDH2 cDNA. Two of the three siRNAs were active, yielding a 65-75% reduction of ALDH2 activity. Based on the most promising siRNA sequence, three short hairpin RNA (shRNA) genes driven by the human U6 RNA promoter were designed and cloned in a plasmid. After transfection of HEK-293 cells, one of the genes was shown to be active, yielding a 50% reduction of ALDH2 activity. This effect is consistent with a 50% reduction in ALDH2 mRNA, whereas neither beta-actin mRNA nor the interferon-inducible transmembrane protein-1 mRNA levels were affected. This study describes chemically synthesized siRNAs and an endogenously synthesized shRNA, which reduce ALDH2 activity and constitute tools that should be of value for further alcohol research.

  7. A comparison of nitrophenyl esters and lactones as substrates of cytosolic aldehyde dehydrogenase.

    PubMed Central

    Kitson, T M; Kitson, K E

    1996-01-01

    1. p-Nitrophenyl (PNP) acetate and propionate show a burst of p-nitrophenoxide release when their hydrolysis is catalysed by sheep liver cytosolic aldehyde dehydrogenase. This is not seen in the presence of NAD+ or NADH, implying a change in ratedetermining step. 2. 6-Nitrodihydrocoumarin (6-NDC) shows no burst of absorbance in the visible region. We propose that the pKa of the transient "reporter group' produced during the hydrolysis of this lactone is high (approx. 10) and that the incipient covalently linked p-nitrophenoxide moiety is protonated immediately on formation. The small burst seen in the hydrolysis of 5-nitro-2-coumaranone (5-NC) suggests that the pKa of its reporter group is about 8.5. 3. NADH markedly enhances the steady-state rate with the lactones. 5-NC shows a large rapid burst of colour development in the presence of NADH; this implies that NADH decreases the pKa of the reporter group to 7-7.5. 4. In the presence of NAD+, 5-NC and 6-NDC give an unusual "negative burst' in the stopped-flow traces. We propose that, under these circumstances, acylation of the enzyme is extremely fast and that the first event seen in the stopped-flow traces is protonation of the reporter group. NAD+ also greatly increases the steady-state rate. 5. With the lactones in the presence of NADH, the kcat value (nearly 6 s-1), a measure of the deacylation rate, is compatible with the single-site model for dehydrogenase and esterase activities. PMID:8645209

  8. Geometric specificity of alcohol dehydrogenases and its potential for separation of trans and cis isomers of unsaturated aldehydes.

    PubMed Central

    Klibanov, A M; Giannousis, P P

    1982-01-01

    The geometric specificity of three different alcohol dehydrogenases (alcohol:NAD+ oxidoreductase, EC 1.1.1.1) (from yeast, from horse liver, and from Leuconostoc mesenteroides) in the reduction of trans- and cis-cinnamaldehydes has been investigated. All three enzymes display a remarkable trans specificity: they react with the trans isomer 7 to 647 times faster than with its cis counterpart. Experiments with the enzymatic reduction of 3-phenylpropionaldehyde, a saturated analog of cinnamaldehyde, have revealed that whereas trans-cinnamaldehyde possesses the "right" configuration for the active centers of the alcohol dehydrogenases, the cis isomer apparently does not fit the active centers well. All three alcohol dehydrogenases studied also exhibit a marked trans specificity in the reaction with alpha-methylcinnamaldehyde. The geometric specificity of alcohol dehydrogenases can be used for the production of otherwise hard to synthesize cis isomers of unsaturated aldehydes from their readily available trans counterparts: trans-cinnamaldehyde was irradiated with ultraviolet light (which converted it to a mixture of trans and cis isomers) then treated with NADH and yeast alcohol dehydrogenase (which selectively reduces only trans aldehyde into the alcohol), and finally the mixture of cis-cinnamaldehyde and trans-cinnamyl alcohol was separated easily by preparative column chromatography. PMID:7048306

  9. Aldehyde dehydrogenase 1B1: molecular cloning and characterization of a novel mitochondrial acetaldehyde-metabolizing enzyme.

    PubMed

    Stagos, Dimitrios; Chen, Ying; Brocker, Chad; Donald, Elizabeth; Jackson, Brian C; Orlicky, David J; Thompson, David C; Vasiliou, Vasilis

    2010-10-01

    Ethanol-induced damage is largely attributed to its toxic metabolite, acetaldehyde. Clearance of acetaldehyde is achieved by its oxidation, primarily catalyzed by the mitochondrial class II aldehyde dehydrogenase (ALDH2). ALDH1B1 is another mitochondrial aldehyde dehydrogenase (ALDH) that shares 75% peptide sequence homology with ALDH2. Recent population studies in whites suggest a role for ALDH1B1 in ethanol metabolism. However, to date, no formal documentation of the biochemical properties of ALDH1B1 has been forthcoming. In this current study, we cloned and expressed human recombinant ALDH1B1 in Sf9 insect cells. The resultant enzyme was purified by affinity chromatography to homogeneity. The kinetic properties of purified human ALDH1B1 were assessed using a wide range of aldehyde substrates. Human ALDH1B1 had an exclusive preference for NAD(+) as the cofactor and was catalytically active toward short- and medium-chain aliphatic aldehydes, aromatic aldehydes, and the products of lipid peroxidation, 4-hydroxynonenal and malondialdehyde. Most importantly, human ALDH1B1 exhibited an apparent K(m) of 55 μM for acetaldehyde, making it the second low K(m) ALDH for metabolism of this substrate. The dehydrogenase activity of ALDH1B1 was sensitive to disulfiram inhibition, a feature also shared with ALDH2. The tissue distribution of ALDH1B1 in C57BL/6J mice and humans was examined by quantitative polymerase chain reaction, Western blotting, and immunohistochemical analysis. The highest expression occurred in the liver, followed by the intestinal tract, implying a potential physiological role for ALDH1B1 in these tissues. The current study is the first report on the expression, purification, and biochemical characterization of human ALDH1B1 protein.

  10. Aldehyde Dehydrogenase 1B1: Molecular Cloning and Characterization of a Novel Mitochondrial Acetaldehyde-Metabolizing Enzyme

    PubMed Central

    Stagos, Dimitrios; Chen, Ying; Brocker, Chad; Donald, Elizabeth; Jackson, Brian C.; Orlicky, David J.; Thompson, David C.

    2010-01-01

    Ethanol-induced damage is largely attributed to its toxic metabolite, acetaldehyde. Clearance of acetaldehyde is achieved by its oxidation, primarily catalyzed by the mitochondrial class II aldehyde dehydrogenase (ALDH2). ALDH1B1 is another mitochondrial aldehyde dehydrogenase (ALDH) that shares 75% peptide sequence homology with ALDH2. Recent population studies in whites suggest a role for ALDH1B1 in ethanol metabolism. However, to date, no formal documentation of the biochemical properties of ALDH1B1 has been forthcoming. In this current study, we cloned and expressed human recombinant ALDH1B1 in Sf9 insect cells. The resultant enzyme was purified by affinity chromatography to homogeneity. The kinetic properties of purified human ALDH1B1 were assessed using a wide range of aldehyde substrates. Human ALDH1B1 had an exclusive preference for NAD+ as the cofactor and was catalytically active toward short- and medium-chain aliphatic aldehydes, aromatic aldehydes, and the products of lipid peroxidation, 4-hydroxynonenal and malondialdehyde. Most importantly, human ALDH1B1 exhibited an apparent Km of 55 μM for acetaldehyde, making it the second low Km ALDH for metabolism of this substrate. The dehydrogenase activity of ALDH1B1 was sensitive to disulfiram inhibition, a feature also shared with ALDH2. The tissue distribution of ALDH1B1 in C57BL/6J mice and humans was examined by quantitative polymerase chain reaction, Western blotting, and immunohistochemical analysis. The highest expression occurred in the liver, followed by the intestinal tract, implying a potential physiological role for ALDH1B1 in these tissues. The current study is the first report on the expression, purification, and biochemical characterization of human ALDH1B1 protein. PMID:20616185

  11. NOTCH-induced aldehyde dehydrogenase 1A1 deacetylation promotes breast cancer stem cells.

    PubMed

    Zhao, Di; Mo, Yan; Li, Meng-Tian; Zou, Shao-Wu; Cheng, Zhou-Li; Sun, Yi-Ping; Xiong, Yue; Guan, Kun-Liang; Lei, Qun-Ying

    2014-12-01

    High aldehyde dehydrogenase (ALDH) activity is a marker commonly used to isolate stem cells, particularly breast cancer stem cells (CSCs). Here, we determined that ALDH1A1 activity is inhibited by acetylation of lysine 353 (K353) and that acetyltransferase P300/CBP-associated factor (PCAF) and deacetylase sirtuin 2 (SIRT2) are responsible for regulating the acetylation state of ALDH1A1 K353. Evaluation of breast carcinoma tissues from patients revealed that cells with high ALDH1 activity have low ALDH1A1 acetylation and are capable of self-renewal. Acetylation of ALDH1A1 inhibited both the stem cell population and self-renewal properties in breast cancer. Moreover, NOTCH signaling activated ALDH1A1 through the induction of SIRT2, leading to ALDH1A1 deacetylation and enzymatic activation to promote breast CSCs. In breast cancer xenograft models, replacement of endogenous ALDH1A1 with an acetylation mimetic mutant inhibited tumorigenesis and tumor growth. Together, the results from our study reveal a function and mechanism of ALDH1A1 acetylation in regulating breast CSCs.

  12. Aldehyde dehydrogenase expression in Metaphire posthuma as a bioindicator to monitor heavy metal pollution in soil.

    PubMed

    Panday, Raju; Bhatt, Padam Shekhar; Bhattarai, Tribikram; Shakya, Kumudini; Sreerama, Lakshmaiah

    2016-11-21

    Soil contamination and associated pollution plays a detrimental role in soil flora and fauna. Soil is processed and remodeled by subterranean earthworms, accordingly are referred to as soil chemical engineers. These worms, besides processing carbon and nitrogen, serve as minors for processing metals. In heavy metal contaminated soils, they accumulate heavy metals, which in turn cause altered gene expression, including aldehyde dehydrogenase (ALDH) enzymes. This study explores the possibility of ALDH expression in earthworms as a novel biomarker for the heavy metal contamination of soil. Earthworms cultured in contaminated soils accumulated significantly higher levels of Pb and Cd. Similarly, significantly higher levels of ALDH enzyme activities were observed in earthworms cultured in soils contaminated with Pb and Cd. The ALDH activity was found to be highest in worms cultured in 5 ppm heavy metal contaminated soils. Although, ALDH activities decreased as the heavy metal concentration in soil increased, they were significantly higher when compared to control worms cultured in uncontaminated soils. The accumulation of heavy metal in earthworms measured after 28 days decreased as the heavy metal concentration in soil increased. Levels of ALDH expression correlated with total Pb and Cd concentration in the earthworm tissue. This study showed that the ALDH activity in earthworms could potentially be used as a biomarker to show heavy metal pollution in soil.

  13. Inhibition of Aldehyde Dehydrogenase Expands Hematopoietic Stem Cells with Radioprotective Capacity†

    PubMed Central

    Muramoto, Garrett G.; Russell, J. Lauren; Safi, Rachid; Salter, Alice B.; Himburg, Heather A.; Daher, Pamela; Meadows, Sarah K.; Doan, Phuong; Storms, Robert W.; Chao, Nelson J.; McDonnell, Donald P.; Chute, John P.

    2013-01-01

    Hematopoietic stem cells (HSCs) are enriched for aldehyde dehydrogenase (ALDH) activity and ALDH is a selectable marker for human HSCs. However, the function of ALDH in HSC biology is not well understood. We sought to determine the function of ALDH in regulating HSC fate. Pharmacologic inhibition of ALDH with diethylaminobenzaldehyde (DEAB) impeded the differentiation of murine CD34−c-kit+Sca-1+lineage− (34−KSL) HSCs in culture and facilitated a ninefold expansion of cells capable of radioprotecting lethally irradiated mice compared to input 34−KSL cells. Treatment of bone marrow (BM) 34−KSL cells with DEAB caused a fourfold increase in 4-week competitive repopulating units, verifying the amplification of short-term HSCs (ST-HSCs) in response to ALDH inhibition. Targeted siRNA of ALDH1a1 in BM HSCs caused a comparable expansion of radioprotective progenitor cells in culture compared to DEAB treatment, confirming that ALDH1a1 was the target of DEAB inhibition. The addition of all trans retinoic acid blocked DEAB-mediated expansion of ST-HSCs in culture, suggesting that ALDH1a1 regulates HSC differentiation via augmentation of retinoid signaling. Pharmacologic inhibition of ALDH has therapeutic potential as a means to amplify ST-HSCs for transplantation purposes. PMID:20054864

  14. Aldehyde Dehydrogenase Activity Identifies a Population of Human Skeletal Muscle Cells With High Myogenic Capacities

    PubMed Central

    Vauchez, Karine; Marolleau, Jean-Pierre; Schmid, Michel; Khattar, Patricia; Chapel, Alain; Catelain, Cyril; Lecourt, Séverine; Larghéro, Jérôme; Fiszman, Marc; Vilquin, Jean-Thomas

    2009-01-01

    Aldehyde dehydrogenase 1A1 (ALDH) activity is one hallmark of human bone marrow (BM), umbilical cord blood (UCB), and peripheral blood (PB) primitive progenitors presenting high reconstitution capacities in vivo. In this study, we have identified ALDH+ cells within human skeletal muscles, and have analyzed their phenotypical and functional characteristics. Immunohistofluorescence analysis of human muscle tissue sections revealed rare endomysial cells. Flow cytometry analysis using the fluorescent substrate of ALDH, Aldefluor, identified brightly stained (ALDHbr) cells with low side scatter (SSClo), in enzymatically dissociated muscle biopsies, thereafter abbreviated as SMALD+ (for skeletal muscle ALDH+) cells. Phenotypical analysis discriminated two sub-populations according to CD34 expression: SMALD+/CD34− and SMALD+/CD34+ cells. These sub-populations did not initially express endothelial (CD31), hematopoietic (CD45), and myogenic (CD56) markers. Upon sorting, however, whereas SMALD+/CD34+ cells developed in vitro as a heterogeneous population of CD56− cells able to differentiate in adipoblasts, the SMALD+/CD34− fraction developed in vitro as a highly enriched population of CD56+ myoblasts able to form myotubes. Moreover, only the SMALD+/CD34− population maintained a strong myogenic potential in vivo upon intramuscular transplantation. Our results suggest that ALDH activity is a novel marker for a population of new human skeletal muscle progenitors presenting a potential for cell biology and cell therapy. PMID:19738599

  15. Aldehyde dehydrogenase 2 overexpression inhibits neuronal apoptosis after spinal cord ischemia/reperfusion injury

    PubMed Central

    Liu, Xing-zhen; Sun, Xin; Shen, Kang-ping; Jin, Wen-jie; Fu, Zhi-yi; Tao, Hai-rong; Xu, Zhi-xing

    2017-01-01

    Aldehyde dehydrogenase 2 (ALDH2) is an important factor in inhibiting oxidative stress and has been shown to protect against renal ischemia/reperfusion injury. Therefore, we hypothesized that ALDH2 could reduce spinal cord ischemia/reperfusion injury. Spinal cord ischemia/reperfusion injury was induced in rats using the modified Zivin's method of clamping the abdominal aorta. After successful model establishment, the agonist group was administered a daily consumption of 2.5% alcohol. At 7 days post-surgery, the Basso, Beattie, and Bresnahan score significantly increased in the agonist group compared with the spinal cord ischemia/reperfusion injury group. ALDH2 expression also significantly increased and the number of apoptotic cells significantly decreased in the agonist group than in the spinal cord ischemia/reperfusion injury group. Correlation analysis revealed that ALDH2 expression negatively correlated with the percentage of TUNEL-positive cells (r = −0.485, P < 0.01). In summary, increased ALDH2 expression protected the rat spinal cord against ischemia/reperfusion injury by inhibiting apoptosis. PMID:28852401

  16. Combined effects of aldehyde dehydrogenase variants and maternal mitochondrial genes on alcohol consumption.

    PubMed

    Israel, Yedy; Quintanilla, María E; Sapag, Amalia; Tampier, Lutske

    2006-01-01

    Two lines of rats bred to differ in their voluntary alcohol consumption--the alcohol-abstaining UChA rats and the alcohol-drinking UChB rats--differ in how effectively toxic acetaldehyde is removed during alcohol metabolism. UChB animals carry efficient variants of the aldehyde dehydrogenase 2 (ALDH2) genes and have active mitochondria, resulting in fast removal of acetaldehyde. UChA animals, in contrast, carry less efficient ALDH2 variants and less active mitochondria, which result in transient elevations of acetaldehyde levels after alcohol ingestion. Cross-breeding studies have demonstrated that the presence of active mitochondria inherited from UChB females can fully abolish the reduction of alcohol consumption associated with the presence of less efficient ALDH2 variants--a phenomenon known as epistasis. These and other findings suggest that mitochondrial activity during alcohol metabolism should be considered a new modulator of alcohol consumption not only in rats but also in other species, including humans.

  17. Aldehyde dehydrogenase 1 positive glioblastoma cells show brain tumor stem cell capacity.

    PubMed

    Rasper, Michael; Schäfer, Andrea; Piontek, Guido; Teufel, Julian; Brockhoff, Gero; Ringel, Florian; Heindl, Stefan; Zimmer, Claus; Schlegel, Jürgen

    2010-10-01

    Glioblastoma (GBM) is the most aggressive primary brain tumor and is resistant to all therapeutic regimens. Relapse occurs regularly and might be caused by a poorly characterized tumor stem cell (TSC) subpopulation escaping therapy. We suggest aldehyde dehydrogenase 1 (ALDH1) as a novel stem cell marker in human GBM. Using the neurosphere formation assay as a functional method to identify brain TSCs, we show that high protein levels of ALDH1 facilitate neurosphere formation in established GBM cell lines. Even single ALDH1 positive cells give rise to colonies and neurospheres. Consequently, the inhibition of ALDH1 in vitro decreases both the number of neurospheres and their size. Cell lines without expression of ALDH1 do not form tumor spheroids under the same culturing conditions. High levels of ALDH1 seem to keep tumor cells in an undifferentiated, stem cell-like state indicated by the low expression of beta-III-tubulin. In contrast, ALDH1 inhibition induces premature cellular differentiation and reduces clonogenic capacity. Primary cell cultures obtained from fresh tumor samples approve the established GBM cell line results.

  18. Tumour budding and the expression of cancer stem cell marker aldehyde dehydrogenase 1 in nasopharyngeal carcinoma.

    PubMed

    Luo, Wei-Ren; Gao, Fei; Li, Si-Yi; Yao, Kai-Tai

    2012-12-01

    To detect the prognostic significance of tumour budding and its expression of aldehyde dehydrogenase 1 (ALDH1) in nasopharyngeal carcinoma (NPC). Tumour budding was investigated in 105 patients with NPC by immunohistochemistry for pan-cytokeratin (AE1/AE3). The intensity of budding correlated strongly with T classification (P=0.008), lymphatic invasion (P<0.001), vascular invasion (P=0.029), lymph node metastasis (P < 0.001), and clinical stage (P=0.010). Univariate analysis revealed that patients with high budding grade had poorer survival than those with low grade (P=0.002). Multivariate analysis showed that tumour budding was an independent predictor of survival (P=0.001). Furthermore, budding cells showed high-level expression of the cancer stem cell (CSC) marker ALDH1. Budding cells with high-level ALDH1 expression contributed to several aggressive behaviours and poor survival (P=0.000).   We describe, for the first time, the presence of tumour budding and its correlation with aggressive tumour behaviour and poor patient survival in NPC. The degree of tumour budding could be a valuable predictive factor in NPC. In addition, we show, also for the first time, that budding cells in NPC might possess the invasive and metastatic properties of CSCs. © 2012 Blackwell Publishing Limited.

  19. Roles of histamine on the expression of aldehyde dehydrogenase 1 in endometrioid adenocarcinoma cell line.

    PubMed

    Wang, Yi; Jiang, Yang; Ikeda, Jun-Ichiro; Tian, Tian; Sato, Atsushi; Ohtsu, Hiroshi; Morii, Eiichi

    2014-10-01

    Cancer-initiating cells (CICs) are a limited number of cells that are essential for maintenance, recurrence, and metastasis of tumors. Aldehyde dehydrogenase 1 (ALDH1) has been recognized as a marker of CICs. We previously reported that ALDH1-high cases of uterine endometrioid adenocarcinoma showed poor prognosis, and that ALDH1 high population was more tumorigenic, invasive, and resistant to apoptosis than ALDH1 low population. Histamine plays a critical role in cancer cell proliferation, migration, and invasion. Here, we examined the effect of histamine on ALDH1 expression in endometrioid adenocarcinoma cell line. The addition of histamine increased ALDH1 high population, which was consistent with the result that histamine enhanced the invasive ability and the resistance to anticancer drug. Among 4 types of histamine receptors, histamine H1 and H2 receptor (H1R and H2R) were expressed in endometrioid adenocarcinoma cell line. The addition of H1R agonist but not H2R agonist increased ALDH1. The antagonist H1R but not H2R inhibited the effect of histamine on ALDH1 expression. These results indicated that histamine increased the expression of ALDH1 via H1R but not H2R. These findings may provide the evidence for exploring a new strategy to suppress CICs by inhibiting ALDH1 expression with histamine.

  20. Aldehyde dehydrogenase 1a3 defines a subset of failing pancreatic β cells in diabetic mice

    PubMed Central

    Kim-Muller, Ja Young; Fan, Jason; Kim, Young Jung R.; Lee, Seung-Ah; Ishida, Emi; Blaner, William S.; Accili, Domenico

    2016-01-01

    Insulin-producing β cells become dedifferentiated during diabetes progression. An impaired ability to select substrates for oxidative phosphorylation, or metabolic inflexibility, initiates progression from β-cell dysfunction to β-cell dedifferentiation. The identification of pathways involved in dedifferentiation may provide clues to its reversal. Here we isolate and functionally characterize failing β cells from various experimental models of diabetes and report a striking enrichment in the expression of aldehyde dehydrogenase 1 isoform A3 (ALDH+) as β cells become dedifferentiated. Flow-sorted ALDH+ islet cells demonstrate impaired glucose-induced insulin secretion, are depleted of Foxo1 and MafA, and include a Neurogenin3-positive subset. RNA sequencing analysis demonstrates that ALDH+ cells are characterized by: (i) impaired oxidative phosphorylation and mitochondrial complex I, IV and V; (ii) activated RICTOR; and (iii) progenitor cell markers. We propose that impaired mitochondrial function marks the progression from metabolic inflexibility to dedifferentiation in the natural history of β-cell failure. PMID:27572106

  1. Identification of Tumor Endothelial Cells with High Aldehyde Dehydrogenase Activity and a Highly Angiogenic Phenotype

    PubMed Central

    Maishi, Nako; Ohga, Noritaka; Hida, Yasuhiro; Kawamoto, Taisuke; Iida, Junichiro; Shindoh, Masanobu; Tsuchiya, Kunihiko; Shinohara, Nobuo; Hida, Kyoko

    2014-01-01

    Tumor blood vessels play an important role in tumor progression and metastasis. It has been reported that tumor endothelial cells (TECs) exhibit highly angiogenic phenotypes compared with those of normal endothelial cells (NECs). TECs show higher proliferative and migratory abilities than those NECs, together with upregulation of vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR2). Furthermore, compared with NECs, stem cell markers such as Sca-1, CD90, and multidrug resistance 1 are upregulated in TECs, suggesting that stem-like cells exist in tumor blood vessels. In this study, to reveal the biological role of stem-like TECs, we analyzed expression of the stem cell marker aldehyde dehydrogenase (ALDH) in TECs and characterized ALDHhigh TECs. TECs and NECs were isolated from melanoma-xenografted nude mice and normal dermis, respectively. ALDH mRNA expression and activity were higher in TECs than those in NECs. Next, ALDHhigh/low TECs were isolated by fluorescence-activated cell sorting to compare their characteristics. Compared with ALDHlow TECs, ALDHhigh TECs formed more tubes on Matrigel-coated plates and sustained the tubular networks longer. Furthermore, VEGFR2 expression was higher in ALDHhigh TECs than that in ALDHlow TECs. In addition, ALDH was expressed in the tumor blood vessels of in vivo mouse models of melanoma and oral carcinoma, but not in normal blood vessels. These findings indicate that ALDHhigh TECs exhibit an angiogenic phenotype. Stem-like TECs may have an essential role in tumor angiogenesis. PMID:25437864

  2. Revascularization of ischemic limbs after transplantation of human bone marrow cells with high aldehyde dehydrogenase activity

    PubMed Central

    Capoccia, Benjamin J.; Robson, Debra L.; Levac, Krysta D.; Maxwell, Dustin J.; Hohm, Sarah A.; Neelamkavil, Marian J.; Bell, Gillian I.; Xenocostas, Anargyros; Link, Daniel C.; Piwnica-Worms, David; Nolta, Jan A.

    2009-01-01

    The development of cell therapies to treat peripheral vascular disease has proven difficult because of the contribution of multiple cell types that coordinate revascularization. We characterized the vascular regenerative potential of transplanted human bone marrow (BM) cells purified by high aldehyde dehydrogenase (ALDHhi) activity, a progenitor cell function conserved between several lineages. BM ALDHhi cells were enriched for myelo-erythroid progenitors that produced multipotent hematopoietic reconstitution after transplantation and contained nonhematopoietic precursors that established colonies in mesenchymal-stromal and endothelial culture conditions. The regenerative capacity of human ALDHhi cells was assessed by intravenous transplantation into immune-deficient mice with limb ischemia induced by femoral artery ligation/transection. Compared with recipients injected with unpurified nucleated cells containing the equivalent of 2- to 4-fold more ALDHhi cells, mice transplanted with purified ALDHhi cells showed augmented recovery of perfusion and increased blood vessel density in ischemic limbs. ALDHhi cells transiently recruited to ischemic regions but did not significantly integrate into ischemic tissue, suggesting that transient ALDHhi cell engraftment stimulated endogenous revascularization. Thus, human BM ALDHhi cells represent a progenitor-enriched population of several cell lineages that improves perfusion in ischemic limbs after transplantation. These clinically relevant cells may prove useful in the treatment of critical ischemia in humans. PMID:19324906

  3. Aldehyde dehydrogenase variation enhances effect of pesticides associated with Parkinson disease

    PubMed Central

    Fitzmaurice, Arthur G.; Rhodes, Shannon L.; Cockburn, Myles; Ritz, Beate

    2014-01-01

    Objective: The objective of this study was to determine whether environmental and genetic alterations of neuronal aldehyde dehydrogenase (ALDH) enzymes were associated with increased Parkinson disease (PD) risk in an epidemiologic study. Methods: A novel ex vivo assay was developed to identify pesticides that can inhibit neuronal ALDH activity. These were investigated for PD associations in a population-based case-control study, the Parkinson's Environment & Genes (PEG) Study. Common variants in the mitochondrial ALDH2 gene were genotyped to assess effect measure modification (statistical interaction) of the pesticide effects by genetic variation. Results: All of the metal-coordinating dithiocarbamates tested (e.g., maneb, ziram), 2 imidazoles (benomyl, triflumizole), 2 dicarboxymides (captan, folpet), and 1 organochlorine (dieldrin) inhibited ALDH activity, potentially via metabolic byproducts (e.g., carbon disulfide, thiophosgene). Fifteen screened pesticides did not inhibit ALDH. Exposures to ALDH-inhibiting pesticides were associated with 2- to 6-fold increases in PD risk; genetic variation in ALDH2 exacerbated PD risk in subjects exposed to ALDH-inhibiting pesticides. Conclusion: ALDH inhibition appears to be an important mechanism through which environmental toxicants contribute to PD pathogenesis, especially in genetically vulnerable individuals, suggesting several potential interventions to reduce PD occurrence or slow or reverse its progression. PMID:24491970

  4. Vasodilatory effect of nitroglycerin in Japanese subjects with different aldehyde dehydrogenase 2 (ALDH2) genotypes.

    PubMed

    Miura, Takeshi; Nishinaka, Toru; Terada, Tomoyuki; Yonezawa, Kazuya

    2017-03-23

    The functional genetic polymorphism of aldehyde dehydrogenase 2 (ALDH2) influences the enzymatic activities of its wild type (Glu504 encoded by ALDH2*1) and mutant type (Lys504 encoded by ALDH2*2) proteins. The enzymatic activities of mutant-type ALDH2 are limited compared with those of the wild type. ALDH2 has been suggested as a critical factor for nitroglycerin-mediated vasodilation by some human studies and in vitro studies. Currently, there is no research on direct observations of the vasodilatory effect of nitroglycerin sublingual tablets, which is the generally used dosage form. In the present study, the contribution of ALDH2 to the vasodilatory effect of nitroglycerin sublingual tablets was investigated among three genotype groups (ALDH2*1/*1, ALDH2*1/*2, and ALDH2*2/*2) in Japanese. The results by direct assessments of in vivo nitroglycerin-mediated dilation showed no apparent difference in vasodilation among all genotypes of ALDH2. Furthermore, to analyze the effect of other factors (age and flow-mediated dilation), multiple regression analysis and Pearson's correlation coefficient analysis were carried out. These analyses also indicated that the genotypes of ALDH2 were not related to the degree of vasodilation. These results suggest the existence of other predominant pathway(s) for nitroglycerin biotransformation, at least with regard to clinical nitroglycerin (e.g., a sublingual tablet) in Japanese subjects.

  5. Aldehyde dehydrogenase 2 is associated with cognitive functions in patients with Parkinson’s disease

    PubMed Central

    Yu, Rwei-Ling; Tan, Chun-Hsiang; Lu, Ying-Che; Wu, Ruey-Meei

    2016-01-01

    Neurotransmitter degradation has been proposed to cause the accumulation of neurotoxic metabolites. The metabolism of these metabolites involves aldehyde dehydrogenase 2 (ALDH2). The Asian-specific single nucleotide polymorphism rs671 causes reduced enzyme activity. This study aims to explore whether Parkinson’s disease (PD) patients with reduced ALDH2 activity owing to the rs671 polymorphism are at risk for neuropsychological impairments. A total of 139 PD patients were recruited. Each participant was assessed for medical characteristics and their ALDH2 genotype. The Mini-Mental State Examination (MMSE), the Clinical Dementia Rating Scale and the Frontal Behavioral Inventory were used to measure neuropsychological functions. We found that the MMSE scores were significantly lower in patients with inactive ALDH2 (U = 1873.5, p = 0.02). The presence of cognitive impairments was significantly more frequent in the inactive ALDH2 group (46.0%) than in the active ALDH2 group (26.3%) (χ2 = 5.886, p = 0.01). The inactive group showed significant deterioration in hobbies and exhibited more severe “disorganization” and “hyper-sexuality” behaviours. The additive effects of the allele on the development of cognitive impairments in PD patients may be an important finding that provides further insight into the pathogenic mechanism of cognitive dysfunction in PD. PMID:27453488

  6. Cloning and heterologous expression of two aryl-aldehyde dehydrogenases from the white-rot basidiomycete Phanerochaete chrysosporium

    SciTech Connect

    Nakamura, Tomofumi; Ichinose, Hirofumi; Wariishi, Hiroyuki

    2010-04-09

    We identified two aryl-aldehyde dehydrogenase proteins (PcALDH1 and PcALDH2) from the white-rot basidiomycete Phanerochaete chrysosporium. Both PcALDHs were translationally up-regulated in response to exogenous addition of vanillin, one of the key aromatic compounds in the pathway of lignin degradation by basidiomycetes. To clarify the catalytic functions of PcALDHs, we isolated full-length cDNAs encoding these proteins and heterologously expressed the recombinant enzymes using a pET/Escherichia coli system. The open reading frames of both PcALDH1 and PcALDH2 consisted of 1503 nucleotides. The deduced amino acid sequences of both proteins showed high homologies with aryl-aldehyde dehydrogenases from other organisms and contained ten conserved domains of ALDHs. Moreover, a novel glycine-rich motif 'GxGxxxG' was located at the NAD{sup +}-binding site. The recombinant PcALDHs catalyzed dehydrogenation reactions of several aryl-aldehyde compounds, including vanillin, to their corresponding aromatic acids. These results strongly suggested that PcALDHs metabolize aryl-aldehyde compounds generated during fungal degradation of lignin and various aromatic xenobiotics.

  7. Structural shifts of aldehyde dehydrogenase enzymes were instrumental for the early evolution of retinoid-dependent axial patterning in metazoans

    PubMed Central

    Sobreira, Tiago J. P.; Marlétaz, Ferdinand; Simões-Costa, Marcos; Schechtman, Deborah; Pereira, Alexandre C.; Brunet, Frédéric; Sweeney, Sarah; Pani, Ariel; Aronowicz, Jochanan; Lowe, Christopher J.; Davidson, Bradley; Laudet, Vincent; Bronner, Marianne; de Oliveira, Paulo S. L.; Schubert, Michael; Xavier-Neto, José

    2011-01-01

    Aldehyde dehydrogenases (ALDHs) catabolize toxic aldehydes and process the vitamin A-derived retinaldehyde into retinoic acid (RA), a small diffusible molecule and a pivotal chordate morphogen. In this study, we combine phylogenetic, structural, genomic, and developmental gene expression analyses to examine the evolutionary origins of ALDH substrate preference. Structural modeling reveals that processing of small aldehydes, such as acetaldehyde, by ALDH2, versus large aldehydes, including retinaldehyde, by ALDH1A is associated with small versus large substrate entry channels (SECs), respectively. Moreover, we show that metazoan ALDH1s and ALDH2s are members of a single ALDH1/2 clade and that during evolution, eukaryote ALDH1/2s often switched between large and small SECs after gene duplication, transforming constricted channels into wide opened ones and vice versa. Ancestral sequence reconstructions suggest that during the evolutionary emergence of RA signaling, the ancestral, narrow-channeled metazoan ALDH1/2 gave rise to large ALDH1 channels capable of accommodating bulky aldehydes, such as retinaldehyde, supporting the view that retinoid-dependent signaling arose from ancestral cellular detoxification mechanisms. Our analyses also indicate that, on a more restricted evolutionary scale, ALDH1 duplicates from invertebrate chordates (amphioxus and ascidian tunicates) underwent switches to smaller and narrower SECs. When combined with alterations in gene expression, these switches led to neofunctionalization from ALDH1-like roles in embryonic patterning to systemic, ALDH2-like roles, suggesting functional shifts from signaling to detoxification. PMID:21169504

  8. Thymoquinone binds and activates human salivary aldehyde dehydrogenase: Potential therapy for the mitigation of aldehyde toxicity and maintenance of oral health.

    PubMed

    Laskar, Amaj Ahmed; Khan, Masood Alam; Askari, Fizza; Younus, Hina

    2017-10-01

    Human salivary aldehyde dehydrogenase (hsALDH) is a very important anti-oxidant enzyme present in the saliva. It is involved in the detoxification of toxic aldehydes and maintenance of oral health. Reduced level of hsALDH activity is a risk factor for oral cancer development. Thymoquinone (TQ) has many pharmacological activities and health benefits. This study aimed to examine the activation of hsALDH by TQ. The effect of TQ on the activity and kinetics of hsALDH was studied. The binding of TQ with the enzyme was examined by different biophysical methods and molecular docking analysis. TQ enhanced the dehydrogenase activity of crude and purified hsALDH by 3.2 and 2.9 fold, respectively. The Km of the purified enzyme decreased and the Vmax increased. The esterase activity also increased by 1.2 fold. No significant change in the nucleophilicity of the catalytic cysteine residue was observed. TQ forms a strong complex with hsALDH without altering the secondary structures of the enzyme. It fits in the active site of ALDH3A1 close to Cys 243 and the other highly conserved amino acid residues which lead to enhancement of substrate binding affinity and catalytic efficiency of the enzyme. TQ is expected to give better protection from toxic aldehydes in the oral cavity and to reduce the risk of oral cancer development through the activation of hsALDH. Therefore, the addition of TQ in the diet and other oral formulations is expected to be beneficial for health. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Modulation of aldehyde dehydrogenase activity affects (±)-4-hydroxy-2E-nonenal (HNE) toxicity and HNE-protein adduct levels in PC12 cells.

    PubMed

    Kong, Dehe; Kotraiah, Vinayaka

    2012-07-01

    Oxidative stress is known to be one of the major factors underlying Parkinson's disease (PD). One of the consequences of oxidative stress is lipid peroxidation. A toxic product of lipid peroxidation, (±)-4-hydroxy-2E-nonenal (HNE) leads to membrane disruption and formation of HNE-protein adducts and such adducts have been detected in PD brain tissues. Aldehyde dehydrogenases (ALDHs) are involved in metabolizing HNE and other endogenous aldehydes. Interestingly, the cytosolic aldehyde dehydrogenase 1A1 (ALDH1A1) has been reported to be down-regulated in brain tissues affected in PD which could result in enhancement of HNE toxicity. We sought to first establish the role of ALDH1A1 in mediating HNE toxicity in PC12 cells by overexpressing ALDH1A1 and by using disulfiram, an ALDH inhibitor. Overexpression and inhibition of ALDH1A1 activity resulted in reduced and increased HNE toxicity, respectively. We then established conditions for detecting HNE-protein adducts following HNE treatment and showed that overexpression and inhibition of ALDH activity resulted in reduced and increased formation of HNE-protein adducts, respectively. We also show that 6-methyl-2-(phenylazo)-3-pyridinol, previously identified as an activator of ALDH1A1, can protect PC12 cells against HNE-mediated toxicity and can cause a small but significant decrease in levels of HNE-protein adducts. Our results should encourage identification of more potent ALDH activators and their testing in the PC12-HNE model. Such cytoprotective compounds could then be tested for their neuroprotective activity in in vivo models of oxidative stress-induced PD.

  10. Formation of Nitric Oxide by Aldehyde Dehydrogenase-2 Is Necessary and Sufficient for Vascular Bioactivation of Nitroglycerin*

    PubMed Central

    Opelt, Marissa; Eroglu, Emrah; Waldeck-Weiermair, Markus; Russwurm, Michael; Koesling, Doris; Malli, Roland; Graier, Wolfgang F.; Fassett, John T.; Schrammel, Astrid; Mayer, Bernd

    2016-01-01

    Aldehyde dehydrogenase-2 (ALDH2) catalyzes vascular bioactivation of the antianginal drug nitroglycerin (GTN), resulting in activation of soluble guanylate cyclase (sGC) and cGMP-mediated vasodilation. We have previously shown that a minor reaction of ALDH2-catalyzed GTN bioconversion, accounting for about 5% of the main clearance-based turnover yielding inorganic nitrite, results in direct NO formation and concluded that this minor pathway could provide the link between vascular GTN metabolism and activation of sGC. However, lack of detectable NO at therapeutically relevant GTN concentrations (≤1 μm) in vascular tissue called into question the biological significance of NO formation by purified ALDH2. We addressed this issue and used a novel, highly sensitive genetically encoded fluorescent NO probe (geNOp) to visualize intracellular NO formation at low GTN concentrations (≤1 μm) in cultured vascular smooth muscle cells (VSMC) expressing an ALDH2 mutant that reduces GTN to NO but lacks clearance-based GTN denitration activity. NO formation was compared with GTN-induced activation of sGC. The addition of 1 μm GTN to VSMC expressing either wild-type or C301S/C303S ALDH2 resulted in pronounced intracellular NO elevation, with maximal concentrations of 7 and 17 nm, respectively. Formation of GTN-derived NO correlated well with activation of purified sGC in VSMC lysates and cGMP accumulation in intact porcine aortic endothelial cells infected with wild-type or mutant ALDH2. Formation of NO and cGMP accumulation were inhibited by ALDH inhibitors chloral hydrate and daidzin. The present study demonstrates that ALDH2-catalyzed NO formation is necessary and sufficient for GTN bioactivation in VSMC. PMID:27679490

  11. A Novel NADPH-Dependent Aldehyde Reductase Gene from Saccharomyces cerevisiae NRRL Y-12632 Involved in the Detoxification of Aldehyde Inhibitors Derived from Lignocellulosic Biomass Conversion

    USDA-ARS?s Scientific Manuscript database

    Aldehyde inhibitors such as furfural, 5-hydroxymethylfurfural (HMF), anisaldehyde, benzaldehyde, cinnamaldehyde, and phenylaldehyde are commonly generated during lignocellulosic biomass conversion process for low-cost cellulosic ethanol production that interferes with subsequent microbial growth and...

  12. Aldose and aldehyde reductases : structure-function studies on the coenzyme and inhibitor-binding sites.

    SciTech Connect

    El-Kabbani, O.; Old, S. E.; Ginell, S. L.; Carper, D. A.; Biosciences Division; Monash Univ.; NIH

    1999-09-03

    PURPOSE: To identify the structural features responsible for the differences in coenzyme and inhibitor specificities of aldose and aldehyde reductases. METHODS: The crystal structure of porcine aldehyde reductase in complex with NADPH and the aldose reductase inhibitor sorbinil was determined. The contribution of each amino acid lining the coenzyme-binding site to the binding of NADPH was calculated using the Discover package. In human aldose reductase, the role of the non-conserved Pro 216 (Ser in aldehyde reductase) in the binding of coenzyme was examined by site-directed mutagenesis. RESULTS: Sorbinil binds to the active site of aldehyde reductase and is hydrogen-bonded to Trp 22, Tyr 50, His 113, and the non-conserved Arg 312. Unlike tolrestat, the binding of sorbinil does not induce a change in the side chain conformation of Arg 312. Mutation of Pro 216 to Ser in aldose reductase makes the binding of coenzyme more similar to that of aldehyde reductase. CONCLUSIONS: The participation of non-conserved active site residues in the binding of inhibitors and the differences in the structural changes required for the binding to occur are responsible for the differences in the potency of inhibition of aldose and aldehyde reductases. We report that the non-conserved Pro 216 in aldose reductase contributes to the tight binding of NADPH.

  13. Characterization of Cardiac-Resident Progenitor Cells Expressing High Aldehyde Dehydrogenase Activity

    PubMed Central

    Roehrich, Marc-Estienne; Spicher, Albert; Milano, Giuseppina; Vassalli, Giuseppe

    2013-01-01

    High aldehyde dehydrogenase (ALDH) activity has been associated with stem and progenitor cells in various tissues. Human cord blood and bone marrow ALDH-bright (ALDHbr) cells have displayed angiogenic activity in preclinical studies and have been shown to be safe in clinical trials in patients with ischemic cardiovascular disease. The presence of ALDHbr cells in the heart has not been evaluated so far. We have characterized ALDHbr cells isolated from mouse hearts. One percent of nonmyocytic cells from neonatal and adult hearts were ALDHbr. ALDHvery-br cells were more frequent in neonatal hearts than adult. ALDHbr cells were more frequent in atria than ventricles. Expression of ALDH1A1 isozyme transcripts was highest in ALDHvery-br cells, intermediate in ALDHbr cells, and lowest in ALDHdim cells. ALDH1A2 expression was highest in ALDHvery-br cells, intermediate in ALDHdim cells, and lowest in ALDHbr cells. ALDH1A3 and ALDH2 expression was detectable in ALDHvery-br and ALDHbr cells, unlike ALDHdim cells, albeit at lower levels compared with ALDH1A1 and ALDH1A2. Freshly isolated ALDHbr cells were enriched for cells expressing stem cell antigen-1, CD34, CD90, CD44, and CD106. ALDHbr cells, unlike ALDHdim cells, could be grown in culture for more than 40 passages. They expressed sarcomeric α-actinin and could be differentiated along multiple mesenchymal lineages. However, the proportion of ALDHbr cells declined with cell passage. In conclusion, the cardiac-derived ALDHbr population is enriched for progenitor cells that exhibit mesenchymal progenitor-like characteristics and can be expanded in culture. The regenerative potential of cardiac-derived ALDHbr cells remains to be evaluated. PMID:23484127

  14. Kinetic involvement of acetaldehyde substrate inhibition on the rate equation of yeast aldehyde dehydrogenase.

    PubMed

    Eggert, Matthew W; Byrne, Mark E; Chambers, Robert P

    2012-10-01

    In order to evaluate the effectiveness of aldehyde dehydrogenase (ALDH) from Saccharomyces cerevisiae as a catalyst for the conversion of acetaldehyde into its physiologically and biologically less toxic acetate, the kinetics over broad concentrations were studied to develop a suitable kinetic rate expression. Even with literature accounts of the binding complexations, the yeast ALDH currently lacks a quantitative kinetic rate expression accounting for simultaneous inhibition parameters under higher acetaldehyde concentrations. Both substrate acetaldehyde and product NADH were observed as individual sources of inhibition with the combined effect of a ternary complex of acetaldehyde and the coenzyme leading to experimental rates as little as an eighth of the expected activity. Furthermore, the onset and strength of inhibition from each component were directly affected by the concentration of the co-substrate NAD. While acetaldehyde inhibition of ALDH is initiated below concentrations of 0.05 mM in the presence of 0.5 mM NAD or less, the acetaldehyde inhibition onset shifts to 0.2 mM with as much as 1.6 mM NAD. The convenience of the statistical software package JMP allowed for effective determination of experimental kinetic constants and simplification to a suitable rate expression: v = Vmax(AB)/(K(ia)K(b) + K(b)A + K(a)B + AB + B(2)/K(I-Ald) + B(2)Q/K(I-Ald-NADH) + BQ/K(I-NADH)) where the last three terms represent the inhibition complex terms for acetaldehyde, acetaldehyde-NADH, and NADH, respectively. The corresponding values of K(I-Ald), K(I-Ald-NADH), and K(I-NADH) for yeast ALDH are 2.55, 0.0269, and 0.162 mM at 22 °C and pH 7.8.

  15. Characterization of betaine aldehyde dehydrogenase (BetB) as an essential virulence factor of Brucella abortus.

    PubMed

    Lee, Jin Ju; Kim, Jae Hong; Kim, Dae Geun; Kim, Dong Hyeok; Simborio, Hannah Leah; Min, Won Gi; Rhee, Man Hee; Lim, Jong Hwan; Chang, Hong Hee; Kim, Suk

    2014-01-10

    The pathogenic mechanisms of Brucellosis used to adapt to the harsh intracellular environment of the host cell are not fully understood. The present study investigated the in vitro and in vivo characteristics of B. abortus betaine aldehyde dehydrogenase (BetB) (Gene Bank ID: 006932) using a betB deletion mutant constructed from virulent B. abortus 544. In test under stress conditions, including osmotic- and acid stress-resistance, the betB mutant had a lower osmotic-resistance than B. abortus wild-type. In addition, the betB mutant showed higher internalization rates compared to the wild-type strain; however, it also displayed replication failures in HeLa cells and RAW 264.7 macrophages. During internalization, compared to the wild-type strain, the betB mutant was more adherent to the host surface and showed enhanced phosphorylation of protein kinases, two processes that promote phagocytic activity, in host cells. During intracellular trafficking, colocalization of B. abortus-containing phagosomes with LAMP-1 was elevated in betB mutant-infected cells compared to the wild-type cells. In mice, the betB mutant was predominantly cleared from spleens compared to the wild-type strain after 2 weeks post-infection, and the vaccination test with the live betB mutant showed effective protection against challenge infection with the virulent wild-type strain. These findings suggested that the B. abortus betB gene substantially affects the phagocytic pathway in human phagocytes and in host cells in mice. Furthermore, this study highlights the potential use of the B. abortus betB mutant as a live vaccine for the control of brucellosis. Copyright © 2013 Elsevier B.V. All rights reserved.

  16. Development and validation of a rapid, aldehyde dehydrogenase bright–based cord blood potency assay

    PubMed Central

    Noldner, Pamela; Troy, Jesse D.; Cheatham, Lynn; Parrish, Amanda; Page, Kristin; Gentry, Tracy; Balber, Andrew E.; Kurtzberg, Joanne

    2016-01-01

    Banked, unrelated umbilical cord blood provides access to hematopoietic stem cell transplantation for patients lacking matched bone marrow donors, yet 10% to 15% of patients experience graft failure or delayed engraftment. This may be due, at least in part, to inadequate potency of the selected cord blood unit (CBU). CBU potency is typically assessed before cryopreservation, neglecting changes in potency occurring during freezing and thawing. Colony-forming units (CFUs) have been previously shown to predict CBU potency, defined as the ability to engraft in patients by day 42 posttransplant. However, the CFU assay is difficult to standardize and requires 2 weeks to perform. Consequently, we developed a rapid multiparameter flow cytometric CBU potency assay that enumerates cells expressing high levels of the enzyme aldehyde dehydrogenase (ALDH bright [ALDHbr]), along with viable CD45+ or CD34+ cell content. These measurements are made on a segment that was attached to a cryopreserved CBU. We validated the assay with prespecified criteria testing accuracy, specificity, repeatability, intermediate precision, and linearity. We then prospectively examined the correlations among ALDHbr, CD34+, and CFU content of 3908 segments over a 5-year period. ALDHbr (r = 0.78; 95% confidence interval [CI], 0.76-0.79), but not CD34+ (r = 0.25; 95% CI, 0.22-0.28), was strongly correlated with CFU content as well as ALDHbr content of the CBU. These results suggest that the ALDHbr segment assay (based on unit characteristics measured before release) is a reliable assessment of potency that allows rapid selection and release of CBUs from the cord blood bank to the transplant center for transplantation. PMID:26968535

  17. Decreased Vesicular Storage and Aldehyde Dehydrogenase Activity in Multiple System Atrophy

    PubMed Central

    Goldstein, David S.; Sullivan, Patricia; Holmes, Courtney; Kopin, Irwin J.; Sharabi, Yehonatan; Mash, Deborah C.

    2015-01-01

    Background Parkinson disease (PD) and multiple system atrophy (MSA) share some neuropathologic findings (nigrostriatal dopaminergic lesion, alpha-synuclein deposition) but not others (Lewy bodies in PD, glial cytoplasmic inclusions in MSA). In PD evidence has accrued for a vesicular storage defect and aldehyde dehydrogenase (ALDH) inhibition in residual dopaminergic terminals, resulting in accumulation of the toxic dopamine (DA) metabolite 3,4-dihydroxyphenylacetaldehyde (DOPAL). In this study we asked whether MSA entails a similar abnormal neurochemical pattern. Methods DA and its main neuronal metabolite 3,4-dihydroxyphenylacetic acid (DOPAC), norepinephrine (NE) and its main neuronal metabolite 3,4-dihydroxyphenylglycol (DHPG), the catecholamine precursor DOPA, and DOPAL were measured in striatal and frontal cortical tissue from patients with pathologically proven end-stage MSA (N=15), sporadic PD (N=17), and control subjects (N=18). Results Compared to the control group, the MSA and PD groups had similarly decreased putamen DA (by 96% and 93%, p<0.0001), DOPAC (97% and 95%, p<0.0001), NE (91% and 74%, p<0.0001), and DHPG (81% and 74%, p<0.0001). In the MSA and PD groups, ratios of DOPAL:DA were 2.3 and 3.5 times control and DHPG:NE 3.1 and 2.6 times control, while DOPAC:DOPAL ratios were decreased by 61% and 74%. In both diseases cortical NE and DHPG were decreased, while DA and DOPAC were not. Conclusions MSA and PD entail a catecholamine metabolic profile indicating impaired vesicular storage, decreased ALDH activity, and DOPAL buildup, which may be part of a common pathway in catecholamine neuronal death. Targeting this pathway by interfering with catecholaldehyde production or effects constitutes a novel treatment approach. PMID:25829070

  18. Expanded Hematopoietic Progenitor Cells Reselected for High Aldehyde Dehydrogenase Activity Demonstrate Islet Regenerative Functions.

    PubMed

    Seneviratne, Ayesh K; Bell, Gillian I; Sherman, Stephen E; Cooper, Tyler T; Putman, David M; Hess, David A

    2016-04-01

    Human umbilical cord blood (UCB) hematopoietic progenitor cells (HPC) purified for high aldehyde dehydrogenase activity (ALDH(hi) ) stimulate islet regeneration after transplantation into mice with streptozotocin-induced β cell deletion. However, ALDH(hi) cells represent a rare progenitor subset and widespread use of UCB ALDH(hi) cells to stimulate islet regeneration will require progenitor cell expansion without loss of islet regenerative functions. Here we demonstrate that prospectively purified UCB ALDH(hi) cells expand efficiently under serum-free, xeno-free conditions with minimal growth factor supplementation. Consistent with the concept that ALDH-activity is decreased as progenitor cells differentiate, kinetic analyses over 9 days revealed the frequency of ALDH(hi) cells diminished as culture time progressed such that total ALDH(hi) cell number was maximal (increased 3-fold) at day 6. Subsequently, day 6 expanded cells (bulk cells) were sorted after culture to reselect differentiated progeny with low ALDH-activity (ALDH(lo) subset) from less differentiated progeny with high ALDH-activity (ALDH(hi) subset). The ALDH(hi) subset retained primitive cell surface marker coexpression (32.0% ± 7.0% CD34(+) /CD38(-) cells, 37.0% ± 6.9% CD34(+) /CD133(+) cells), and demonstrated increased hematopoietic colony forming cell function compared with the ALDH(lo) subset. Notably, bulk cells or ALDH(lo) cells did not possess the functional capacity to lower hyperglycemia after transplantation into streptozotocin-treated NOD/SCID mice. However, transplantation of the repurified ALDH(hi) subset significantly reduced hyperglycemia, improved glucose tolerance, and increased islet-associated cell proliferation and capillary formation. Thus, expansion and delivery of reselected UCB cells that retain high ALDH-activity after short-term culture represents an improved strategy for the development of cellular therapies to enhance islet regeneration in situ.

  19. Mesenchymal glioma stem cells are maintained by activated glycolytic metabolism involving aldehyde dehydrogenase 1A3

    PubMed Central

    Mao, Ping; Joshi, Kaushal; Li, Jianfeng; Kim, Sung-Hak; Li, Peipei; Santana-Santos, Lucas; Luthra, Soumya; Chandran, Uma R.; Benos, Panayiotis V.; Smith, Luke; Wang, Maode; Hu, Bo; Cheng, Shi-Yuan; Sobol, Robert W.; Nakano, Ichiro

    2013-01-01

    Tumor heterogeneity of high-grade glioma (HGG) is recognized by four clinically relevant subtypes based on core gene signatures. However, molecular signaling in glioma stem cells (GSCs) in individual HGG subtypes is poorly characterized. Here we identified and characterized two mutually exclusive GSC subtypes with distinct dysregulated signaling pathways. Analysis of mRNA profiles distinguished proneural (PN) from mesenchymal (Mes) GSCs and revealed a pronounced correlation with the corresponding PN or Mes HGGs. Mes GSCs displayed more aggressive phenotypes in vitro and as intracranial xenografts in mice. Further, Mes GSCs were markedly resistant to radiation compared with PN GSCs. The glycolytic pathway, comprising aldehyde dehydrogenase (ALDH) family genes and in particular ALDH1A3, were enriched in Mes GSCs. Glycolytic activity and ALDH activity were significantly elevated in Mes GSCs but not in PN GSCs. Expression of ALDH1A3 was also increased in clinical HGG compared with low-grade glioma or normal brain tissue. Moreover, inhibition of ALDH1A3 attenuated the growth of Mes but not PN GSCs. Last, radiation treatment of PN GSCs up-regulated Mes-associated markers and down-regulated PN-associated markers, whereas inhibition of ALDH1A3 attenuated an irradiation-induced gain of Mes identity in PN GSCs. Taken together, our data suggest that two subtypes of GSCs, harboring distinct metabolic signaling pathways, represent intertumoral glioma heterogeneity and highlight previously unidentified roles of ALDH1A3-associated signaling that promotes aberrant proliferation of Mes HGGs and GSCs. Inhibition of ALDH1A3-mediated pathways therefore might provide a promising therapeutic approach for a subset of HGGs with the Mes signature. PMID:23650391

  20. Aldehyde dehydrogenase 3 converts farnesal into farnesoic acid in the corpora allata of mosquitoes.

    PubMed

    Rivera-Perez, Crisalejandra; Nouzova, Marcela; Clifton, Mark E; Garcia, Elena Martin; LeBlanc, Elizabeth; Noriega, Fernando G

    2013-08-01

    The juvenile hormones (JHs) play a central role in insect reproduction, development and behavior. Interrupting JH biosynthesis has long been considered a promising strategy for the development of target-specific insecticides. Using a combination of RNAi, in vivo and in vitro studies we characterized the last unknown biosynthetic enzyme of the JH pathway, a fatty aldehyde dehydrogenase (AaALDH3) that oxidizes farnesal into farnesoic acid (FA) in the corpora allata (CA) of mosquitoes. The AaALDH3 is structurally and functionally a NAD(+)-dependent class 3 ALDH showing tissue- and developmental-stage-specific splice variants. Members of the ALDH3 family play critical roles in the development of cancer and Sjögren-Larsson syndrome in humans, but have not been studies in groups other than mammals. Using a newly developed assay utilizing fluorescent tags, we demonstrated that AaALDH3 activity, as well as the concentrations of farnesol, farnesal and FA were different in CA of sugar and blood-fed females. In CA of blood-fed females the low catalytic activity of AaALDH3 limited the flux of precursors and caused a remarkable increase in the pool of farnesal with a decrease in FA and JH synthesis. The accumulation of the potentially toxic farnesal stimulated the activity of a reductase that converted farnesal back into farnesol, resulting in farnesol leaking out of the CA. Our studies indicated AaALDH3 plays a key role in the regulation of JH synthesis in blood-fed females and mosquitoes seem to have developed a "trade-off" system to balance the key role of farnesal as a JH precursor with its potential toxicity.

  1. Development and validation of a rapid, aldehyde dehydrogenase bright-based cord blood potency assay.

    PubMed

    Shoulars, Kevin; Noldner, Pamela; Troy, Jesse D; Cheatham, Lynn; Parrish, Amanda; Page, Kristin; Gentry, Tracy; Balber, Andrew E; Kurtzberg, Joanne

    2016-05-12

    Banked, unrelated umbilical cord blood provides access to hematopoietic stem cell transplantation for patients lacking matched bone marrow donors, yet 10% to 15% of patients experience graft failure or delayed engraftment. This may be due, at least in part, to inadequate potency of the selected cord blood unit (CBU). CBU potency is typically assessed before cryopreservation, neglecting changes in potency occurring during freezing and thawing. Colony-forming units (CFUs) have been previously shown to predict CBU potency, defined as the ability to engraft in patients by day 42 posttransplant. However, the CFU assay is difficult to standardize and requires 2 weeks to perform. Consequently, we developed a rapid multiparameter flow cytometric CBU potency assay that enumerates cells expressing high levels of the enzyme aldehyde dehydrogenase (ALDH bright [ALDH(br)]), along with viable CD45(+) or CD34(+) cell content. These measurements are made on a segment that was attached to a cryopreserved CBU. We validated the assay with prespecified criteria testing accuracy, specificity, repeatability, intermediate precision, and linearity. We then prospectively examined the correlations among ALDH(br), CD34(+), and CFU content of 3908 segments over a 5-year period. ALDH(br) (r = 0.78; 95% confidence interval [CI], 0.76-0.79), but not CD34(+) (r = 0.25; 95% CI, 0.22-0.28), was strongly correlated with CFU content as well as ALDH(br) content of the CBU. These results suggest that the ALDH(br) segment assay (based on unit characteristics measured before release) is a reliable assessment of potency that allows rapid selection and release of CBUs from the cord blood bank to the transplant center for transplantation. © 2016 by The American Society of Hematology.

  2. Regulation of Human Mitochondrial Aldehyde Dehydrogenase (ALDH-2) Activity by Electrophiles in Vitro*

    PubMed Central

    Oelze, Matthias; Knorr, Maike; Schell, Richard; Kamuf, Jens; Pautz, Andrea; Art, Julia; Wenzel, Philip; Münzel, Thomas; Kleinert, Hartmut; Daiber, Andreas

    2011-01-01

    Recently, mitochondrial aldehyde dehydrogenase (ALDH-2) was reported to reduce ischemic damage in an experimental myocardial infarction model. ALDH-2 activity is redox-sensitive. Therefore, we here compared effects of various electrophiles (organic nitrates, reactive fatty acid metabolites, or oxidants) on the activity of ALDH-2 with special emphasis on organic nitrate-induced inactivation of the enzyme, the biochemical correlate of nitrate tolerance. Recombinant human ALDH-2 was overexpressed in Escherichia coli; activity was determined with an HPLC-based assay, and reactive oxygen and nitrogen species formation was determined by chemiluminescence, fluorescence, protein tyrosine nitration, and diaminonaphthalene nitrosation. The organic nitrate glyceryl trinitrate caused a severe concentration-dependent decrease in enzyme activity, whereas incubation with pentaerythritol tetranitrate had only minor effects. 4-Hydroxynonenal, an oxidized prostaglandin J2, and 9- or 10-nitrooleate caused a significant inhibition of ALDH-2 activity, which was improved in the presence of Mg2+ and Ca2+. Hydrogen peroxide and NO generation caused only minor inhibition of ALDH-2 activity, whereas peroxynitrite generation or bolus additions lead to severe impairment of the enzymatic activity, which was prevented by the thioredoxin/thioredoxin reductase (Trx/TrxR) system. In the presence of glyceryl trinitrate and to a lesser extent pentaerythritol tetranitrate, ALDH-2 may be switched to a peroxynitrite synthase. Electrophiles of different nature potently regulate the enzymatic activity of ALDH-2 and thereby may influence the resistance to ischemic damage in response to myocardial infarction. The Trx/TrxR system may play an important role in this process because it not only prevents inhibition of ALDH-2 but is also inhibited by the ALDH-2 substrate 4-hydroxynonenal. PMID:21252222

  3. Aldehyde Dehydrogenase 1A1: Friend or Foe to Female Metabolism?

    PubMed Central

    Petrosino, Jennifer M.; DiSilvestro, David; Ziouzenkova, Ouliana

    2014-01-01

    In this review, we summarize recent advances in understanding vitamin A-dependent regulation of sex-specific differences in metabolic diseases, inflammation, and certain cancers. We focus on the characterization of the aldehyde dehydrogenase-1 family of enzymes (ALDH1A1, ALDH1A2, ALDH1A3) that catalyze conversion of retinaldehyde to retinoic acid. Additionally, we propose a “horizontal transfer of signaling” from estrogen to retinoids through the action of ALDH1A1. Although estrogen does not directly influence expression of Aldh1a1, it has the ability to suppress Aldh1a2 and Aldh1a3, thereby establishing a female-specific mechanism for retinoic acid generation in target tissues. ALDH1A1 regulates adipogenesis, abdominal fat formation, glucose tolerance, and suppression of thermogenesis in adipocytes; in B cells, ALDH1A1 plays a protective role by inducing oncogene suppressors Rara and Pparg. Considering the conflicting responses of Aldh1a1 in a multitude of physiological processes, only tissue-specific regulation of Aldh1a1 can result in therapeutic effects. We have shown through successful implantation of tissue-specific Aldh1a1−/− preadipocytes that thermogenesis can be induced in wild-type adipose tissues to resolve diet-induced visceral obesity in females. We will briefly discuss the emerging role of ALDH1A1 in multiple myeloma, the regulation of reproduction, and immune responses, and conclude by discussing the role of ALDH1A1 in future therapeutic applications. PMID:24594504

  4. Characterization of aldehyde dehydrogenase 1 high ovarian cancer cells: Towards targeted stem cell therapy.

    PubMed

    Sharrow, Allison C; Perkins, Brandy; Collector, Michael I; Yu, Wayne; Simons, Brian W; Jones, Richard J

    2016-08-01

    The cancer stem cell (CSC) paradigm hypothesizes that successful clinical eradication of CSCs may lead to durable remission for patients with ovarian cancer. Despite mounting evidence in support of ovarian CSCs, their phenotype and clinical relevance remain unclear. We and others have found high aldehyde dehydrogenase 1 (ALDH(high)) expression in a variety of normal and malignant stem cells, and sought to better characterize ALDH(high) cells in ovarian cancer. We compared ALDH(high) to ALDH(low) cells in two ovarian cancer models representing distinct subtypes: FNAR-C1 cells, derived from a spontaneous rat endometrioid carcinoma, and the human SKOV3 cell line (described as both serous and clear cell subtypes). We assessed these populations for stem cell features then analyzed expression by microarray and qPCR. ALDH(high) cells displayed CSC properties, including: smaller size, quiescence, regenerating the phenotypic diversity of the cell lines in vitro, lack of contact inhibition, nonadherent growth, multi-drug resistance, and in vivo tumorigenicity. Microarray and qPCR analysis of the expression of markers reported by others to enrich for ovarian CSCs revealed that ALDH(high) cells of both models showed downregulation of CD24, but inconsistent expression of CD44, KIT and CD133. However, the following druggable targets were consistently expressed in the ALDH(high) cells from both models: mTOR signaling, her-2/neu, CD47 and FGF18/FGFR3. Based on functional characterization, ALDH(high) ovarian cancer cells represent an ovarian CSC population. Differential gene expression identified druggable targets that have the potential for therapeutic efficacy against ovarian CSCs from multiple subtypes. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Serelaxin Treatment Reduces Oxidative Stress and Increases Aldehyde Dehydrogenase-2 to Attenuate Nitrate Tolerance

    PubMed Central

    Leo, Chen Huei; Fernando, Dhanushke T.; Tran, Lillie; Ng, Hooi Hooi; Marshall, Sarah A.; Parry, Laura J.

    2017-01-01

    Background: Glyceryl trinitrate (GTN) is a commonly prescribed treatment for acute heart failure patients. However, prolonged GTN treatment induces tolerance, largely due to increased oxidative stress and reduced aldehyde dehydrogenase-2 (ALDH-2) expression. Serelaxin has several vasoprotective properties, which include reducing oxidative stress and augmenting endothelial function. We therefore tested the hypothesis in rodents that serelaxin treatment could attenuate low-dose GTN-induced tolerance. Methods and Results: Co-incubation of mouse aortic rings ex vivo with GTN (10 μM) and serelaxin (10 nM) for 1 h, restored GTN responses, suggesting that serelaxin prevented the development of GTN tolerance. Male Wistar rats were subcutaneously infused with ethanol (control), low-dose GTN+placebo or low-dose GTN+serelaxin via osmotic minipumps for 3 days. Aortic vascular function and superoxide levels were assessed using wire myography and lucigenin-enhanced chemiluminescence assay respectively. Changes in aortic ALDH-2 expression were measured by qPCR and Western blot respectively. GTN+placebo infusion significantly increased superoxide levels, decreased ALDH-2 and attenuated GTN-mediated vascular relaxation. Serelaxin co-treatment with GTN significantly enhanced GTN-mediated vascular relaxation, reduced superoxide levels and increased ALDH-2 expression compared to GTN+placebo-treated rats. Conclusion: Our data demonstrate that a combination of serelaxin treatment with low dose GTN attenuates the development of GTN-induced tolerance by reducing superoxide production and increasing ALDH-2 expression in the rat aorta. We suggest that serelaxin may improve nitrate efficacy in a clinical setting. PMID:28377719

  6. Ethanol reduces lifespan, body weight, and serum alanine aminotransferase level of aldehyde dehydrogenase 2 knockout mouse.

    PubMed

    Matsumoto, Akiko; Vasiliou, Vasilis; Kawamoto, Toshihiro; Tanaka, Keitaro; Ichiba, Masayoshi

    2014-07-01

    The aldehyde dehydrogenase 2 (Aldh2) knockout mouse is an animal model of a polymorphism at the human ALDH2 locus (ALDH2*2). To detect differences in the basic phenotype of this animal model, lifespan, body weight (BW), and serum alanine aminotransferase (ALT) level were evaluated. Aldh2(+/+) , Aldh2(+/-) , and Aldh2(-/-) mice were maintained, from 10 weeks of age, on standard solid food, with liquid supplied as ethanol (EtOH) solution at a concentration of 0 to 20% (forced EtOH consumption). For animals provided with water (without EtOH), mice of the distinct genotypes exhibited no difference in lifespan, with the mean values ranging from 90 to 96 weeks for female mice and 97 to 105 weeks for male mice. For animals provided with EtOH, there was a dose-dependent reduction of lifespan in Aldh2(-/-) mice with p for trend <0.001. For example, the mean lifespans of the Aldh2(-/-) females in the 0, 3, 10, and 20% groups were 95, 85, 70, and 29 weeks, respectively. No influence on lifespan was found for Aldh2(+/+) and Aldh2(+/-) mice. BW and ALT level of Aldh2(-/-) mice were significantly lower than those of Aldh2(+/+) mice when the mice were treated with EtOH. While multiple regression analysis suggested that the BW and ALT level in Aldh2(-/-) mice correlated with lifespan, adjustment for EtOH concentration revealed that this correlation was not significant (i.e., reflected EtOH dependence). Aldh2(-/-) mice were unchanged in terms of their basic phenotype under standard laboratory conditions. However, chronic EtOH administration (forced consumption) in these mice resulted in dose-dependent reductions in lifespan, BW, and serum ALT level. Copyright © 2014 by the Research Society on Alcoholism.

  7. Mitochondrial aldehyde dehydrogenase 2 deficiency aggravates energy metabolism disturbance and diastolic dysfunction in diabetic mice.

    PubMed

    Wang, Cong; Fan, Fan; Cao, Quan; Shen, Cheng; Zhu, Hong; Wang, Peng; Zhao, Xiaona; Sun, Xiaolei; Dong, Zhen; Ma, Xin; Liu, Xiangwei; Han, Shasha; Wu, Chaoneng; Zou, Yunzeng; Hu, Kai; Ge, Junbo; Sun, Aijun

    2016-11-01

    Diabetes causes energy metabolism disturbance and may lead to cardiac dysfunction. Mitochondrial aldehyde dehydrogenase 2 (ALDH2) protects cardiac function from myocardial damage. Therefore, understanding of its roles in diabetic heart is critical for developing new therapeutics targeting ALDH2 and mitochondrial function for diabetic hearts. This study investigated the impact of ALDH2 deficiency on diastolic function and energy metabolism in diabetic mice. Diabetes was induced in ALDH2 knockout and wild-type mice by streptozotocin. Cardiac function was determined by echocardiography. Glucose uptake, energy status, and metabolic profiles were used to evaluate cardiac energy metabolism. The association between ALDH2 polymorphism and diabetes was also analyzed in patients. Echocardiography revealed preserved systolic function and impaired diastolic function in diabetic ALDH2-deficient mice. Energy reserves (phosphocreatine/adenosine triphosphate ratio) were reduced in the diabetic mutants and were associated with diastolic dysfunction. Western blot analysis showed that diabetes induces accumulated lipid peroxidation products and escalated AMP-activated protein kinase-LKB1 pathway. Further, ALDH2 deficiency exacerbated the diabetes-induced deficient myocardial glucose uptake and other perturbations of metabolic profiles. Finally, ALDH2 mutations were associated with worse diastolic dysfunction in diabetic patients. Together, our results demonstrate that ALDH2 deficiency and resulting energy metabolism disturbance is a part of pathology of diastolic dysfunction of diabetic hearts, and suggest that patients with ALDH2 mutations are vulnerable to diabetic damage. ALDH2 deficiency exacerbates diastolic dysfunction in early diabetic hearts. ALDH2 deficiency triggers decompensation of metabolic reserves and energy metabolism disturbances in early diabetic hearts. ALDH2 deficiency potentiates oxidative stress and AMPK phosphorylation induced by diabetes via post

  8. Proteomic alteration of mitochondrial aldehyde dehydrogenase 2 in sepsis regulated by heat shock response.

    PubMed

    Chen, Hsiang-Wen; Kuo, Hung-Tien; Hwang, Long-Chih; Kuo, Mei-Fang; Yang, Rei-Cheng

    2007-12-01

    The present study was designed to investigate the proteomic alteration of hepatic mitochondria during sepsis and to explore the possible effects induced by heat shock treatment. Sepsis was induced by cecal ligation and puncture in Sprague-Dawley rats. Liver mitochondrial proteins were isolated and evaluated by 2-dimensional electrophoresis with broad pH-ranged (pH 3 - 10) immobile DryStrip and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The protein spots were visualized with silver stain and analyzed by Bio-2D software. Results showed that around 120 dominant spots could be separated and visualized distinctly by 2-dimensional electrophoresis analysis. Among them, three spots with the same molecular weight (56.4 kd), mitochondrial protein 1 (MP1), MP2, and MP3, were significantly altered in septic specimens. When analyzed by liquid chromatography-tandem mass spectrometry, the three spots all revealed to be an identical enzyme: aldehyde dehydrogenase 2 (ALDH2, EC 1.2.1.3). During sepsis, MP1 and MP2 were downregulated, whereas MP3 was upregulated concomitantly. Interestingly, heat shock treatment could reverse this phenomenon. Phosphoprotein staining showed that the degree of phosphorylation is higher in MP1 and MP2 than that in MP3. The enzyme activity assay showed that ALDH2 activity was downregulated in nonheated septic rats of 18 h after cecal ligation and puncture operation, and preserved in heated septic rats. The results of this study suggest that posttranslation modification, highly possible the phosphorylation, in ALDH2 may play a functional role in the pathogenesis of sepsis and provide a novel protective mechanism of heat shock treatment.

  9. High current density PQQ-dependent alcohol and aldehyde dehydrogenase bioanodes.

    PubMed

    Aquino Neto, Sidney; Hickey, David P; Milton, Ross D; De Andrade, Adalgisa R; Minteer, Shelley D

    2015-10-15

    In this paper, we explore the bioelectrooxidation of ethanol using pyrroloquinoline quinone (PQQ)-dependent alcohol and aldehyde dehydrogenase (ADH and AldDH) enzymes for biofuel cell applications. The bioanode architectures were designed with both direct electron transfer (DET) and mediated electron transfer (MET) mechanisms employing high surface area materials such as multi-walled carbon nanotubes (MWCNTs) and MWCNT-decorated gold nanoparticles, along with different immobilization techniques. Three different polymeric matrices were tested (tetrabutyl ammonium bromide (TBAB)-modified Nafion; octyl-modified linear polyethyleneimine (C8-LPEI); and cellulose) in the DET studies. The modified Nafion membrane provided the best electrical communication between enzymes and the electrode surface, with catalytic currents as high as 16.8 ± 2.1 µA cm(-2). Then, a series of ferrocene redox polymers were evaluated for MET. The redox polymer 1,1'-dimethylferrocene-modified linear polyethyleneimine (FcMe2-C3-LPEI) provided the best electrochemical response. Using this polymer, the electrochemical assays conducted in the presence of MWCNTs and MWCNTs-Au indicated a Jmax of 781 ± 59 µA cm(-2) and 925 ± 68 µA cm(-2), respectively. Overall, from the results obtained here, DET using the PQQ-dependent ADH and AldDH still lacks high current density, while the bioanodes that operate via MET employing ferrocene-modified LPEI redox polymers show efficient energy conversion capability in ethanol/air biofuel cells. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Inhibition of human alcohol and aldehyde dehydrogenases by acetaminophen: Assessment of the effects on first-pass metabolism of ethanol.

    PubMed

    Lee, Yung-Pin; Liao, Jian-Tong; Cheng, Ya-Wen; Wu, Ting-Lun; Lee, Shou-Lun; Liu, Jong-Kang; Yin, Shih-Jiun

    2013-11-01

    Acetaminophen is one of the most widely used over-the-counter analgesic, antipyretic medications. Use of acetaminophen and alcohol are commonly associated. Previous studies showed that acetaminophen might affect bioavailability of ethanol by inhibiting gastric alcohol dehydrogenase (ADH). However, potential inhibitions by acetaminophen of first-pass metabolism (FPM) of ethanol, catalyzed by the human ADH family and by relevant aldehyde dehydrogenase (ALDH) isozymes, remain undefined. ADH and ALDH both exhibit racially distinct allozymes and tissue-specific distribution of isozymes, and are principal enzymes responsible for ethanol metabolism in humans. In this study, we investigated acetaminophen inhibition of ethanol oxidation with recombinant human ADH1A, ADH1B1, ADH1B2, ADH1B3, ADH1C1, ADH1C2, ADH2, and ADH4, and inhibition of acetaldehyde oxidation with recombinant human ALDH1A1 and ALDH2. The investigations were done at near physiological pH 7.5 and with a cytoplasmic coenzyme concentration of 0.5 mM NAD(+). Acetaminophen acted as a noncompetitive inhibitor for ADH enzymes, with the slope inhibition constants (Kis) ranging from 0.90 mM (ADH2) to 20 mM (ADH1A), and the intercept inhibition constants (Kii) ranging from 1.4 mM (ADH1C allozymes) to 19 mM (ADH1A). Acetaminophen exhibited noncompetitive inhibition for ALDH2 (Kis = 3.0 mM and Kii = 2.2 mM), but competitive inhibition for ALDH1A1 (Kis = 0.96 mM). The metabolic interactions between acetaminophen and ethanol/acetaldehyde were assessed by computer simulation using inhibition equations and the determined kinetic constants. At therapeutic to subtoxic plasma levels of acetaminophen (i.e., 0.2-0.5 mM) and physiologically relevant concentrations of ethanol (10 mM) and acetaldehyde (10 μm) in target tissues, acetaminophen could inhibit ADH1C allozymes (12-26%) and ADH2 (14-28%) in the liver and small intestine, ADH4 (15-31%) in the stomach, and ALDH1A1 (16-33%) and ALDH2 (8.3-19%) in all 3 tissues. The

  11. Insight into Coenzyme A cofactor binding and the mechanism of acyl-transfer in an acylating aldehyde dehydrogenase from Clostridium phytofermentans

    PubMed Central

    Tuck, Laura R.; Altenbach, Kirsten; Ang, Thiau Fu; Crawshaw, Adam D.; Campopiano, Dominic J.; Clarke, David J.; Marles-Wright, Jon

    2016-01-01

    The breakdown of fucose and rhamnose released from plant cell walls by the cellulolytic soil bacterium Clostridium phytofermentans produces toxic aldehyde intermediates. To enable growth on these carbon sources, the pathway for the breakdown of fucose and rhamnose is encapsulated within a bacterial microcompartment (BMC). These proteinaceous organelles sequester the toxic aldehyde intermediates and allow the efficient action of acylating aldehyde dehydrogenase enzymes to produce an acyl-CoA that is ultimately used in substrate-level phosphorylation to produce ATP. Here we analyse the kinetics of the aldehyde dehydrogenase enzyme from the fucose/rhamnose utilisation BMC with different short-chain fatty aldehydes and show that it has activity against substrates with up to six carbon atoms, with optimal activity against propionaldehyde. We have also determined the X-ray crystal structure of this enzyme in complex with CoA and show that the adenine nucleotide of this cofactor is bound in a distinct pocket to the same group in NAD+. This work is the first report of the structure of CoA bound to an aldehyde dehydrogenase enzyme and our crystallographic model provides important insight into the differences within the active site that distinguish the acylating from non-acylating aldehyde dehydrogenase enzymes. PMID:26899032

  12. Saturation transfer difference NMR studies on substrates and inhibitors of succinic semialdehyde dehydrogenases

    SciTech Connect

    Jaeger, Martin Rothacker, Boris; Ilg, Thomas

    2008-08-01

    Saturation transfer difference (STD) NMR experiments on Escherichia coli and Drosophila melanogaster succinic semialdehyde dehydrogenase (SSADH, EC1.2.1.24) suggest that only the aldehyde forms and not the gem-diol forms of the specific substrate succinic semialdehyde (SSA), of selected aldehyde substrates, and of the inhibitor 3-tolualdehyde bind to these enzymes. Site-directed mutagenesis of the active site cysteine311 to alanine in D. melanogaster SSADH leads to an inactive product binding both SSA aldehyde and gem-diol. Thus, the residue cysteine311 is crucial for their discrimination. STD experiments on SSADH and NAD{sup +}/NADP{sup +} indicate differential affinity in agreement with the respective cosubstrate properties. Epitope mapping by STD points to a strong interaction of the NAD{sup +}/NADP{sup +} adenine H2 proton with SSADH. Adenine H8, nicotinamide H2, H4, and H6 also show STD signals. Saturation transfer to the ribose moieties is limited to the anomeric protons of E. coli SSADH suggesting that the NAD{sup +}/NADP{sup +} adenine and nicotinamide, but not the ribose moieties are important for the binding of the coenzymes.

  13. The metabolism of ethanol-derived acetaldehyde by alcohol dehydrogenase (EC 1.1.1.1) and aldehyde dehydrogenase (EC 1.2.1.3) in Drosophila melanogaster larvae.

    PubMed Central

    Heinstra, P W; Geer, B W; Seykens, D; Langevin, M

    1989-01-01

    Both aldehyde dehydrogenase (ALDH, EC 1.2.1.3) and the aldehyde dehydrogenase activity of alcohol dehydrogenase (ADH, EC 1.1.1.1) were found to coexist in Drosophila melanogaster larvae. The enzymes, however, showed different inhibition patterns with respect to pyrazole, cyanamide and disulphiram. ALDH-1 and ALDH-2 isoenzymes were detected in larvae by electrophoretic methods. Nonetheless, in tracer studies in vivo, more than 75% of the acetaldehyde converted to acetate by the ADH ethanol-degrading pathway appeared to be also catalysed by the ADH enzyme. The larval fat body probably was the major site of this pathway. Images Fig. 1. Fig. 2. PMID:2499314

  14. Induction of mitochondrial aldehyde dehydrogenase by submergence facilitates oxidation of acetaldehyde during re-aeration in rice.

    PubMed

    Tsuji, Hiroyuki; Meguro, Naoki; Suzuki, Yasuhiro; Tsutsumi, Nobuhiro; Hirai, Atsushi; Nakazono, Mikio

    2003-07-10

    Post-hypoxic injuries in plants are primarily caused by bursts of reactive oxygen species and acetaldehyde. In agreement with previous studies, we found accumulations of acetaldehyde in rice during re-aeration following submergence. During re-aeration, acetaldehyde-oxidizing aldehyde dehydrogenase (ALDH) activity increased, thereby causing the acetaldehyde content to decrease in rice. Interestingly, re-aerated rice plants showed an intense mitochondrial ALDH2a protein induction, even though ALDH2a mRNA was submergence induced and declined upon re-aeration. This suggests that rice ALDH2a mRNA is accumulated in order to quickly metabolize acetaldehyde that is produced upon re-aeration.

  15. Aldehyde dehydrogenase activity selects for the holoclone phenotype in prostate cancer cells

    SciTech Connect

    Doherty, R.E.; Haywood-Small, S.L.; Sisley, K.; Cross, N.A.

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer Isolated ALDH{sup Hi} PC3 cells preferentially form primitive holoclone-type colonies. Black-Right-Pointing-Pointer Primitive holoclone colonies are predominantly ALDH{sup Lo} but contain rare ALDH{sup Hi} cells. Black-Right-Pointing-Pointer Holoclone-forming cells are not restricted to the ALDH{sup Hi} population. Black-Right-Pointing-Pointer ALDH phenotypic plasticity occurs in PC3 cells (ALDH{sup Lo} to ALDH{sup Hi} and vice versa). Black-Right-Pointing-Pointer ALDH{sup Hi} cells are observed but very rare in PC3 spheroids grown in stem cell medium. -- Abstract: Aldehyde dehydrogenase 1 (ALDH) activity is considered to be a marker of cancer stem cells (CSCs) in many tumour models, since these cells are more proliferative and tumourigenic than ALDH{sup Lo} cells in experimental models. However it is unclear whether all CSC-like cells are within the ALDH{sup Hi} population, or whether all ALDH{sup Hi} cells are highly proliferative and tumourigenic. The ability to establish a stem cell hierarchy in vitro, whereby sub-populations of cells have differing proliferative and differentiation capacities, is an alternate indication of the presence of stem cell-like populations within cell lines. In this study, we have examined the interaction between ALDH status and the ability to establish a stem cell hierarchy in PC3 prostate cancer cells. We demonstrate that PC3 cells contain a stem cell hierarchy, and isolation of ALDH{sup Hi} cells enriches for the most primitive holoclone population, however holoclone formation is not restricted to ALDH{sup Hi} cells. In addition, we show that ALDH activity undergoes phenotypic plasticity, since the ALDH{sup Lo} population can develop ALDH{sup Hi} populations comparable to parental cells within 2 weeks in culture. Furthermore, we show that the majority of ALDH{sup Hi} cells are found within the least primitive paraclone population, which is circumvented by culturing PC3 cells as spheroids in

  16. Activation of Human Salivary Aldehyde Dehydrogenase by Sulforaphane: Mechanism and Significance

    PubMed Central

    Alam, Md. Fazle; Laskar, Amaj Ahmed; Maryam, Lubna

    2016-01-01

    Cruciferous vegetables contain the bio-active compound sulforaphane (SF) which has been reported to protect individuals against various diseases by a number of mechanisms, including activation of the phase II detoxification enzymes. In this study, we show that the extracts of five cruciferous vegetables that we commonly consume and SF activate human salivary aldehyde dehydrogenase (hsALDH), which is a very important detoxifying enzyme in the mouth. Maximum activation was observed at 1 μg/ml of cabbage extract with 2.6 fold increase in the activity. There was a ~1.9 fold increase in the activity of hsALDH at SF concentration of ≥ 100 nM. The concentration of SF at half the maximum response (EC50 value) was determined to be 52 ± 2 nM. There was an increase in the Vmax and a decrease in the Km of the enzyme in the presence of SF. Hence, SF interacts with the enzyme and increases its affinity for the substrate. UV absorbance, fluorescence and CD studies revealed that SF binds to hsALDH and does not disrupt its native structure. SF binds with the enzyme with a binding constant of 1.23 x 107 M-1. There is one binding site on hsALDH for SF, and the thermodynamic parameters indicate the formation of a spontaneous strong complex between the two. Molecular docking analysis depicted that SF fits into the active site of ALDH3A1, and facilitates the catalytic mechanism of the enzyme. SF being an antioxidant, is very likely to protect the catalytic Cys 243 residue from oxidation, which leads to the increase in the catalytic efficiency and hence the activation of the enzyme. Further, hsALDH which is virtually inactive towards acetaldehyde exhibited significant activity towards it in the presence of SF. It is therefore very likely that consumption of large quantities of cruciferous vegetables or SF supplements, through their activating effect on hsALDH can protect individuals who are alcohol intolerant against acetaldehyde toxicity and also lower the risk of oral cancer

  17. Changes in aldehyde dehydrogenase-1 expression during neoadjuvant chemotherapy predict outcome in locally advanced breast cancer

    PubMed Central

    2014-01-01

    Introduction Although neoadjuvant chemotherapy (NAC) for locally advanced breast cancer can improve operability and local disease control, there is a lack of reliable biomarkers that predict response to chemotherapy or long-term survival. Since expression of aldehyde dehydrogenase-1 (ALDH1) is associated with the stem-like properties of self-renewal and innate chemoresistance in breast cancer, we asked whether expression in serial tumor samples treated with NAC could identify women more likely to benefit from this therapy. Methods Women with locally advanced breast cancer were randomly assigned to receive four cycles of anthracycline-based chemotherapy, followed by four cycles of taxane therapy (Arm A), or the same regimen in reverse order (Arm B). Tumor specimens were collected at baseline, after four cycles, and then at surgical resection. ALDH1 expression was determined by immunohistochemistry and correlated with tumor response using Fisher’s exact test while Kaplan-Meier method was used to calculate survival. Results A hundred and nineteen women were enrolled into the study. Fifty seven (48%) were randomized to Arm A and 62 (52%) to Arm B. Most of the women (90%) had ductal carcinoma and 10% had lobular carcinoma. Of these, 26 (22%) achieved a pathological complete response (pCR) after NAC. There was no correlation between baseline ALDH1 expression and tumor grade, stage, hormone receptor, human epidermal growth factor receptor 2 (HER2) status and Ki67 index. ALDH1 negativity at baseline was significantly associated with pCR (P = 0.004). The presence of ALDH1(+) cells in the residual tumor cells in non-responding women was strongly predictive of worse overall survival (P = 0.024). Moreover, serial analysis of specimens from non-responders showed a marked increase in tumor-specific ALDH1 expression (P = 0.028). Overall, there was no survival difference according to the chemotherapy sequence. However, poorly responding tumours from women receiving

  18. Aldehyde dehydrogenase (ALDH) 3A1 expression by the human keratocyte and its repair phenotypes.

    PubMed

    Pei, Ying; Reins, Rose Y; McDermott, Alison M

    2006-11-01

    Transparency is essential for normal corneal function. Recent studies suggest that corneal cells express high levels of so-called corneal crystallins, such as aldehyde dehydrogenase (ALDH) and transketolase (TKT) that contribute to maintaining cellular transparency. Stromal injury leads to the appearance of repair phenotype keratocytes, the corneal fibroblast and myofibroblast. Previous studies on keratocytes from species such as bovine and rabbit indicate that the transformation from the normal to repair phenotype is accompanied by a loss of corneal crystallin expression, which may be associated with loss of cellular transparency. Here we investigated if a similar loss occurs with human keratocyte repair phenotypes. Human corneal epithelial cells were collected by scraping and keratocytes were isolated by collagenase digestion from cadaveric corneas. The cells were either processed immediately (freshly isolated keratocytes) or were cultured in the presence of 10% fetal bovine serum or transforming growth factor-beta to induce transformation to the corneal fibroblast and myofibroblast phenotypes, respectively. RT-PCR, western blotting and immunolabeling were used to detect mRNA and protein expression of ALDH isozymes and TKT. ALDH enzyme activity was also quantitated and immunolabeling was performed to determine the expression of ALDH3A1 in human corneal tissue sections from normal and diseased corneas. Human corneal keratocytes isolated from three donors expressed ALDH1A1 and ALDH3A1 mRNA, and one donor also expressed ALDH2 and TKT. Corneal epithelial cells expressed ALDH1A1, ALDH2, ALDH3A1 and TKT. Compared to normal keratocytes, corneal fibroblast expression of ALDH3A1 mRNA was reduced by 27% (n=5). ALDH3A1 protein expression as detected by western blotting was markedly reduced in passage zero fibroblasts and undetectable in higher passages (n=3). TKT protein expression was reduced in fibroblasts compared to keratocytes (n=2). ALDH3A1 enzyme activity was not

  19. Aldehyde dehydrogenase 1 a1 regulates energy metabolism in adipocytes from different species.

    PubMed

    Yang, Kefeng; Adin, Christopher; Shen, Qiwen; Lee, Ly James; Yu, Lianbo; Fadda, Paolo; Samogyi, Arpad; Ham, Kathleen; Xu, Lu; Gilor, Chen; Ziouzenkova, Ouliana

    2017-09-01

    Survival and longevity of xenotransplants depend on immune function and ability to integrate energy metabolism between cells from different species. However, mechanisms for interspecies cross talk in energy metabolism are not well understood. White adipose tissue stores energy and is capable of mobilization and dissipation of energy as heat (thermogenesis) by adipocytes expressing uncoupling protein 1 (Ucp1). Both pathways are under the control of vitamin A metabolizing enzymes. Deficient retinoic acid production in aldehyde dehydrogenase 1 A1 (Aldh1a1) knockout adipocytes (KO) inhibits adipogenesis and increases thermogenesis. Here we test the role Aldh1a1 in regulation of lipid metabolism in xenocultures. Murine wide-type (WT) and KO pre-adipocytes were encapsulated into a poly-L-lysine polymer that allows exchange of humoral factors <32kD via nanopores. Encapsulated murine adipocytes were co-incubated with primary differentiated canine adipocytes. Then, expression of adipogenic and thermogenic genes in differentiated canine adipocytes was detected by real-time polymerase chain reaction (PCR). The regulatory factors in WT and KO cells were identified by comparison of secretome using proteomics and in transcriptome by gene microarray. Co-culture of encapsulated mouse KO vs WT adipocytes increased expression of peroxisome proliferator-activated receptor gamma (Pparg), but reduced expression of its target genes fatty acid binding protein 4 (Fabp4), and adipose triglyceride lipase (Atgl) in canine adipocytes, suggesting inhibition of PPARγ activation. Co-culture with KO adipocytes also induced expression of Ucp1 in canine adipocytes compared to expression in WT adipocytes. Cumulatively, murine KO compared to WT adipocytes decreased lipid accumulation in canine adipocytes. Comparative proteomics revealed significantly higher levels of vitamin A carriers, retinol binding protein 4 (RBP4), and lipokalin 2 (LCN2) in KO vs WT adipocytes. Our data demonstrate the

  20. Dioxin receptor regulates aldehyde dehydrogenase to block melanoma tumorigenesis and metastasis.

    PubMed

    Contador-Troca, María; Alvarez-Barrientos, Alberto; Merino, Jaime M; Morales-Hernández, Antonio; Rodríguez, María I; Rey-Barroso, Javier; Barrasa, Eva; Cerezo-Guisado, María I; Catalina-Fernández, Inmaculada; Sáenz-Santamaría, Javier; Oliver, Francisco J; Fernandez-Salguero, Pedro M

    2015-08-05

    The dioxin (AhR) receptor can have oncogenic or tumor suppressor activities depending on the phenotype of the target cell. We have shown that AhR knockdown promotes melanoma primary tumorigenesis and lung metastasis in the mouse and that human metastatic melanomas had reduced AhR levels with respect to benign nevi. Mouse melanoma B16F10 cells were engineered by retroviral transduction to stably downregulate AhR expression, Aldh1a1 expression or both. They were characterized for Aldh1a1 activity, stem cell markers and migration and invasion in vitro. Their tumorigenicity in vivo was analyzed using xenografts and lung metastasis assays as well as in vivo imaging. Depletion of aldehyde dehydrogenase 1a1 (Aldh1a1) impairs the pro-tumorigenic and pro-metastatic advantage of melanoma cells lacking AhR expression (sh-AhR). Thus, Aldh1a1 knockdown in sh-AhR cells (sh-AhR + sh-Aldh1a1) diminished their migration and invasion potentials and blocked tumor growth and metastasis to the lungs in immunocompetent AhR+/+ recipient mice. However, Aldh1a1 downmodulation in AhR-expressing B16F10 cells did not significantly affect tumor growth in vivo. Aldh1a1 knockdown reduced the high levels of CD133(+)/CD29(+)/CD44(+) cells, melanosphere size and the expression of the pluripotency marker Sox2 in sh-AhR cells. Interestingly, Sox2 increased Aldh1a1 expression in sh-AhR but not in sh-AhR + sh-Aldh1a1 cells, suggesting that Aldh1a1 and Sox2 may be co-regulated in melanoma cells. In vivo imaging revealed that mice inoculated with AhR + Aldh1a1 knockdown cells had reduced tumor burden and enhanced survival than those receiving Aldh1a1-expressing sh-AhR cells. Aldh1a1 overactivation in an AhR-deficient background enhances melanoma progression. Since AhR may antagonize the protumoral effects of Aldh1a1, the AhR(low)-Aldh1a1(high) phenotype could be indicative of bad outcome in melanoma.

  1. Substrate specificity, substrate channeling, and allostery in BphJ: an acylating aldehyde dehydrogenase associated with the pyruvate aldolase BphI.

    PubMed

    Baker, Perrin; Carere, Jason; Seah, Stephen Y K

    2012-06-05

    BphJ, a nonphosphorylating acylating aldehyde dehydrogenase, catalyzes the conversion of aldehydes to form acyl-coenzyme A in the presence of NAD(+) and coenzyme A (CoA). The enzyme is structurally related to the nonacylating aldehyde dehydrogenases, aspartate-β-semialdehyde dehydrogenase and phosphorylating glyceraldehyde-3-phosphate dehydrogenase. Cys-131 was identified as the catalytic thiol in BphJ, and pH profiles together with site-specific mutagenesis data demonstrated that the catalytic thiol is not activated by an aspartate residue, as previously proposed. In contrast to the wild-type enzyme that had similar specificities for two- or three-carbon aldehydes, an I195A variant was observed to have a 20-fold higher catalytic efficiency for butyraldehyde and pentaldehyde compared to the catalytic efficiency of the wild type toward its natural substrate, acetaldehyde. BphJ forms a heterotetrameric complex with the class II aldolase BphI that channels aldehydes produced in the aldol cleavage reaction to the dehydrogenase via a molecular tunnel. Replacement of Ile-171 and Ile-195 with bulkier amino acid residues resulted in no more than a 35% reduction in acetaldehyde channeling efficiency, showing that these residues are not critical in gating the exit of the channel. Likewise, the replacement of Asn-170 in BphJ with alanine and aspartate did not substantially alter aldehyde channeling efficiencies. Levels of activation of BphI by BphJ N170A, N170D, and I171A were reduced by ≥3-fold in the presence of NADH and ≥4.5-fold when BphJ was undergoing turnover, indicating that allosteric activation of the aldolase has been compromised in these variants. The results demonstrate that the dehydrogenase coordinates the catalytic activity of BphI through allostery rather than through aldehyde channeling.

  2. Alcohol and Aldehyde Dehydrogenases Contribute to Sex-Related Differences in Clearance of Zolpidem in Rats

    PubMed Central

    Peer, Cody J.; Strope, Jonathan D.; Beedie, Shaunna; Ley, Ariel M.; Holly, Alesia; Calis, Karim; Farkas, Ronald; Parepally, Jagan; Men, Angela; Fadiran, Emmanuel O.; Scott, Pamela; Jenkins, Marjorie; Theodore, William H.; Sissung, Tristan M.

    2016-01-01

    Objectives: The recommended zolpidem starting dose was lowered in females (5 mg vs. 10 mg) since side effects were more frequent and severe than those of males; the mechanism underlying sex differences in pharmacokinetics (PK) is unknown. We hypothesized that such differences were caused by known sex-related variability in alcohol dehydrogenase (ADH) expression. Methods: Male, female, and castrated male rats were administered 2.6 mg/kg zolpidem, ± disulfiram (ADH/ALDH pathway inhibitor) to compare PK changes induced by sex and gonadal hormones. PK analyses were conducted in rat plasma and rat brain. Key findings: Sex differences in PK were evident: females had a higher CMAX (112.4 vs. 68.1 ug/L) and AUC (537.8 vs. 231.8 h∗ug/L) than uncastrated males. Castration induced an earlier TMAX (0.25 vs. 1 h), greater CMAX (109.1 vs. 68.1 ug/L), and a corresponding AUC increase (339.7 vs. 231.8 h∗ug/L). Administration of disulfiram caused more drastic CMAX and TMAX changes in male vs. female rats that mirrored the effects of castration on first-pass metabolism, suggesting that the observed PK differences may be caused by ADH/ALDH expression. Brain concentrations paralleled plasma concentrations. Conclusion: These findings indicate that sex differences in zolpidem PK are influenced by variation in the expression of ADH/ALDH due to gonadal androgens. PMID:27574509

  3. Peptidyl aldehydes as slow-binding inhibitors of dual-specificity phosphatases.

    PubMed

    Park, Junguk; Fu, Hua; Pei, Dehua

    2004-02-09

    Peptidyl aldehydes were tested for inhibition of dual-specificity phosphatases VH1 and VHR. The most potent compound, cinnamaldehyde-Gly-Glu-Glu (Cinn-GEE), acted as a slow-binding inhibitor with K(I)* values of 18 and 288 microM against VH1 and VHR, respectively.

  4. Involvement of alcohol and aldehyde dehydrogenase activities on hepatic retinoid metabolism and its possible participation in the progression of rat liver regeneration.

    PubMed

    López-Valencia, Verónica; Rangel, Pablo; Rodríguez, Sandra; Hernández-Muñoz, Rolando

    2007-02-15

    Liver alcohol dehydrogenase (ADH) activity is decreased towards exogenous substrates after partial hepatectomy (PH), probably due to putative endogenous substrates acting as ADH inhibitors. Hence, retinoids could be suitable candidates as such endogenous substrates. Therefore, cytosolic ADH kinetic analysis using several substrates, liver cytosolic and mitochondrial aldehyde dehydrogenase (ALDH) activities, retinal and retinol content, as well as expression of proteins for ADH and CRBPI (a retinol carrier protein) were determined in liver samples, at two stages of liver regeneration (one- or two-thirds PH). The effect of inhibiting in vivo liver ADH by 4-methylpyrazole (4-MP) was also evaluated after 70%-PH. With 70%-PH, in vitro ADH activity towards exogenous alcohols and aldehydes was diminished, but retinol oxidation was increased and retinal reduction was decreased. These activities that be due to the participation of an ADH type which did not correlate with the amount of immunoreactive ADH protein. Cytosolic and mitochondrial ALDH activities oxidized actively retinal, whereas retinol and CBRP-I expression were reduced in these animals. With 30%-PH, these changes were less evident and sometimes opposite to those found with 70%-PH. In addition, retinol readily inhibited ADH-mediated ethanol oxidation. Interestingly, in vivo 4-MP administration inhibited ADH activity in a dose-dependent manner correlating with a progressive inhibition of liver regeneration. In conclusion, PH-induced inhibition of ADH (mainly type I) seems to be related to ADH-mediated retinoid metabolism during liver proliferation. Thus, results suggest a role of ADH in retinoid metabolism, which is apparently required during rat liver regeneration.

  5. Phytoestrogens as inhibitors of fungal 17beta-hydroxysteroid dehydrogenase.

    PubMed

    Kristan, Katja; Krajnc, Katja; Konc, Janez; Gobec, Stanislav; Stojan, Jure; Rizner, Tea Lanisnik

    2005-09-01

    Different phytoestrogens were tested as inhibitors of 17beta-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17beta-HSDcl), a member of the short-chain dehydrogenase/reductase superfamily. Phytoestrogens inhibited the oxidation of 100 microM 17beta-hydroxyestra-4-en-3-one and the reduction of 100 microM estra-4-en-3,17-dione, the best substrate pair known. The best inhibitors of oxidation, with IC(50) below 1 microM, were flavones hydroxylated at positions 3, 5 and 7: 3-hydroxyflavone, 3,7-dihydroxyflavone, 5,7-dihydroxyflavone (chrysin) and 5-hydroxyflavone, together with 5-methoxyflavone. The best inhibitors of reduction were less potent; 3-hydroxyflavone, 5-methoxyflavone, coumestrol, 3,5,7,4'-tetrahydroxyflavone (kaempferol) and 5-hydroxyflavone all had IC(50) values between 1 and 5 microM. Docking the representative inhibitors chrysin and kaempferol into the active site of 17beta-HSDcl revealed the possible binding mode, in which they are sandwiched between the nicotinamide moiety and Tyr212. The structural features of phytoestrogens, inhibitors of both oxidation and reduction catalyzed by the fungal 17beta-HSD, are similar to the reported structural features of phytoestrogen inhibitors of human 17beta-HSD types 1 and 2.

  6. Phytoestrogens as inhibitors of fungal 17beta-hydroxysteroid dehydrogenase.

    PubMed

    Kristan, Katja; Krajnc, Katja; Konc, Janez; Gobec, Stanislav; Stojan, Jure; Lanisnik Rizner, Tea

    2005-08-01

    Different phytoestrogens were tested as inhibitors of 17beta-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17beta-HSDcl), a member of the short-chain dehydrogenase/reductase superfamily. Phytoestrogens inhibited the oxidation of 100microM 17beta-hydroxyestra-4-en-3-one and the reduction of 100microM estra-4-en-3,17-dione, the best substrate pair known. The best inhibitors of oxidation, with IC(50) below 1microM, were flavones hydroxylated at positions 3, 5 and 7: 3-hydroxyflavone, 3,7-dihydroxyflavone, 5,7-dihydroxyflavone (chrysin) and 5-hydroxyflavone, together with 5-methoxyflavone. The best inhibitors of reduction were less potent; 3-hydroxyflavone, 5-methoxyflavone, coumestrol, 3,5,7,4'-tetrahydroxyflavone (kaempferol) and 5-hydroxyflavone, all had IC(50) values between 1 and 5microM. Docking the representative inhibitors chrysin and kaempferol into the active site of 17beta-HSDcl revealed the possible binding mode, in which they are sandwiched between the nicotinamide moiety and Tyr212. The structural features of phytoestrogens, inhibitors of both oxidation and reduction catalyzed by the fungal 17beta-HSD, are similar to the reported structural features of phytoestrogen inhibitors of human 17beta-HSD types 1 and 2.

  7. Inhibition of etoposide-induced DNA damage and cytotoxicity in L1210 cells by dehydrogenase inhibitors and other agents.

    PubMed

    Wozniak, A J; Glisson, B S; Hande, K R; Ross, W E

    1984-02-01

    The mechanism of action of 4'-demethylepipodophyllotoxin-9-(4,6-O-ethylidene-beta-D-glucopyra noside) (VP-16), an important antitumor agent, is unclear. There is evidence that DNA may be the target of action because VP-16 causes single-strand and double-strand breaks in DNA and produces cytotoxicity over a similar dose range. We have hypothesized that an enzyme system, such as dehydrogenase, catalyzes an oxidation-reduction reaction involving the pendant phenolic group which forms an active metabolite that causes the DNA damage and cytotoxicity. To test our hypothesis, we investigated the effect of disulfiram, an aldehyde dehydrogenase inhibitor, and its metabolite, diethyldithiocarbamate, on VP-16-induced DNA damage in L1210 cells. Using the alkaline elution technique to assay DNA damage, we found that disulfiram and diethyldithiocerbamate inhibited VP-16-induced single-strand breaks. Both compounds were also capable of significantly reducing VP-16-induced cytotoxicity. Oxalic acid, pyrophosphate, and malonic acid, competitive inhibitors of succinate dehydrogenase, and the naturally occurring dehydrogenase substrates, succinic acid, beta-glycerophosphate, and isocitric acid, also blocked the effects of VP-16. Free-radical scavengers were also studied. While sodium benzoate was particularly effective in preventing drug-induced DNA damage and cytotoxicity, a number of other scavengers were not. Our data are consistent with the hypothesis that VP-16 is activated by an enzyme such as a dehydrogenase which transforms it into an active intermediate resulting in DNA damage and, consequently, cell death.

  8. The diagnostic value of alcohol dehydrogenase (ADH) isoenzymes and aldehyde dehydrogenase (ALDH) measurement in the sera of patients with brain tumor

    PubMed Central

    Laniewska-Dunaj, Magdalena; Orywal, Karolina; Kochanowicz, Jan; Rutkowski, Robert; Szmitkowski, Maciej

    2017-01-01

    Introduction Alcohol dehydrogenase (ADH) isoenzymes and aldehyde dehydrogenase (ALDH) exist in the brain. Alcohol dehydrogenase and ALDH are also present in brain tumor cells. Moreover, the activity of class I isoenzymes was significantly higher in cancer than healthy brain cells. The activity of these enzymes in tumor tissue is reflected in the serum and could thus be helpful for diagnostics of brain neoplasms. The aim of this study was to investigate the potential role of ADH and ALDH as markers for brain tumors. Material and methods Serum samples were taken for routine biochemical investigation from 115 patients suffering from brain tumors (65 glioblastomas, 50 meningiomas). For the measurement of the activity of class I and II ADH isoenzymes and ALDH activity, fluorometric methods were used. The total ADH activity and activity of class III and IV isoenzymes were measured by the photometric method. Results There was a significant increase in the activity of ADH I isoenzyme and ADH total in the sera of brain tumor patients compared to the controls. The diagnostic sensitivity for ADH I was 78%, specificity 85%, and positive and negative predictive values were 86% and 76% respectively. The sensitivity and specificity of ADH I increased with the stage of the carcinoma. Area under receiver-operating characteristic curve for ADH I was 0.71. Conclusions The results suggest a potential role for ADH I as a marker for brain tumor. PMID:28261287

  9. East asian variant of aldehyde dehydrogenase 2 is associated with coronary spastic angina: possible roles of reactive aldehydes and implications of alcohol flushing syndrome.

    PubMed

    Mizuno, Yuji; Harada, Eisaku; Morita, Sumio; Kinoshita, Kenji; Hayashida, Mariko; Shono, Makoto; Morikawa, Yoshinobu; Murohara, Toyoaki; Nakayama, Masafumi; Yoshimura, Michihiro; Yasue, Hirofumi

    2015-05-12

    Coronary spastic angina (CSA) is a common disease among East Asians, including Japanese. The prevalence of alcohol flushing syndrome associated with deficient activity of the variant aldehyde dehydrogenase 2 (ALDH2*2) genotype is prevalent among East Asians. We examined whether CSA is associated with the ALDH2*2 genotype in Japanese. The study subjects consisted of 202 patients in whom intracoronary injection of acetylcholine was performed by angiography on suspicion of CSA (119 men and 83 women; mean age, 66.2±11.4 years). They were divided into CSA (112 patients) and control groups (90 patients). ALDH2 genotyping was performed by the direct application of the TaqMan polymerase chain reaction system on dried whole blood. Clinical and laboratory data were examined using conventional methods. The frequencies of male sex, ALDH2*2 genotype carriers, alcohol flushing syndrome, tobacco smoking, and the plasma level of uric acid were higher (P<0.001, P<0.001, P<0.001, P<0.001, and P=0.007, respectively) and the plasma high-density lipoprotein cholesterol levels were lower (P<0.001) in the CSA group than in the control group. The multivariable logistic regression analysis revealed that ALDH2*2 genotype and smoking were significantly associated with CSA (P<0.001 and P=0.024, respectively). East Asian variant ALDH2*2 genotypes and, hence, deficient ALDH2 activity were associated with CSA in Japanese. These data support further investigation of treatment targeting aldehydes for CSA. © 2015 American Heart Association, Inc.

  10. Comparative study of the aldehyde dehydrogenase (ALDH) gene superfamily in the glycophyte Arabidopsis thaliana and Eutrema halophytes

    PubMed Central

    Hou, Quancan; Bartels, Dorothea

    2015-01-01

    Background and Aims Stresses such as drought or salinity induce the generation of reactive oxygen species, which subsequently cause excessive accumulation of aldehydes in plant cells. Aldehyde dehydrogenases (ALDHs) are considered as ‘aldehyde scavengers’ to eliminate toxic aldehydes caused by oxidative stress. The completion of the genome sequencing projects of the halophytes Eutrema parvulum and E. salsugineum has paved the way to explore the relationships and the roles of ALDH genes in the glycophyte Arabidopsis thaliana and halophyte model plants. Methods Protein sequences of all plant ALDH families were used as queries to search E. parvulum and E. salsugineum genome databases. Evolutionary analyses compared the phylogenetic relationships of ALDHs from A. thaliana and Eutrema. Expression patterns of several stress-associated ALDH genes were investigated under different salt conditions using reverse transcription–PCR. Putative cis-elements in the promoters of ALDH10A8 from A. thaliana and E. salsugineum were compared in silico. Key Results Sixteen and 17 members of ten ALDH families were identified from E. parvulum and E. salsugineum genomes, respectively. Phylogenetic analysis of ALDH protein sequences indicated that Eutrema ALDHs are closely related to those of Arabidopsis, and members within these species possess nearly identical exon–intron structures. Gene expression analysis under different salt conditions showed that most of the ALDH genes have similar expression profiles in Arabidopsis and E. salsugineum, except for ALDH7B4 and ALDH10A8. In silico analysis of promoter regions of ALDH10A8 revealed different distributions of cis-elements in E. salsugineum and Arabidopsis. Conclusions Genomic organization, copy number, sub-cellular localization and expression profiles of ALDH genes are conserved in Arabidopsis, E. parvulum and E. salsugineum. The different expression patterns of ALDH7B4 and ALDH10A8 in Arabidopsis and E. salsugineum suggest that E

  11. Elucidating the contributions of multiple aldehyde/alcohol dehydrogenases to butanol and ethanol production in Clostridium acetobutylicum

    PubMed Central

    Dai, Zongjie; Dong, Hongjun; Zhang, Yanping; Li, Yin

    2016-01-01

    Ethanol and butanol biosynthesis in Clostridium acetobutylicum share common aldehyde/alcohol dehydrogenases. However, little is known about the relative contributions of these multiple dehydrogenases to ethanol and butanol production respectively. The contributions of six aldehyde/alcohol dehydrogenases of C. acetobutylicum on butanol and ethanol production were evaluated through inactivation of the corresponding genes respectively. For butanol production, the relative contributions from these enzymes were: AdhE1 > BdhB > BdhA ≈ YqhD > SMB_P058 > AdhE2. For ethanol production, the contributions were: AdhE1 > BdhB > YqhD > SMB_P058 > AdhE2 > BdhA. AdhE1 and BdhB are two essential enzymes for butanol and ethanol production. AdhE1 was relatively specific for butanol production over ethanol, while BdhB, YqhD, and SMB_P058 favor ethanol production over butanol. Butanol synthesis was increased in the adhE2 mutant, which had a higher butanol/ethanol ratio (8.15:1) compared with wild type strain (6.65:1). Both the SMB_P058 mutant and yqhD mutant produced less ethanol without loss of butanol formation, which led to higher butanol/ethanol ratio, 10.12:1 and 10.17:1, respectively. To engineer a more efficient butanol-producing strain, adhE1 could be overexpressed, furthermore, adhE2, SMB_P058, yqhD are promising gene inactivation targets. This work provides useful information guiding future strain improvement for butanol production. PMID:27321949

  12. Ubiquitin-aldehyde: a general inhibitor of ubiquitin-recycling processes.

    PubMed Central

    Hershko, A; Rose, I A

    1987-01-01

    The generation and characterization of ubiquitin (Ub)-aldehyde, a potent inhibitor of Ub-C-terminal hydrolase, has previously been reported. We now examine the action of this compound on the Ub-mediated proteolytic pathway using the system derived from rabbit reticulocytes. Addition of Ub-aldehyde was found to strongly inhibit breakdown of added 125I-labeled lysozyme, but inhibition was overcome by increasing concentrations of Ub. The following evidence shows the effect of Ub-aldehyde on protein breakdown to be indirectly caused by its interference with the recycling of Ub, leading to exhaustion of the supply of free Ub: Ub-aldehyde markedly increased the accumulation of Ub-protein conjugates coincident with a much decreased rate of conjugate breakdown. release of Ub from isolated Ub-protein conjugates in the absence of ATP (and therefore not coupled to protein degradation) is markedly inhibited by Ub-aldehyde. On the other hand, the ATP-dependent degradation of the protein moiety of Ub conjugates, which is an integral part of the proteolytic process, is not inhibited by this agent. Direct measurement of levels of free Ub showed a rapid disappearance caused by the inhibitor. The Ub is found to be distributed in derivatives of a wide range of molecular weight classes. It thus seems that Ub-aldehyde, previously demonstrated to inhibit the hydrolysis of Ub conjugates of small molecules, also inhibits the activity of a series of enzymes that regenerate free Ub from adducts with proteins and intermediates in protein breakdown. Images PMID:3031653

  13. Ubiquitin-aldehyde: a general inhibitor of ubiquitin-recycling processes

    SciTech Connect

    Hershko, A.; Rose, I.A.

    1987-04-01

    The generation and characterization of ubiquitin (Ub)-aldehyde, a potent inhibitor of Ub-C-terminal hydrolase, has previously been reported. The authors examine the action of this compound on the Ub-mediated proteolytic pathway using the system derived from rabbit reticulocytes. Addition of Ub-aldehyde was found to strongly inhibit breakdown of added /sup 125/I-labeled lysozyme, but inhibition was overcome by increasing concentrations of Ub. The following evidence shows the effect of Ub-aldehyde on protein breakdown to be indirectly caused by its interference with the recycling of Ub, leading to exhaustion of the supply of free Ub: (i) Ub-aldehyde markedly increased the accumulation of Ub-protein conjugates coincident with a much decreased rate of conjugate breakdown; (ii) release of Ub from isolated Ub-protein conjugates in the absence of ATP (and therefore not coupled to protein degradation) is markedly inhibited by Ub-aldehyde. On the other hand, the ATP-dependent degradation of the protein moiety of Ub conjugates, which is an integral part of the proteolytic process, is not inhibited by this agent; (iii) direct measurement of levels of free Ub showed a rapid disappearance caused by the inhibitor. The Ub is found to be distributed in derivatives of a wide range of molecular weight classes. It thus seems that Ub-aldehyde, previously demonstrated to inhibit the hydrolysis of Ub conjugates of small molecules, also inhibits the activity of a series of enzymes that regenerate free Ub from adducts with proteins and intermediates in protein breakdown.

  14. Comparative genomics of aldehyde dehydrogenase 5a1 (succinate semialdehyde dehydrogenase) and accumulation of gamma-hydroxybutyrate associated with its deficiency

    PubMed Central

    2009-01-01

    Succinic semialdehyde dehydrogenase (SSADH; aldehyde dehydrogenase 5A1 [ALDH5A1]; locus 6p22) occupies a central position in central nervous system (CNS) neurotransmitter metabolism as one of two enzymes necessary for γ-aminobutyric acid (GABA) recycling from the synaptic cleft. Its importance is highlighted by the neurometabolic disease associated with its inherited deficiency in humans, as well as the severe epileptic phenotype observed in Aldh5a1-/- knockout mice. Expanding evidence now suggests, however, that even subtle decreases in human SSADH activity, associated with rare and common single nucleotide polymorphisms, may produce subclinical pathological effects. SSADH, in conjunction with aldo-keto reductase 7A2 (AKR7A2), represent two neural enzymes responsible for further catabolism of succinic semialdehyde, producing either succinate (SSADH) or γ-hydroxybutyrate (GHB; AKR7A2). A GABA analogue, GHB is a short-chain fatty alcohol with unusual properties in the CNS and a long pharmacological history. Moreover, SSADH occupies a further role in the CNS as the enzyme responsible for further metabolism of the lipid peroxidation aldehyde 4-hydroxy-2-nonenal (4-HNE), an intermediate known to induce oxidant stress. Accordingly, subtle decreases in SSADH activity may have the capacity to lead to regional accumulation of neurotoxic intermediates (GHB, 4-HNE). Polymorphisms in SSADH gene structure may also associate with quantitative traits, including intelligence quotient and life expectancy. Further population-based studies of human SSADH activity promise to reveal additional properties of its function and additional roles in CNS tissue. PMID:19164088

  15. Human aldehyde dehydrogenase 3A1 (ALDH3A1): biochemical characterization and immunohistochemical localization in the cornea.

    PubMed Central

    Pappa, Aglaia; Estey, Tia; Manzer, Rizwan; Brown, Donald; Vasiliou, Vasilis

    2003-01-01

    ALDH3A1 (aldehyde dehydrogenase 3A1) is expressed at high concentrations in the mammalian cornea and it is believed that it protects this vital tissue and the rest of the eye against UV-light-induced damage. The precise biological function(s) and cellular distribution of ALDH3A1 in the corneal tissue remain to be elucidated. Among the hypotheses proposed for ALDH3A1 function in cornea is detoxification of aldehydes formed during UV-induced lipid peroxidation. To investigate in detail the biochemical properties and distribution of this protein in the human cornea, we expressed human ALDH3A1 in Sf9 insect cells using a baculovirus vector and raised monoclonal antibodies against ALDH3A1. Recombinant ALDH3A1 protein was purified to homogeneity with a single-step affinity chromatography method using 5'-AMP-Sepharose 4B. Human ALDH3A1 demonstrated high substrate specificity for medium-chain (6 carbons and more) saturated and unsaturated aldehydes, including 4-hydroxy-2-nonenal, which are generated by the peroxidation of cellular lipids. Short-chain aliphatic aldehydes, such as acetaldehyde, propionaldehyde and malondialdehyde, were found to be very poor substrates for human ALDH3A1. In addition, ALDH3A1 metabolized glyceraldehyde poorly and did not metabolize glucose 6-phosphate, 6-phosphoglucono-delta-lactone and 6-phosphogluconate at all, suggesting that this enzyme is not involved in either glycolysis or the pentose phosphate pathway. Immunohistochemistry in human corneas, using the monoclonal antibodies described herein, revealed ALDH3A1 expression in epithelial cells and stromal keratocytes, but not in endothelial cells. Overall, these cumulative findings support the metabolic function of ALDH3A1 as a part of a corneal cellular defence mechanism against oxidative damage caused by aldehydic products of lipid peroxidation. Both recombinant human ALDH3A1 and the highly specific monoclonal antibodies described in the present paper may prove to be useful in probing

  16. The activity of class I, II, III and IV alcohol dehydrogenase isoenzymes and aldehyde dehydrogenase in ovarian cancer and ovarian cysts.

    PubMed

    Orywal, K; Jelski, W; Zdrodowski, M; Szmitkowski, M

    2013-01-01

    The metabolism of cancerous cells is in many ways different than in healthy cells. In ovarian cancer, cells exhibit activity of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), which participate in metabolism of many biological substances. The aim of this study was to compare the metabolism of ovarian cancer cells, ovarian cysts and normal ovarian cells by measurement of ADH isoenzymes and ALDH activities. The study material consisted of 36 cancerous ovarian tissues. Class III, IV of ADH and total ADH activity was measured by the photometric method and class I, II ADH and ALDH activity by the fluorometric method with class-specific fluorogenic substrates. The activity of the class I ADH isoenzyme and the total ADH was significantly higher in ovarian cancer as compared to ovarian cysts and healthy tissues but there are no significant differences between ovarian cysts and healthy cells. The other classes of ADH tested, did not show significant differences between activity of cancerous cells and healthy ovary. The increased activity of total ADH in ovarian cancer, especially the class I isoenzyme and normal activity of ALDH, may be the factor for the disturbances in important biological substances metabolism and could increase the concentration of highly carcinogenic acetaldehyde.

  17. HEPATOCYTE EXPRESION OF TUMOR ASSOCIATED ALDEHYDE DEHYDROGENASE (ALDH-3) AND P21 RAS FOLLOWING DIETHYLNITROSAMINE (DEN) INITIATION AND CHRONIC EXPOSURE TO DI(2-ETHYLHEXYL) PHTHALATE (DHEP)

    EPA Science Inventory

    Phthalate esters such as di(2-ethylhexyl)phthalate (DEHP)either promote or inhibit rat liver tumorigenesis depending on the carcinogenesis protocol. In this study, we examined the expression of two histochemical markers, the tumor associated isozyme of aldehyde dehydrogenase (ALD...

  18. Aldehyde dehydrogenase activity identifies a subpopulation of canine adipose-derived stem cells with higher differentiation potential.

    PubMed

    Itoh, Harumichi; Nishikawa, Shimpei; Haraguchi, Tomoya; Arikawa, Yu; Hiyama, Masato; Eto, Shotaro; Iseri, Toshie; Itoh, Yoshiki; Tani, Kenji; Nakaichi, Munekazu; Taura, Yasuho; Itamoto, Kazuhito

    2017-09-12

    Adipose-derived stem cells (ADSCs) are abundant and readily obtained, and have been studied for their clinical applicability in regenerative medicine. Some surface antigens have been identified as markers of different ADSC subpopulations in mice and humans. However, it is unclear whether functionally distinct subpopulations exist in dogs. To address this issue, we evaluated aldehyde dehydrogenase (ALDH) activity-a widely used stem cell marker in mice and humans-by flow cytometry. Approximately 20% of bulk ADSCs showed high ALDH activity. Compared to cells with low activity (ALDH(Lo)), the high-activity (ALDH(Hi)) subpopulation exhibited a higher capacity for adipogenic and osteogenic differentiation. This is the first report of distinct ADSC subpopulations in dogs that differ in terms of adipogenic and osteogenic differentiation potential.

  19. The mechanism of discrimination between oxidized and reduced coenzyme in the aldehyde dehydrogenase domain of Aldh1l1.

    PubMed

    Tsybovsky, Yaroslav; Malakhau, Yuryi; Strickland, Kyle C; Krupenko, Sergey A

    2013-02-25

    Aldh1l1, also known as 10-formyltetrahydrofolate dehydrogenase (FDH), contains the carboxy-terminal domain (Ct-FDH), which is a structural and functional homolog of aldehyde dehydrogenases (ALDHs). This domain is capable of catalyzing the NADP(+)-dependent oxidation of short chain aldehydes to their corresponding acids, and similar to most ALDHs it has two conserved catalytic residues, Cys707 and Glu673. Previously, we demonstrated that in the Ct-FDH mechanism these residues define the conformation of the bound coenzyme and the affinity of its interaction with the protein. Specifically, the replacement of Cys707 with an alanine resulted in the enzyme lacking the ability to differentiate between the oxidized and reduced coenzyme. We suggested that this was due to the loss of a covalent bond between the cysteine and the C4N atom of nicotinamide ring of NADP(+) formed during Ct-FDH catalysis. To obtain further insight into the functional significance of the covalent bond between Cys707 and the coenzyme, and the overall role of the two catalytic residues in the coenzyme binding and positioning, we have now solved crystal structures of Ct-FDH in the complex with thio-NADP(+) and the complexes of the C707S mutant with NADP(+) and NADPH. This study has allowed us to trap the coenzyme in the contracted conformation, which provided a snapshot of the conformational processing of the coenzyme during the transition from oxidized to reduced form. Overall, the results of this study further support the previously proposed mechanism by which Cys707 helps to differentiate between the oxidized and reduced coenzyme during ALDH catalysis.

  20. The mechanism of discrimination between oxidized and reduced coenzyme in the aldehyde dehydrogenase domain of Aldh1l1

    PubMed Central

    Tsybovsky, Yaroslav; Malakhau, Yuryi; Strickland, Kyle C.; Krupenko, Sergey A.

    2013-01-01

    Aldh1l1, also known as 10-formyltetrahydrofolate dehydrogenase (FDH), contains the carboxy-terminal domain (Ct-FDH), which is a structural and functional homolog of aldehyde dehydrogenases (ALDHs). This domain is capable of catalyzing the NADP+-dependent oxidation of short chain aldehydes to their corresponding acids, and similar to most ALDHs it has two conserved catalytic residues, Cys707 and Glu673. Previously, we demonstrated that in the Ct-FDH mechanism these residues define the conformation of the bound coenzyme and the affinity of its interaction with the protein. Specifically, the replacement of Cys707 with an alanine resulted in the enzyme lacking the ability to differentiate between the oxidized and reduced coenzyme. We suggested that this was due to the loss of a covalent bond between the cysteine and the C4N atom of nicotinamide ring of NADP+ formed during Ct-FDH catalysis. To obtain further insight into the functional significance of the covalent bond between Cys707 and the coenzyme, and the overall role of the two catalytic residues in the coenzyme binding and positioning, we have now solved crystal structures of Ct-FDH in the complex with thio-NADP+ and the complexes of the C707S mutant with NADP+ and NADPH. This study has allowed us to trap the coenzyme in the contracted conformation, which provided a snapshot of the conformational processing of the coenzyme during the transition from oxidized to reduced form. Overall, the results of this study further support the previously proposed mechanism by which Cys707 helps to differentiate between the oxidized and reduced coenzyme during ALDH catalysis. PMID:23295222

  1. Preventive effects of Chlorella on skeletal muscle atrophy in muscle-specific mitochondrial aldehyde dehydrogenase 2 activity-deficient mice.

    PubMed

    Nakashima, Yuya; Ohsawa, Ikuroh; Nishimaki, Kiyomi; Kumamoto, Shoichiro; Maruyama, Isao; Suzuki, Yoshihiko; Ohta, Shigeo

    2014-10-11

    Oxidative stress is involved in age-related muscle atrophy, such as sarcopenia. Since Chlorella, a unicellular green alga, contains various antioxidant substances, we used a mouse model of enhanced oxidative stress to investigate whether Chlorella could prevent muscle atrophy. Aldehyde dehydrogenase 2 (ALDH2) is an anti-oxidative enzyme that detoxifies reactive aldehydes derived from lipid peroxides such as 4-hydroxy-2-nonenal (4-HNE). We therefore used transgenic mice expressing a dominant-negative form of ALDH2 (ALDH2*2 Tg mice) to selectively decrease ALDH2 activity in the muscles. To evaluate the effect of Chlorella, the mice were fed a Chlorella-supplemented diet (CSD) for 6 months. ALDH2*2 Tg mice exhibited small body size, muscle atrophy, decreased fat content, osteopenia, and kyphosis, accompanied by increased muscular 4-HNE levels. The CSD helped in recovery of body weight, enhanced oxidative stress, and increased levels of a muscle impairment marker, creatine phosphokinase (CPK) induced by ALDH2*2. Furthermore, histological and histochemical analyses revealed that the consumption of the CSD improved skeletal muscle atrophy and the activity of the mitochondrial cytochrome c oxidase. This study suggests that long-term consumption of Chlorella has the potential to prevent age-related muscle atrophy.

  2. Targeted therapy for a subset of acute myeloid leukemias that lack expression of aldehyde dehydrogenase 1A1.

    PubMed

    Gasparetto, Maura; Pei, Shanshan; Minhajuddin, Mohammad; Khan, Nabilah; Pollyea, Daniel A; Myers, Jason R; Ashton, John M; Becker, Michael W; Vasiliou, Vasilis; Humphries, Keith R; Jordan, Craig T; Smith, Clayton A

    2017-03-09

    Aldehyde dehydrogenase 1A1 (ALDH1A1) activity is high in hematopoietic stem cells and functions in part to protect stem cells from reactive aldehydes and other toxic compounds. In contrast, we found that ~25% of all acute myeloid leukemias expressed low or undetectable levels of ALDH1A1 and that this ALDH1A1- subset of leukemias correlates with good prognosis cytogenetics. ALDH1A1- cell lines as well as primary leukemia cells were found to be sensitive to treatment with compounds that directly and indirectly generate toxic ALDH substrates including 4-hydroxynonenal and the clinically relevant compounds arsenic trioxide and 4-hydroperoxycyclophosphamide. In contrast, normal hematopoietic stem cells were relatively resistant to these compounds. Using a murine xenotransplant model to emulate a clinical treatment strategy, established ALDH1A1- leukemias were also sensitive to in vivo treatment with cyclophosphamide combined with arsenic trioxide. These results demonstrate that targeting ALDH1A1- leukemic cells with toxic ALDH1A1 substrates such as arsenic and cyclophosphamide may be a novel targeted therapeutic strategy for this subset of acute myeloid leukemias.

  3. Expression of a betaine aldehyde dehydrogenase gene in rice, a glycinebetaine nonaccumulator, and possible localization of its protein in peroxisomes.

    PubMed

    Nakamura, T; Yokota, S; Muramoto, Y; Tsutsui, K; Oguri, Y; Fukui, K; Takabe, T

    1997-05-01

    Betaine aldehyde dehydrogenase (BADH) catalyzes the last step in the plant biosynthetic pathway that leads to glycinebetaine. Rice plants (Oryza sativa L.), albeit considered a typical non-glycinebetaine accumulating species, have been found to express this enzyme at low levels. This observation evokes an interest in phylogenic evolution of the enzyme in the plant kingdom. It is reported here that rice plants possess the ability to take up exogenously added betaine aldehyde through the roots and convert it to glycinebetaine, resulting in an enhanced salt-tolerance of the plants. A gene encoding a putative BADH from the rice genome was also cloned and sequenced. The gene was found to contain 14 introns, and the overall nucleotide sequence of the coding region is c. 78% identical to that of the barley BADH cDNA. Cloning of a partial BADH cDNA from rice was accomplished by reverse transcription-polymerase chain reaction (RT-PCR). The nucleotide sequence of the cloned fragment was found to be identical to the corresponding exon regions of the rice genomic BADH gene. The deduced amino acid sequences of rice and barley BADH both contain a C-terminal tripeptide SKL, a signal known to target preproteins to microbodies. This localization was confirmed by an immuno-gold labeling study of transgenic tobacco harboring barley cDNA, which showed BADH protein inside peroxisomes. Northern blot analysis revealed that the level of BADH mRNA is salt-inducible.

  4. ABA3 is a molybdenum cofactor sulfurase required for activation of aldehyde oxidase and xanthine dehydrogenase in Arabidopsis thaliana.

    PubMed

    Bittner, F; Oreb, M; Mendel, R R

    2001-11-02

    The xanthine oxidase class of molybdenum enzyzmes requires a terminal sulfur ligand at the active site. It has been proposed that a special sulfurase catalyzes the insertion of this ligand thereby activating the enzymes. Previous analyses of mutants in plants indicated that the genetic locus aba3 is involved in this step leading to activation of the molybdenum enzymes aldehyde oxidase and xanthine dehydrogenase. Here we report the cloning of the aba3 gene from Arabidopsis thaliana and the biochemical characterization of the purified protein. ABA3 is a two-domain protein with a N-terminal NifS-like sulfurase domain and a C-terminal domain that might be involved in recognizing the target enzymes. Molecular analysis of three aba3 mutants identified mutations in both domains. ABA3 contains highly conserved binding motifs for pyridoxal phosphate and for a persulfide. The purified recombinant protein possesses a cysteine desulfurase activity, is yellow in color, and shows a NifS-like change in absorbance in the presence of L-cysteine. Pretreatment of ABA3 with a thiol-specific alkylating reagent inhibited its desulfurase activity. These data indicate a transsulfuration reaction similar to bacterial NifS. In a fully defined in vitro system, the purified protein was able to activate aldehyde oxidase by using L-cysteine as sulfur donor. Finally, we show that the expression of the aba3 gene is inducible by drought-stress.

  5. Aldehyde dehydrogenase 3A1 protects airway epithelial cells from cigarette smoke-induced DNA damage and cytotoxicity.

    PubMed

    Jang, Jun-Ho; Bruse, Shannon; Liu, Yushi; Duffy, Veronica; Zhang, Chunyu; Oyamada, Nathaniel; Randell, Scott; Matsumoto, Akiko; Thompson, David C; Lin, Yong; Vasiliou, Vasilis; Tesfaigzi, Yohannes; Nyunoya, Toru

    2014-03-01

    Aldehyde dehydrogenase 3A1 (ALDH3A1), an ALDH superfamily member, catalyzes the oxidation of reactive aldehydes, highly toxic components of cigarette smoke (CS). Even so, the role of ALDH3A1 in CS-induced cytotoxicity and DNA damage has not been examined. Among all of the ALDH superfamily members, ALDH3A1 mRNA levels showed the greatest induction in response to CS extract (CSE) exposure of primary human bronchial epithelial cells (HBECs). ALDH3A1 protein accumulation was accompanied by increased ALDH enzymatic activity in CSE-exposed immortalized HBECs. The effects of overexpression or suppression of ALDH3A1 on CSE-induced cytotoxicity and DNA damage (γH2AX) were evaluated in cultured immortalized HBECs. Enforced expression of ALDH3A1 attenuated cytotoxicity and downregulated γH2AX. SiRNA-mediated suppression of ALDH3A1 blocked ALDH enzymatic activity and augmented cytotoxicity in CSE-exposed cells. Our results suggest that the availability of ALDH3A1 is important for cell survival against CSE in HBECs.

  6. Structure and mechanism of action of the hydroxy aryl aldehyde class of IRE1 endoribonuclease inhibitors

    PubMed Central

    Sanches, Mario; Duffy, Nicole M.; Talukdar, Manisha; Thevakumaran, Nero; Chiovitti, David; Canny, Marella D.; Lee, Kenneth; Kurinov, Igor; Uehling, David; Al-awar, Rima; Poda, Gennadiy; Prakesch, Michael; Wilson, Brian; Tam, Victor; Schweitzer, Colleen; Toro, Andras; Lucas, Julie L.; Vuga, Danka; Lehmann, Lynn; Durocher, Daniel; Zeng, Qingping; Patterson, John B.; Sicheri, Frank

    2014-01-01

    Endoplasmic reticulum (ER) stress activates the unfolded protein response and its dysfunction is linked to multiple diseases. The stress transducer IRE1α is a transmembrane kinase endoribonuclease (RNase) that cleaves mRNA substrates to re-establish ER homeostasis. Aromatic ring systems containing hydroxy-aldehyde moieties, termed hydroxy aryl aldehydes (HAA), selectively inhibit IRE1α RNase and thus represent a novel chemical series for therapeutic development. We solved crystal structures of murine IRE1α in complex with three HAA inhibitors. HAA inhibitors engage a shallow pocket at the RNase active site through pi-stacking interactions with His910 and Phe889, an essential Schiff base with Lys907 and a H-bond with Tyr892. Structure activity studies and mutational analysis of contact residues define the optimal chemical space of inhibitors and validate the inhibitor binding site. These studies lay the foundation for understanding both the biochemical and cellular functions of IRE1α using small molecule inhibitors and suggest new avenues for inhibitor design. PMID:25164867

  7. Triazaspirodimethoxybenzoyls as Selective Inhibitors of Mycobacterial Lipoamide Dehydrogenase

    SciTech Connect

    Bryk, Ruslana; Arango, Nancy; Venugopal, Aditya; Warren, J. David; Park, Yun-Hee; Patel, Mulchand S.; Lima, Christopher D.; Nathan, Carl

    2010-06-25

    Mycobacterium tuberculosis (Mtb) remains the leading single cause of death from bacterial infection. Here we explored the possibility of species-selective inhibition of lipoamide dehydrogenase (Lpd), an enzyme central to Mtb's intermediary metabolism and antioxidant defense. High-throughput screening of combinatorial chemical libraries identified triazaspirodimethoxybenzoyls as high-nanomolar inhibitors of Mtb's Lpd that were noncompetitive versus NADH, NAD{sup +}, and lipoamide and >100-fold selective compared to human Lpd. Efficacy required the dimethoxy and dichlorophenyl groups. The structure of an Lpd-inhibitor complex was resolved to 2.42 {angstrom} by X-ray crystallography, revealing that the inhibitor occupied a pocket adjacent to the Lpd NADH/NAD{sup +} binding site. The inhibitor did not overlap with the adenosine moiety of NADH/NAD{sup +} but did overlap with positions predicted to bind the nicotinamide rings in NADH and NAD{sup +} complexes. The dimethoxy ring occupied a deep pocket adjacent to the FAD flavin ring where it would block coordination of the NADH nicotinamide ring, while the dichlorophenyl group occupied a more exposed pocket predicted to coordinate the NAD{sup +} nicotinamide. Several residues that are not conserved between the bacterial enzyme and its human homologue were predicted to contribute both to inhibitor binding and to species selectivity, as confirmed for three residues by analysis of the corresponding mutant Mtb Lpd proteins. Thus, nonconservation of residues lining the electron-transfer tunnel in Mtb Lpd can be exploited for development of species-selective Lpd inhibitors.

  8. Triazaspirodimethoxybenzoyls as selective inhibitors of mycobacterial lipoamide dehydrogenase .

    PubMed

    Bryk, Ruslana; Arango, Nancy; Venugopal, Aditya; Warren, J David; Park, Yun-Hee; Patel, Mulchand S; Lima, Christopher D; Nathan, Carl

    2010-03-02

    Mycobacterium tuberculosis (Mtb) remains the leading single cause of death from bacterial infection. Here we explored the possibility of species-selective inhibition of lipoamide dehydrogenase (Lpd), an enzyme central to Mtb's intermediary metabolism and antioxidant defense. High-throughput screening of combinatorial chemical libraries identified triazaspirodimethoxybenzoyls as high-nanomolar inhibitors of Mtb's Lpd that were noncompetitive versus NADH, NAD(+), and lipoamide and >100-fold selective compared to human Lpd. Efficacy required the dimethoxy and dichlorophenyl groups. The structure of an Lpd-inhibitor complex was resolved to 2.42 A by X-ray crystallography, revealing that the inhibitor occupied a pocket adjacent to the Lpd NADH/NAD(+) binding site. The inhibitor did not overlap with the adenosine moiety of NADH/NAD(+) but did overlap with positions predicted to bind the nicotinamide rings in NADH and NAD(+) complexes. The dimethoxy ring occupied a deep pocket adjacent to the FAD flavin ring where it would block coordination of the NADH nicotinamide ring, while the dichlorophenyl group occupied a more exposed pocket predicted to coordinate the NAD(+) nicotinamide. Several residues that are not conserved between the bacterial enzyme and its human homologue were predicted to contribute both to inhibitor binding and to species selectivity, as confirmed for three residues by analysis of the corresponding mutant Mtb Lpd proteins. Thus, nonconservation of residues lining the electron-transfer tunnel in Mtb Lpd can be exploited for development of species-selective Lpd inhibitors.

  9. Isolated tumoral pyruvate dehydrogenase can synthesize acetoin which inhibits pyruvate oxidation as well as other aldehydes.

    PubMed

    Baggetto, L G; Lehninger, A L

    1987-05-29

    Oxidation of 1 mM pyruvate by Ehrlich and AS30-D tumor mitochondria is inhibited by acetoin, an unusual and important metabolite of pyruvate utilization by cancer cells, by acetaldehyde, methylglyoxal and excess pyruvate. The respiratory inhibition is reversed by other substrates added to pyruvate and also by 0.5 mM ATP. Kinetic properties of pyruvate dehydrogenase complex isolated from these tumor mitochondria have been studied. This complex appears to be able to synthesize acetoin from acetaldehyde plus pyruvate and is competitively inhibited by acetoin. The role of a new regulatory pattern for tumoral pyruvate dehydrogenase is presented.

  10. Highly efficient asymmetric reduction of arylpropionic aldehydes by horse liver alcohol dehydrogenase through dynamic kinetic resolution.

    PubMed

    Giacomini, Daria; Galletti, Paola; Quintavalla, Arianna; Gucciardo, Gabriele; Paradisi, Francesca

    2007-10-21

    The enantioselective synthesis of (2S)-2-phenylpropanol and (2S)-2-(4-iso-butylphenyl)propanol ((S)-Ibuprofenol) has been achieved by means of Horse Liver Alcohol Dehydrogenase (HLADH) in buffered aqueous solution or buffered organic solvent mixtures; under the reaction conditions, a dynamic kinetic resolution (DKR) process was realized with good reaction yields and enantiomeric ratios.

  11. Expression of a heat-stable NADPH-dependent alcohol dehydrogenase in Caldicellulosiruptor bescii results in furan aldehyde detoxification

    DOE PAGES

    Chung, Daehwan; Verbeke, Tobin J.; Cross, Karissa L.; ...

    2015-07-22

    Compounds such as furfural and 5-hydroxymethylfurfural (5-HMF) are generated through the dehydration of xylose and glucose, respectively, during dilute-acid pretreatment of lignocellulosic biomass and are also potent microbial growth and fermentation inhibitors. The enzymatic reduction of these furan aldehydes to their corresponding, and less toxic, alcohols is an engineering approach that has been successfully implemented in both Saccharomyces cerevisiae and ethanologenicEscherichia coli, but has not yet been investigated in thermophiles relevant to biofuel production through consolidated bioprocessing (CBP). Developing CBP-relevant biocatalysts that are either naturally resistant to such inhibitors, or are amenable to engineered resistance, is therefore, an important componentmore » in making biofuels production from lignocellulosic biomass feasible.« less

  12. Expression of a heat-stable NADPH-dependent alcohol dehydrogenase in Caldicellulosiruptor bescii results in furan aldehyde detoxification

    SciTech Connect

    Chung, Daehwan; Verbeke, Tobin J.; Cross, Karissa L.; Westpheling, Janet; Elkins, James G.

    2015-07-22

    Compounds such as furfural and 5-hydroxymethylfurfural (5-HMF) are generated through the dehydration of xylose and glucose, respectively, during dilute-acid pretreatment of lignocellulosic biomass and are also potent microbial growth and fermentation inhibitors. The enzymatic reduction of these furan aldehydes to their corresponding, and less toxic, alcohols is an engineering approach that has been successfully implemented in both Saccharomyces cerevisiae and ethanologenicEscherichia coli, but has not yet been investigated in thermophiles relevant to biofuel production through consolidated bioprocessing (CBP). Developing CBP-relevant biocatalysts that are either naturally resistant to such inhibitors, or are amenable to engineered resistance, is therefore, an important component in making biofuels production from lignocellulosic biomass feasible.

  13. Structure of Plasmodium falciparum dihydroorotate dehydrogenase with a bound inhibitor.

    PubMed

    Hurt, Darrell E; Widom, Joanne; Clardy, Jon

    2006-03-01

    Membrane-associated dihydroorotate dehydrogenase (DHODH) is an antimalarial therapeutic target without an effective inhibitor. Studies on human DHODH (HsDHODH) led to a structural mechanistic model in which respiratory quinones bind in a tunnel formed by the highly variable N-terminus that leads to the flavin mononucleotide-binding site. The therapeutic agents leflunomide (Arava) and brequinar sodium inhibit HsDHODH by binding in this tunnel. Plasmodium falciparum DHODH (PfDHODH) and HsDHODH have markedly different sensitivities to the two drugs. To understand the structural basis of this differential sensitivity and begin a structure-based drug-design cycle for PfDHODH inhibitors, the three-dimensional structure (2.4 Angstroms, R = 20.1%) of PfDHODH bound to the active metabolite of leflunomide was determined by X-ray crystallography. Comparison of the structures of HsDHODH and PfDHODH reveals a completely different binding mode for the same inhibitor in these two catalytically identical enzymes and explains the previously observed species-specific preferential binding. Because no effective inhibitors have been described for PfDHODH, this structure provides critical insight for the design of potential antimalarials.

  14. Relationships of alcohol dehydrogenase 1B (ADH1B) and aldehyde dehydrogenase 2 (ALDH2) genotypes with alcohol sensitivity, drinking behavior and problem drinking in Japanese older men.

    PubMed

    Hashimoto, Marowa; Watanabe, Masutaka; Uematsu, Yuji; Hattori, Sonomi; Miyai, Nobuyuki; Utsumi, Miyoko; Oka, Mayumi; Hayashida, Mariko; Kinoshita, Kenji; Arita, Mikio; Takeshita, Tatsuya

    2016-05-01

    Many East Asians have the genetic polymorphisms rs1229984 in alcohol dehydrogenase 1B (ADH1B) and rs671 in aldehyde dehydrogenase 2 (ALDH2). Here we analyzed the relationships of the two genotypes with alcohol sensitivity, drinking behavior and problem drinking among older and younger men living in rural areas of Japan. The subjects were 718 Japanese men aged 63.3 ± 10.8 (mean ± SD), categorized into the older (≥65 years, n = 357) and younger (<65 years, n = 361) groups. Facial flushing frequency, drinking behavior and positive CAGE results were compared among the genotypes using Bonferroni-corrected χ(2) test and a multivariate logistic regression analysis adjusting for age, BMI and lifestyle factors. The frequency of 'always' facial flushing among the ADH1B*1/*2 carriers was significantly lower than that among the ADH1B*2/*2 carriers in the older group (P < 0.01). The alcohol consumption (unit/day) in the ADH1B*1/*2 carriers tended to be higher compared with that in the ADH1B*2/*2 carriers among the older group (P = 0.050). In the younger group, no significant differences in alcohol sensitivity and drinking habits were generally found among the ADH1B genotypes. The ADH1B*1/*1 genotype tended to be positively associated with problem drinking in the older group (P = 0.080) but not in the younger group. The ALDH2 genotypes consistently and strongly affected the alcohol sensitivity, drinking behavior and problem drinking in both the younger and older group. We for the first time observed a significant difference in alcohol sensitivity between ADH1B*1/*2 and ADH1B*2/*2 in older men aged 65 and above.

  15. Alcohol dehydrogenase and aldehyde dehydrogenase gene polymorphisms, alcohol intake and the risk of colorectal cancer in the European Prospective Investigation into Cancer and Nutrition study.

    PubMed

    Ferrari, P; McKay, J D; Jenab, M; Brennan, P; Canzian, F; Vogel, U; Tjønneland, A; Overvad, K; Tolstrup, J S; Boutron-Ruault, M-C; Clavel-Chapelon, F; Morois, S; Kaaks, R; Boeing, H; Bergmann, M; Trichopoulou, A; Katsoulis, M; Trichopoulos, D; Krogh, V; Panico, S; Sacerdote, C; Palli, D; Tumino, R; Peeters, P H; van Gils, C H; Bueno-de-Mesquita, B; Vrieling, A; Lund, E; Hjartåker, A; Agudo, A; Suarez, L R; Arriola, L; Chirlaque, M-D; Ardanaz, E; Sánchez, M-J; Manjer, J; Lindkvist, B; Hallmans, G; Palmqvist, R; Allen, N; Key, T; Khaw, K-T; Slimani, N; Rinaldi, S; Romieu, I; Boffetta, P; Romaguera, D; Norat, T; Riboli, E

    2012-12-01

    Heavy alcohol drinking is a risk factor of colorectal cancer (CRC), but little is known on the effect of polymorphisms in the alcohol-metabolizing enzymes, alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) on the alcohol-related risk of CRC in Caucasian populations. A nested case-control study (1269 cases matched to 2107 controls by sex, age, study centre and date of blood collection) was conducted within the European Prospective Investigation into Cancer and Nutrition (EPIC) to evaluate the impact of rs1229984 (ADH1B), rs1573496 (ADH7) and rs441 (ALDH2) polymorphisms on CRC risk. Using the wild-type variant of each polymorphism as reference category, CRC risk estimates were calculated using conditional logistic regression, with adjustment for matching factors. Individuals carrying one copy of the rs1229984(A) (ADH1B) allele (fast metabolizers) showed an average daily alcohol intake of 4.3 g per day lower than subjects with two copies of the rs1229984(G) allele (slow metabolizers) (P(diff)<0.01). None of the polymorphisms was associated with risk of CRC or cancers of the colon or rectum. Heavy alcohol intake was more strongly associated with CRC risk among carriers of the rs1573496(C) allele, with odds ratio equal to 2.13 (95% confidence interval: 1.26-3.59) compared with wild-type subjects with low alcohol consumption (P(interaction)=0.07). The rs1229984(A) (ADH1B) allele was associated with a reduction in alcohol consumption. The rs1229984 (ADH1B), rs1573496 (ADH7) and rs441 (ALDH2) polymorphisms were not associated with CRC risk overall in Western-European populations. However, the relationship between alcohol and CRC risk might be modulated by the rs1573496 (ADH7) polymorphism.

  16. Overview of the role of alcohol dehydrogenase and aldehyde dehydrogenase and their variants in the genesis of alcohol-related pathology.

    PubMed

    Crabb, David W; Matsumoto, Michinaga; Chang, David; You, Min

    2004-02-01

    Alcohol dehydrogenase (ADH) and mitochondrial aldehyde dehydrogenase (ALDH2) are responsible for metabolizing the bulk of ethanol consumed as part of the diet and their activities contribute to the rate of ethanol elimination from the blood. They are expressed at highest levels in liver, but at lower levels in many tissues. This pathway probably evolved as a detoxification mechanism for environmental alcohols. However, with the consumption of large amounts of ethanol, the oxidation of ethanol can become a major energy source and, particularly in the liver, interferes with the metabolism of other nutrients. Polymorphic variants of the genes for these enzymes encode enzymes with altered kinetic properties. The pathophysiological effects of these variants may be mediated by accumulation of acetaldehyde; high-activity ADH variants are predicted to increase the rate of acetaldehyde generation, while the low-activity ALDH2 variant is associated with an inability to metabolize this compound. The effects of acetaldehyde may be expressed either in the cells generating it, or by delivery of acetaldehyde to various tissues by the bloodstream or even saliva. Inheritance of the high-activity ADH beta2, encoded by the ADH2*2 gene, and the inactive ALDH2*2 gene product have been conclusively associated with reduced risk of alcoholism. This association is influenced by gene-environment interactions, such as religion and national origin. The variants have also been studied for association with alcoholic liver disease, cancer, fetal alcohol syndrome, CVD, gout, asthma and clearance of xenobiotics. The strongest correlations found to date have been those between the ALDH2*2 allele and cancers of the oro-pharynx and oesophagus. It will be important to replicate other interesting associations between these variants and other cancers and heart disease, and to determine the biochemical mechanisms underlying the associations.

  17. The activity of class I, II, III and IV alcohol dehydrogenase isoenzymes and aldehyde dehydrogenase in the sera of bladder cancer patients.

    PubMed

    Orywal, Karolina; Jelski, Wojciech; Werel, Tadeusz; Szmitkowski, Maciej

    2017-01-01

    Studies on alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) activity in the sera of patients with malignant neoplasms show that cancer cells in many organs may release ADH isoenzymes into the blood. The aim of this study was to investigate the differences in the activity of ADH isoenzymes and ALDH in the sera of patients with bladder cancer (BCa), and with different grades of the disease. Blood samples were taken from 39 patients with BCa (15 patients with low-grade and 24 with high-grade BCa) and from 60 healthy subjects. Class III and IV of ADH and total ADH activity were measured using the photometric method, while class I and II ADH and ALDH activity using the fluorometric method with class-specific fluorogenic substrates. The activity of the class I ADH isoenzyme and total ADH was significantly higher in the sera of BCa patients as compared to control group. Analysis of ALDH activity did not show statistically significant differences between the tested groups. Significantly higher total activity of ADH in comparison to control was found in both, low-grade and high-grade BCa group. The activity of ADH class I was also significantly higher in high-grade BCa group when compared to low-grade patients and controls. The increase of total ADH activity in the sera of BCa patients seems to be caused by isoenzymes released from cancerous cells. The higher activity of ADH I probably resulted from metastatic tumors as significant increase was detected only in the sera of high-grade bladder cancer patients.

  18. Genome-wide identification and analysis of the aldehyde dehydrogenase (ALDH) gene superfamily in apple (Malus × domestica Borkh.).

    PubMed

    Li, Xiaoqin; Guo, Rongrong; Li, Jun; Singer, Stacy D; Zhang, Yucheng; Yin, Xiangjing; Zheng, Yi; Fan, Chonghui; Wang, Xiping

    2013-10-01

    Aldehyde dehydrogenases (ALDHs) represent a protein superfamily encoding NAD(P)(+)-dependent enzymes that oxidize a wide range of endogenous and exogenous aliphatic and aromatic aldehydes. In plants, they are involved in many biological processes and play a role in the response to environmental stress. In this study, a total of 39 ALDH genes from ten families were identified in the apple (Malus × domestica Borkh.) genome. Synteny analysis of the apple ALDH (MdALDH) genes indicated that segmental and tandem duplications, as well as whole genome duplications, have likely contributed to the expansion and evolution of these gene families in apple. Moreover, synteny analysis between apple and Arabidopsis demonstrated that several MdALDH genes were found in the corresponding syntenic blocks of Arabidopsis, suggesting that these genes appeared before the divergence of lineages that led to apple and Arabidopsis. In addition, phylogenetic analysis, as well as comparisons of exon-intron and protein structures, provided further insight into both their evolutionary relationships and their putative functions. Tissue-specific expression analysis of the MdALDH genes demonstrated diverse spatiotemporal expression patterns, while their expression profiles under abiotic stress and various hormone treatments indicated that many MdALDH genes were responsive to high salinity and drought, as well as different plant hormones. This genome-wide identification, as well as characterization of evolutionary relationships and expression profiles, of the apple MdALDH genes will not only be useful for the further analysis of ALDH genes and their roles in stress response, but may also aid in the future improvement of apple stress tolerance. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  19. Neuroprotective effects of aldehyde dehydrogenase 2 activation in rotenone-induced cellular and animal models of parkinsonism

    PubMed Central

    Chiu, Ching-Chi; Yeh, Tu-Hsueh; Lai, Szu-Chia; Wu-Chou, Yah-Huei; Chen, Che-Hong; Mochly-Rosen, Daria; Huang, Yin-Cheng; Chen, Yu-Jie; Chen, Chao-Lang; Chang, Ya-Ming; Wang, Hung-Li; Lu, Chin-Song

    2015-01-01

    Many studies have shown that mitochondrial aldehyde dehydrogenase 2 (ALDH2) functions as a cellular protector against oxidative stress by detoxification of cytotoxic aldehydes. Within dopaminergic neurons, dopamine is metabolized by monoamine oxidase to yield 3,4-dihydroxyphenylacetaldehyde (DOPAL) then converts to a less toxic acid product by ALDH. The highly toxic and reactive DOPAL has been hypothesized to contribute to the selective neurodegeneration in Parkinson’s disease (PD). In this study, we investigated the neuroprotective mechanism and therapeutic effect of ALDH2 in rotenone models for parkinsonism. Overexpression of wild-type ALDH2 gene, but not the enzymatically deficient mutant ALDH2*2 (E504K), reduced rotenone-induced cell death. Application of a potent activator of ALDH2, Alda-1, was effective in protecting against rotenone-induced apoptotic cell death in both SH-SY5Y cells and primary cultured substantia nigra (SN) dopaminergic neurons. In addition, intraperitoneal administration of Alda-1 significantly reduced rotenone- or MPTP-induced death of SN tyrosine hydroxylase (TH)-positive dopaminergic neurons. The attenuation of rotenone-induced apoptosis by Alda-1 resulted from decreasing ROS accumulation, reversal of mitochondrial membrane potential depolarization, and inhibition of activation of proteins related to mitochondrial apoptotic pathway. The present study demonstrates that ALDH2 plays a crucial role in maintaining normal mitochondrial function to protect against neurotoxicity and that Alda-1 is effective in ameliorating mitochondrial dysfunction and inhibiting mitochondria-mediated apoptotic pathway. These results indicate that ALDH2 activation could be a neuroprotective therapy for PD. PMID:25263579

  20. Hypoglycemic drugs induce antioxidant aldehyde dehydrogenase activity and remain high in patients with glycemic control in type 2 diabetes.

    PubMed

    Picazo, Alejandra; Jiménez-Osorio, Angélica S; Zúñiga-Mejía, Porfirio; Pedraza-Chaverri, José; Monroy, Adriana; Rodríguez-Arellano, M Eunice; Barrera-Oviedo, Diana

    2017-04-05

    The antioxidant system results essential to control and prevent lipid peroxidation due to stress damage in type 2 diabetes. An example is aldehyde dehydrogenase (ALDH), an enzyme that is involved in the detoxification of aldehydes formed during lipid peroxidation. This study was conducted to evaluate ALDH activity and to determine their association with hypoglycemic treatment in type 2 diabetes patients. The study population consisted of 422 Mexican subjects: a control group and type 2 diabetes patients. Type 2 diabetes patients were re-classified as those with or without hypoglycemic treatment and those with or without glycemic control (according to glycated hemoglobin (HbA1c)). Clinical parameters, antioxidant enzyme activities (ALDH, superoxide dismutase (SOD), catalase and glutathione peroxidase) and oxidative markers (reactive oxygen species and thiobarbituric acid reactive substances (TBARS)) were evaluated. The activity of antioxidant enzymes and oxidative stress markers were higher in type 2 diabetes patients with hypoglycemic treatment and without glycemic control than control group. The activity of ALDH and SOD remained high in type 2 diabetes patients with moderate glycemic control while only ALDH's remained high in type 2 diabetes patients with tight glycemic control. Increased ALDH and SOD activities were associated with hypoglycemic therapy. TBARS levels were associated with glycemic control. The persistence of high ALDH and SOD activities in type 2 diabetes patients with glycemic control may be to avoid a significant damage due to the increase in reactive oxygen species and TBARS. It is possible that this new oxidative status prevented the development the classical complications of diabetes. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Simultaneous involvement of a tungsten-containing aldehyde:ferredoxin oxidoreductase and a phenylacetaldehyde dehydrogenase in anaerobic phenylalanine metabolism.

    PubMed

    Debnar-Daumler, Carlotta; Seubert, Andreas; Schmitt, Georg; Heider, Johann

    2014-01-01

    Anaerobic phenylalanine metabolism in the denitrifying betaproteobacterium Aromatoleum aromaticum is initiated by conversion of phenylalanine to phenylacetate, which is further metabolized via benzoyl-coenzyme A (CoA). The formation of phenylacetate is catalyzed by phenylalanine transaminase, phenylpyruvate decarboxylase, and a phenylacetaldehyde-oxidizing enzyme. The presence of these enzymes was detected in extracts of cells grown with phenylalanine and nitrate. We found that two distinct enzymes are involved in the oxidation of phenylacetaldehyde to phenylacetate, an aldehyde:ferredoxin oxidoreductase (AOR) and a phenylacetaldehyde dehydrogenase (PDH). Based on sequence comparison, growth studies with various tungstate concentrations, and metal analysis of the enriched enzyme, AOR was shown to be a tungsten-containing enzyme, necessitating specific cofactor biosynthetic pathways for molybdenum- and tungsten-dependent enzymes simultaneously. We predict from the genome sequence that most enzymes of molybdopterin biosynthesis are shared, while the molybdate/tungstate uptake systems are duplicated and specialized paralogs of the sulfur-inserting MoaD and the metal-inserting MoeA proteins seem to be involved in dedicating biosynthesis toward molybdenum or tungsten cofactors. We also characterized PDH biochemically and identified both NAD(+) and NADP(+) as electron acceptors. We identified the gene coding for the enzyme and purified a recombinant Strep-tagged PDH variant. The homotetrameric enzyme is highly specific for phenylacetaldehyde, has cooperative kinetics toward the substrate, and shows considerable substrate inhibition. Our data suggest that A. aromaticum utilizes PDH as the primary enzyme during anaerobic phenylalanine degradation, whereas AOR is not essential for the metabolic pathway. We hypothesize a function as a detoxifying enzyme if high aldehyde concentrations accumulate in the cytoplasm, which would lead to substrate inhibition of PDH.

  2. Simultaneous Involvement of a Tungsten-Containing Aldehyde:Ferredoxin Oxidoreductase and a Phenylacetaldehyde Dehydrogenase in Anaerobic Phenylalanine Metabolism

    PubMed Central

    Debnar-Daumler, Carlotta; Seubert, Andreas; Schmitt, Georg

    2014-01-01

    Anaerobic phenylalanine metabolism in the denitrifying betaproteobacterium Aromatoleum aromaticum is initiated by conversion of phenylalanine to phenylacetate, which is further metabolized via benzoyl-coenzyme A (CoA). The formation of phenylacetate is catalyzed by phenylalanine transaminase, phenylpyruvate decarboxylase, and a phenylacetaldehyde-oxidizing enzyme. The presence of these enzymes was detected in extracts of cells grown with phenylalanine and nitrate. We found that two distinct enzymes are involved in the oxidation of phenylacetaldehyde to phenylacetate, an aldehyde:ferredoxin oxidoreductase (AOR) and a phenylacetaldehyde dehydrogenase (PDH). Based on sequence comparison, growth studies with various tungstate concentrations, and metal analysis of the enriched enzyme, AOR was shown to be a tungsten-containing enzyme, necessitating specific cofactor biosynthetic pathways for molybdenum- and tungsten-dependent enzymes simultaneously. We predict from the genome sequence that most enzymes of molybdopterin biosynthesis are shared, while the molybdate/tungstate uptake systems are duplicated and specialized paralogs of the sulfur-inserting MoaD and the metal-inserting MoeA proteins seem to be involved in dedicating biosynthesis toward molybdenum or tungsten cofactors. We also characterized PDH biochemically and identified both NAD+ and NADP+ as electron acceptors. We identified the gene coding for the enzyme and purified a recombinant Strep-tagged PDH variant. The homotetrameric enzyme is highly specific for phenylacetaldehyde, has cooperative kinetics toward the substrate, and shows considerable substrate inhibition. Our data suggest that A. aromaticum utilizes PDH as the primary enzyme during anaerobic phenylalanine degradation, whereas AOR is not essential for the metabolic pathway. We hypothesize a function as a detoxifying enzyme if high aldehyde concentrations accumulate in the cytoplasm, which would lead to substrate inhibition of PDH. PMID:24214948

  3. Cardiac Mitochondrial Respiratory Dysfunction and Tissue Damage in Chronic Hyperglycemia Correlate with Reduced Aldehyde Dehydrogenase-2 Activity

    PubMed Central

    Deshpande, Mandar; Thandavarayan, Rajarajan A.; Xu, Jiang; Yang, Xiao-Ping; Palaniyandi, Suresh S.

    2016-01-01

    Aldehyde dehydrogenase (ALDH) 2 is a mitochondrial isozyme of the heart involved in the metabolism of toxic aldehydes produced from oxidative stress. We hypothesized that hyperglycemia-mediated decrease in ALDH2 activity may impair mitochondrial respiration and ultimately result in cardiac damage. A single dose (65 mg/kg; i.p.) streptozotocin injection to rats resulted in hyperglycemia with blood glucose levels of 443 ± 9 mg/dl versus 121 ± 7 mg/dl in control animals, p<0.0001, N = 7–11. After 6 months of diabetes mellitus (DM) induction, the rats were sacrificed after recording the functionality of their hearts. Increase in the cardiomyocyte cross sectional area (446 ± 32 μm2 Vs 221 ± 10 μm2; p<0.0001) indicated cardiac hypertrophy in DM rats. Both diastolic and systolic dysfunctions were observed with DM rats compared to controls. Most importantly, myocardial ALDH2 activity and levels were reduced, and immunostaining for 4HNE protein adducts was increased in DM hearts compared to controls. The mitochondrial oxygen consumption rate (OCR), an index of mitochondrial respiration, was decreased in mitochondria isolated from DM hearts compared to controls (p<0.0001). Furthermore, the rate of mitochondrial respiration and the increase in carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP)-induced maximal respiration were also decreased with chronic hyperglycemia. Chronic hyperglycemia reduced mitochondrial OXPHOS proteins. Reduced ALDH2 activity was correlated with mitochondrial dysfunction, pathological remodeling and cardiac dysfunction, respectively. Our results suggest that chronic hyperglycemia reduces ALDH2 activity, leading to mitochondrial respiratory dysfunction and consequently cardiac damage and dysfunction. PMID:27736868

  4. Monovalent cations requirements for the stability of betaine aldehyde dehydrogenase from Pseudomonas aeruginosa, porcine kidney and amaranth leaves.

    PubMed

    Valenzuela-Soto, Elisa M; Velasco-García, Roberto; Mújica-Jiménez, Carlos; Gaviria-González, L Laraí; Muñoz-Clares, Rosario A

    2003-02-01

    Betaine aldehyde dehydrogenase from the human pathogen Pseudomonas aeruginosa requires K(+) ions for maintenance of its active conformation. In order to explore if this property is shared by other BADHs of different origins and to further understand the mechanism underlying the effects of these ions, we carried out a comparative study on the stability and quaternary structure of P. aeruginosa, porcine kidney and amaranth leaves BADHs in the absence of K(+) ions. At low enzyme concentrations, the bacterial and porcine enzymes were totally inactivated upon removal of K(+) following biphasic and monophasic kinetics, respectively, whereas the amaranth enzyme retained its activity. Inactivation of P. aeruginosa BADH was much faster than that of the porcine enzyme. The oxidized coenzyme protected both enzymes against inactivation by the absence of K(+), whereas betaine aldehyde afforded partial protection to the bacterial BADH and increased the inactivation rate of the porcine. Reactivation of the inactive enzymes, by adding back to the incubation medium K(+) ions, was dependent on enzyme concentration, suggesting that enzyme dissociation takes place in the absence of K(+). In the bacterial enzyme, NH(4)(+) but not Na(+) ions could mimic the effects of K(+), whereas the three cations tested reactivated porcine BADH, indicating a requirement of this enzyme for high ionic strength rather than for a specific monovalent cation. Size exclusion chromatography of the inactivated enzymes confirmed that K(+) ions or other monovalent cations are required for the maintenance of the quaternary structure of these two BADHs. At pH 7.0, in the absence of K(+) in a buffer of low ionic strength, the active tetrameric form of P. aeruginosa BADH dissociated into inactive monomers and that of porcine kidney BADH into inactive dimers. Once reactivated, both enzymes reassociated into active tetramers.

  5. Association between aldehyde dehydrogenase 2 polymorphisms and the incidence of diabetic retinopathy among Japanese subjects with type 2 diabetes mellitus.

    PubMed

    Morita, Kazunori; Saruwatari, Junji; Miyagawa, Haruna; Uchiyashiki, Yoshihiro; Oniki, Kentaro; Sakata, Misaki; Kajiwara, Ayami; Yoshida, Akira; Jinnouchi, Hideaki; Nakagawa, Kazuko

    2013-09-13

    Mitochondrial aldehyde dehydrogenase 2 (ALDH2) detoxifies reactive aldehydes in the micro- and macrovasculature. These substrates, including methylglyoxal and 4-hydroxynonenal formed from glucose and lipids, cause protein carbonylation and mitochondrial dysfunction, forming advanced glycation end products (AGEs). The present study aimed to confirm the association between the inactive ALDH2*2 allele and diabetic retinopathy (DR). A retrospective longitudinal analysis was conducted, among 234 Japanese patients with type 2 diabetes mellitus (DM) (156 males and 78 females) who had no DR signs at baseline and were treated for more than half a year. The ALDH2*1/*2 alleles were determined using a real-time TaqMan allelic discrimination assay. Multivariate-adjusted hazard ratios (HRs) and 95% confidential intervals (CIs) for the cumulative incidence of the development of DR were examined using a Cox proportional hazard model, taking drinking habits and the serum γ-glutamyltransferase (GGT) level into consideration. The frequency of the ALDH2*2 allele was 22.3%. Fifty-two subjects cumulatively developed DR during the follow-up period of 5.5 ± 2.5 years. The ALDH2*2 allele carriers had a significantly higher incidence of DR than the non-carriers (HR: 1.92; P = 0.02). The incidence of DR was significantly higher in the drinkers with the ALDH2*2 allele than in those with the ALDH2*1/*1 genotype (HR: 2.61; P = 0.03), while the incidence of DR in the non-drinkers did not differ significantly between the ALDH2 genotype groups (P > 0.05). The incidence of DR was significantly higher in the ALDH2*2 allele carriers with a high GGT level than in the non-carriers with a high or low GGT level (HR: 2.45; P = 0.03; and HR: 2.63; P = 0.03, respectively). To the best of our knowledge, this is the first report of a significant association between the ALDH2*2 allele and the incidence of DR. These findings provide additional evidence that ALDH2 protects both microvasculature and

  6. Novel Inhibitors of Mitochondrial sn-Glycerol 3-phosphate Dehydrogenase

    PubMed Central

    Orr, Adam L.; Ashok, Deepthi; Sarantos, Melissa R.; Ng, Ryan; Shi, Tong; Gerencser, Akos A.; Hughes, Robert E.; Brand, Martin D.

    2014-01-01

    Mitochondrial sn-glycerol 3-phosphate dehydrogenase (mGPDH) is a ubiquinone-linked enzyme in the mitochondrial inner membrane best characterized as part of the glycerol phosphate shuttle that transfers reducing equivalents from cytosolic NADH into the mitochondrial electron transport chain. Despite the widespread expression of mGPDH and the availability of mGPDH-null mice, the physiological role of this enzyme remains poorly defined in many tissues, likely because of compensatory pathways for cytosolic regeneration of NAD+ and mechanisms for glycerol phosphate metabolism. Here we describe a novel class of cell-permeant small-molecule inhibitors of mGPDH (iGP) discovered through small-molecule screening. Structure-activity analysis identified a core benzimidazole-phenyl-succinamide structure as being essential to inhibition of mGPDH while modifications to the benzimidazole ring system modulated both potency and off-target effects. Live-cell imaging provided evidence that iGPs penetrate cellular membranes. Two compounds (iGP-1 and iGP-5) were characterized further to determine potency and selectivity and found to be mixed inhibitors with IC50 and Ki values between ∼1–15 µM. These novel mGPDH inhibitors are unique tools to investigate the role of glycerol 3-phosphate metabolism in both isolated and intact systems. PMID:24587137

  7. Aldehyde dehydrogenases of the rat colon: comparison with other tissues of the alimentary tract and the liver.

    PubMed

    Koivisto, T; Salaspuro, M

    1996-05-01

    Intracolonic bacteria have previously been shown to produce substantial amounts of acetaldehyde during ethanol oxidation, and it has been suggested that this acetaldehyde might be associated with alcohol-related colonic disorders, as well as other alcohol-induced organ injuries. The capacity of colonic mucosa to remove this bacterial acetaldehyde by aldehyde dehydrogenase (ALDH) is, however, poorly known. We therefore measured ALDH activities and determined ALDH isoenzyme profiles from different subcellular fractions of rat colonic mucosa. For comparison, hepatic, gastric, and small intestinal samples were studied similarly. Alcohol dehydrogenase (ADH) activities were also measured from all of these tissues. Rat colonic mucosa was found to possess detectable amounts of ALDH activity with both micromolar and millimolar acetaldehyde concentrations and in all subcellular fractions. The ALDH activities of colonic mucosa were, however, generally low when compared with the liver and stomach, and they also tended to be lower than in small intestine. Mitochondrial low K(m) ALDH2 and cytosolic ALDH with low K(m) for acetaldehyde were expressed in the colonic mucosa, whereas some cytosolic high K(m) isoenzymes found in the small intestine and stomach were not detectable in colonic samples. Cytosolic ADH activity corresponded well to ALDH activity in different tissues: in colonic mucosa, it was approximately 6 times lower than in the liver and about one-half of gastric ADH activity. ALDH activity of the colonic mucosa should, thus, be sufficient for the removal of acetaldehyde produced by colonic mucosal ADH during ethanol oxidation. It may, however, be insufficient for the removal of the acetaldehyde produced by intracolonic bacteria. This may lead to the accumulation of acetaldehyde in the colon and colonic mucosa after ingestion of ethanol that might, at least after chronic heavy alcohol consumption, contribute to the development of alcohol-related colonic morbidity

  8. Conserved catalytic residues of the ALDH1L1 aldehyde dehydrogenase domain control binding and discharging of the coenzyme.

    PubMed

    Tsybovsky, Yaroslav; Krupenko, Sergey A

    2011-07-01

    The C-terminal domain (C(t)-FDH) of 10-formyltetrahydrofolate dehydrogenase (FDH, ALDH1L1) is an NADP(+)-dependent oxidoreductase and a structural and functional homolog of aldehyde dehydrogenases. Here we report the crystal structures of several C(t)-FDH mutants in which two essential catalytic residues adjacent to the nicotinamide ring of bound NADP(+), Cys-707 and Glu-673, were replaced separately or simultaneously. The replacement of the glutamate with an alanine causes irreversible binding of the coenzyme without any noticeable conformational changes in the vicinity of the nicotinamide ring. Additional replacement of cysteine 707 with an alanine (E673A/C707A double mutant) did not affect this irreversible binding indicating that the lack of the glutamate is solely responsible for the enhanced interaction between the enzyme and the coenzyme. The substitution of the cysteine with an alanine did not affect binding of NADP(+) but resulted in the enzyme lacking the ability to differentiate between the oxidized and reduced coenzyme: unlike the wild-type C(t)-FDH/NADPH complex, in the C707A mutant the position of NADPH is identical to the position of NADP(+) with the nicotinamide ring well ordered within the catalytic center. Thus, whereas the glutamate restricts the affinity for the coenzyme, the cysteine is the sensor of the coenzyme redox state. These conclusions were confirmed by coenzyme binding experiments. Our study further suggests that the binding of the coenzyme is additionally controlled by a long-range communication between the catalytic center and the coenzyme-binding domain and points toward an α-helix involved in the adenine moiety binding as a participant of this communication.

  9. Conserved Catalytic Residues of the ALDH1L1 Aldehyde Dehydrogenase Domain Control Binding and Discharging of the Coenzyme*

    PubMed Central

    Tsybovsky, Yaroslav; Krupenko, Sergey A.

    2011-01-01

    The C-terminal domain (Ct-FDH) of 10-formyltetrahydrofolate dehydrogenase (FDH, ALDH1L1) is an NADP+-dependent oxidoreductase and a structural and functional homolog of aldehyde dehydrogenases. Here we report the crystal structures of several Ct-FDH mutants in which two essential catalytic residues adjacent to the nicotinamide ring of bound NADP+, Cys-707 and Glu-673, were replaced separately or simultaneously. The replacement of the glutamate with an alanine causes irreversible binding of the coenzyme without any noticeable conformational changes in the vicinity of the nicotinamide ring. Additional replacement of cysteine 707 with an alanine (E673A/C707A double mutant) did not affect this irreversible binding indicating that the lack of the glutamate is solely responsible for the enhanced interaction between the enzyme and the coenzyme. The substitution of the cysteine with an alanine did not affect binding of NADP+ but resulted in the enzyme lacking the ability to differentiate between the oxidized and reduced coenzyme: unlike the wild-type Ct-FDH/NADPH complex, in the C707A mutant the position of NADPH is identical to the position of NADP+ with the nicotinamide ring well ordered within the catalytic center. Thus, whereas the glutamate restricts the affinity for the coenzyme, the cysteine is the sensor of the coenzyme redox state. These conclusions were confirmed by coenzyme binding experiments. Our study further suggests that the binding of the coenzyme is additionally controlled by a long-range communication between the catalytic center and the coenzyme-binding domain and points toward an α-helix involved in the adenine moiety binding as a participant of this communication. PMID:21540484

  10. Arabidopsis aldehyde dehydrogenase 10 family members confer salt tolerance through putrescine-derived 4-aminobutyrate (GABA) production

    PubMed Central

    Zarei, Adel; Trobacher, Christopher P.; Shelp, Barry J.

    2016-01-01

    Polyamines represent a potential source of 4-aminobutyrate (GABA) in plants exposed to abiotic stress. Terminal catabolism of putrescine in Arabidopsis thaliana involves amine oxidase and the production of 4-aminobutanal, which is a substrate for NAD+-dependent aminoaldehyde dehydrogenase (AMADH). Here, two AMADH homologs were chosen (AtALDH10A8 and AtALDH10A9) as candidates for encoding 4-aminobutanal dehydrogenase activity for GABA synthesis. The two genes were cloned and soluble recombinant proteins were produced in Escherichia coli. The pH optima for activity and catalytic efficiency of recombinant AtALDH10A8 with 3-aminopropanal as substrate was 10.5 and 8.5, respectively, whereas the optima for AtALDH10A9 were approximately 9.5. Maximal activity and catalytic efficiency were obtained with NAD+ and 3-aminopropanal, followed by 4-aminobutanal; negligible activity was obtained with betaine aldehyde. NAD+ reduction was accompanied by the production of GABA and β-alanine, respectively, with 4-aminobutanal and 3-aminopropanal as substrates. Transient co-expression systems using Arabidopsis cell suspension protoplasts or onion epidermal cells and several organelle markers revealed that AtALDH10A9 was peroxisomal, but AtALDH10A8 was cytosolic, although the N-terminal 140 amino acid sequence of AtALDH10A8 localized to the plastid. Root growth of single loss-of-function mutants was more sensitive to salinity than wild-type plants, and this was accompanied by reduced GABA accumulation. PMID:27725774

  11. Long-Chain Aldehyde Dehydrogenase That Participates in n-Alkane Utilization and Wax Ester Synthesis in Acinetobacter sp. Strain M-1

    PubMed Central

    Ishige, Takeru; Tani, Akio; Sakai, Yasuyoshi; Kato, Nobuo

    2000-01-01

    A long-chain aldehyde dehydrogenase, Ald1, was found in a soluble fraction of Acinetobacter sp. strain M-1 cells grown on n-hexadecane as a sole carbon source. The gene (ald1) was cloned from the chromosomal DNA of the bacterium. The open reading frame of ald1 was 1,512 bp long, corresponding to a protein of 503 amino acid residues (molecular mass, 55,496 Da), and the deduced amino acid sequence showed high similarity to those of various aldehyde dehydrogenases. The ald1 gene was stably expressed in Escherichia coli, and the gene product (recombinant Ald1 [rAld1]) was purified to apparent homogeneity by gel electrophoresis. rAld1 showed enzyme activity toward n-alkanals (C4 to C14), with a preference for longer carbon chains within the tested range; the highest activity was obtained with tetradecanal. The ald1 gene was disrupted by homologous recombination on the Acinetobacter genome. Although the ald1 disruptant (ald1Δ) strain still had the ability to grow on n-hexadecane to some extent, its aldehyde dehydrogenase activity toward n-tetradecanal was reduced to half the level of the wild-type strain. Under nitrogen-limiting conditions, the accumulation of intracellular wax esters in the ald1Δ strain became much lower than that in the wild-type strain. These and other results imply that a soluble long-chain aldehyde dehydrogenase indeed plays important roles both in growth on n-alkane and in wax ester formation in Acinetobacter sp. strain M-1. PMID:10919810

  12. Electrophilic Aldehydes Generated by Sperm Metabolism Activate Mitochondrial Reactive Oxygen Species Generation and Apoptosis by Targeting Succinate Dehydrogenase*

    PubMed Central

    Aitken, R. John; Whiting, Sara; De Iuliis, Geoffry N.; McClymont, Samantha; Mitchell, Lisa A.; Baker, Mark A.

    2012-01-01

    Oxidative stress is a major cause of defective sperm function in cases of male infertility. Such stress is known to be associated with high levels of superoxide production by the sperm mitochondria; however, the causes of this aberrant activity are unknown. Here we show that electrophilic aldehydes such as 4-hydroxynonenal (4HNE) and acrolein, generated as a result of lipid peroxidation, target the mitochondria of human spermatozoa and stimulate mitochondrial superoxide generation in a dose- and time-dependent manner. The activation of mitochondrial electron leakage by 4HNE is shown to involve the disruption of succinate dehydrogenase activity and subsequent activation of an intrinsic apoptotic cascade beginning with a loss of mitochondrial membrane potential and terminating in oxidative DNA adduct formation, DNA strand breakage, and cell death. A tight correlation between spontaneous mitochondrial superoxide generation and 4HNE content (R2 = 0.89) in untreated populations of human spermatozoa emphasized the pathophysiological significance of these findings. The latter also provide a biochemical explanation for the self-perpetuating nature of oxidative stress in the male germ line, with the products of lipid peroxidation stimulating free radical generation by the sperm mitochondria in a positive feedback loop. PMID:22851170

  13. Deficient expression of aldehyde dehydrogenase 1A1 is consistent with increased sensitivity of Gorlin syndrome patients to radiation carcinogenesis

    SciTech Connect

    Wright, Aaron T.; Magnaldo, Thierry; Sontag, Ryan L.; Anderson, Lindsey N.; Sadler, Natalie C.; Piehowski, Paul D.; Gache, Yannick; Weber, Thomas J.

    2013-11-27

    Human phenotypes that are highly susceptible to radiation carcinogenesis have been identified. Sensitive phenotypes often display robust regulation of molecular features that modify biological response, which can facilitate identification of relevant pathways/networks. Here we interrogate primary dermal fibroblasts isolated from Gorlin syndrome patients (GDFs), who display a pronounced tumorigenic response to radiation, in comparison to normal human dermal fibroblasts (NHDFs). Our approach exploits newly developed thiol-reactive probes with a flexible click chemistry functional group to define changes in protein thiol profiles in live cell studies, which minimizes artifacts associated with cell lysis. We observe qualitative differences in protein thiol profiles by SDS-PAGE analysis when detection by iodoacetamide vs maleimide probe chemistries are compared, and pretreatment of cells with hydrogen peroxide eliminates detection of the majority of SDS-PAGE bands. Redox probes revealed deficient expression of an apparent 55 kDa protein thiol in GDFs from independent donors, compared with NHDFs. Proteomics tentatively identified this protein as aldehyde dehydrogenase 1A1 (ALDH1A1), a key enzyme regulating retinoic acid synthesis, and this deficiency was confirmed by Western blot. Redox probes revealed additional protein thiol differences between GDFs and NHDFs, including radiation responsive annexin family members. Our results indicate a multifactorial basis for the unusual sensitivity of Gorlin syndrome to radiation carcinogenesis, and the pathways identified have plausible implications for radiation health effects.

  14. Aldehyde Dehydrogenase 2 (ALDH2) Polymorphism and the Risk of Alcoholic Liver Cirrhosis among East Asians: A Meta-Analysis

    PubMed Central

    He, Lei; Luo, Hesheng

    2016-01-01

    Purpose The aldehyde dehydrogenase 2 (ALDH2) gene has been implicated in the development of alcoholic liver cirrhosis (ALC) in East Asians. However, the results are inconsistent. In this study, a meta-analysis was performed to assess the associations between the ALDH2 polymorphism and the risk of ALC. Materials and Methods Relevant studies were retrieved by searching PubMed, Web of Science, CNKI, Wanfang and Veipu databases up to January 10, 2015. Pooled odds ratio (OR) and 95% confidence interval (CI) were calculated using either the fixed- or random effects model. Results A total of twelve case-control studies included 1003 cases and 2011 controls were included. Overall, the ALDH2 polymorphism was associated with a decreased risk of ALC (*1/*2 vs. *1/*1: OR=0.78, 95% CI: 0.61–0.99). However, in stratification analysis by country, we failed to detect any association among Chinese, Korean or Japanese populations. Conclusion The pooled evidence suggests that ALDH2 polymorphism may be an important protective factor for ALC in East Asians. PMID:27189280

  15. Deficient expression of aldehyde dehydrogenase 1A1 is consistent with increased sensitivity of Gorlin syndrome patients to radiation carcinogenesis

    DOE PAGES

    Wright, Aaron T.; Magnaldo, Thierry; Sontag, Ryan L.; ...

    2013-11-27

    Human phenotypes that are highly susceptible to radiation carcinogenesis have been identified. Sensitive phenotypes often display robust regulation of molecular features that modify biological response, which can facilitate identification of relevant pathways/networks. Here we interrogate primary dermal fibroblasts isolated from Gorlin syndrome patients (GDFs), who display a pronounced tumorigenic response to radiation, in comparison to normal human dermal fibroblasts (NHDFs). Our approach exploits newly developed thiol-reactive probes with a flexible click chemistry functional group to define changes in protein thiol profiles in live cell studies, which minimizes artifacts associated with cell lysis. We observe qualitative differences in protein thiol profilesmore » by SDS-PAGE analysis when detection by iodoacetamide vs maleimide probe chemistries are compared, and pretreatment of cells with hydrogen peroxide eliminates detection of the majority of SDS-PAGE bands. Redox probes revealed deficient expression of an apparent 55 kDa protein thiol in GDFs from independent donors, compared with NHDFs. Proteomics tentatively identified this protein as aldehyde dehydrogenase 1A1 (ALDH1A1), a key enzyme regulating retinoic acid synthesis, and this deficiency was confirmed by Western blot. Redox probes revealed additional protein thiol differences between GDFs and NHDFs, including radiation responsive annexin family members. Our results indicate a multifactorial basis for the unusual sensitivity of Gorlin syndrome to radiation carcinogenesis, and the pathways identified have plausible implications for radiation health effects.« less

  16. Modeling-Dependent Protein Characterization of the Rice Aldehyde Dehydrogenase (ALDH) Superfamily Reveals Distinct Functional and Structural Features

    PubMed Central

    Kotchoni, Simeon O.; Jimenez-Lopez, Jose C.; Gao, Dongying; Edwards, Vincent; Gachomo, Emma W.; Margam, Venu M.; Seufferheld, Manfredo J.

    2010-01-01

    The completion of the rice genome sequence has made it possible to identify and characterize new genes and to perform comparative genomics studies across taxa. The aldehyde dehydrogenase (ALDH) gene superfamily encoding for NAD(P)+-dependent enzymes is found in all major plant and animal taxa. However, the characterization of plant ALDHs has lagged behind their animal- and prokaryotic-ALDH homologs. In plants, ALDHs are involved in abiotic stress tolerance, male sterility restoration, embryo development and seed viability and maturation. However, there is still no structural property-dependent functional characterization of ALDH protein superfamily in plants. In this paper, we identify members of the rice ALDH gene superfamily and use the evolutionary nesting events of retrotransposons and protein-modeling–based structural reconstitution to report the genetic and molecular and structural features of each member of the rice ALDH superfamily in abiotic/biotic stress responses and developmental processes. Our results indicate that rice-ALDHs are the most expanded plant ALDHs ever characterized. This work represents the first report of specific structural features mediating functionality of the whole families of ALDHs in an organism ever characterized. PMID:20634950

  17. E. coli metabolic protein aldehyde-alcohol dehydrogenase-E binds to the ribosome: a unique moonlighting action revealed

    PubMed Central

    Shasmal, Manidip; Dey, Sandip; Shaikh, Tanvir R.; Bhakta, Sayan; Sengupta, Jayati

    2016-01-01

    It is becoming increasingly evident that a high degree of regulation is involved in the protein synthesis machinery entailing more interacting regulatory factors. A multitude of proteins have been identified recently which show regulatory function upon binding to the ribosome. Here, we identify tight association of a metabolic protein aldehyde-alcohol dehydrogenase E (AdhE) with the E. coli 70S ribosome isolated from cell extract under low salt wash conditions. Cryo-EM reconstruction of the ribosome sample allows us to localize its position on the head of the small subunit, near the mRNA entrance. Our study demonstrates substantial RNA unwinding activity of AdhE which can account for the ability of ribosome to translate through downstream of at least certain mRNA helices. Thus far, in E. coli, no ribosome-associated factor has been identified that shows downstream mRNA helicase activity. Additionally, the cryo-EM map reveals interaction of another extracellular protein, outer membrane protein C (OmpC), with the ribosome at the peripheral solvent side of the 50S subunit. Our result also provides important insight into plausible functional role of OmpC upon ribosome binding. Visualization of the ribosome purified directly from the cell lysate unveils for the first time interactions of additional regulatory proteins with the ribosome. PMID:26822933

  18. Expression of the betaine aldehyde dehydrogenase gene in barley in response to osmotic stress and abscisic acid.

    PubMed

    Ishitani, M; Nakamura, T; Han, S Y; Takabe, T

    1995-01-01

    When subjected to salt stress or drought, some vascular plants such as barley respond with an increased accumulation of the osmoprotectant glycine betaine (betaine), being the last step of betaine synthesis catalyzed by betaine aldehyde dehydrogenase (BADH). We report here cloning and characterization of BADH cDNA from barley, a monocot, and the expression pattern of a BADH transcript. An open reading frame of 1515 bp encoded a protein which showed high homology to BADH enzymes present in other plants (spinach and sugar-beet) and in Escherichia coli. Transgenic tobacco plants harboring the clone expressed high levels of both BADH protein and its enzymatic activity. Northern blot analyses indicated that BADH mRNA levels increased almost 8-fold and 2-fold, respectively, in leaves and roots of barley plants grown in high-salt conditions, and that these levels decreased upon release of the stress, whereas they did not decrease under continuous salt stress. BADH transcripts also accumulate in response to water stress or drought, indicating a common response of the plant to osmotic changes that affect its water status. The addition of abscisic acid (ABA) to plants during growth also increased the levels of BADH transcripts dramatically, although the response was delayed when compared to that found for salt-stressed plants. Removal of plant roots before transferring the plants to high-salt conditions reduced only slightly the accumulation of BADH transcripts in the leaves.

  19. cDNA cloning and analysis of betaine aldehyde dehydrogenase, a salt inducible enzyme in sugar beet

    SciTech Connect

    McCue, K.F.; Hanson, A.D. )

    1990-05-01

    Betaine accumulates and serves as a compatible osmolyte in some plants subjected to drought or salinity stress. The last enzyme in the betaine biosynthetic pathway is betaine aldehyde dehydrogenase (BADH). The activity of BADH increases in response to increasing salinity levels. This increase in activity corresponds to an increase in protein detectable by immunoblotting, and to an increase in the translatable BADH mRNA. BADH was cloned from a cDNA library constructed in {lambda}gt10 using poly(A){sup +} RNA from sugar beets salinized to 500 mM NaCl. cDNAs were size selected (>1kb) before ligation into the vector, and the library was screened with a spinach BADH cDNA probe. Three nearly full length clones obtained were confirmed as BADH by their nucleotide and deduced amino acid homology to spinach BADH. Clones averaged 1.8 kb and contained open reading frames of 500 amino acids at 80% identity with spinach BADH. RNA gel blot analysis of poly(A){sup +} RNA indicated that salinization to 500 mM NaCl resulted in a 5-fold increase of BADH mRNA level.

  20. Salt-induction of betaine aldehyde dehydrogenase mRNA, protein, and enzymatic activity in sugar beet. [Beta vulgaris L

    SciTech Connect

    McCue, K.F.; Hanson, A.D. )

    1991-05-01

    In Chenopodiaceae such as sugar beet (Beta vulgaris L.), glycine betaine (betaine) accumulates in response to drought or salinity stress and functions in the cytoplasm as a compatible osmolyte. The last enzyme in the biosynthetic pathway, betaine aldehyde dehydrogenase (BADH), increases as much as 4-fold in response to rising salinity in the external medium. This increase is accompanied by an increase in both protein and mRNA levels. The steady state increases in BADH were examined at a series of NaCl concentrations from 100 to 500 mM NaCl. BADH protein levels were examined by native PAGE, and by western blot analysis using antibodies raised against BADH purified from spinach. mRNA levels were examined by northern plot analysis of total RNA isolated from the leaves and hybridized with a sugar beet BADH cDNA clone. The time course for BADH mRNA induction was determined in a salt shock experiment utilizing 400 mM NaCl added to the external growth medium. Disappearance of BADH was examined in a salt relief experiment using plants step-wise salinized to 500 mM NaCl and then returned to 0 mM NaCl.

  1. Aldehyde dehydrogenase, Ald4p, is a major component of mitochondrial fluorescent inclusion bodies in the yeast Saccharomyces cerevisiae

    PubMed Central

    Misonou, Yoshiko; Kikuchi, Maiko; Sato, Hiroshi; Inai, Tomomi; Kuroiwa, Tsuneyoshi; Tanaka, Kenji; Miyakawa, Isamu

    2014-01-01

    ABSTRACT When Saccharomyces cerevisiae strain 3626 was cultured to the stationary phase in a medium that contained glucose, needle-like structures that emitted autofluorescence were observed in almost all cells by fluorescence microscopy under UV excitation. The needle-like structures completely overlapped with the profile of straight elongated mitochondria. Therefore, these structures were designated as mitochondrial fluorescent inclusion bodies (MFIBs). The MFIB-enriched mitochondrial fractions were successfully isolated and 2D-gel electrophoresis revealed that a protein of 54 kDa was only highly concentrated in the fractions. Determination of the N-terminal amino acid sequence of the 54-kDa protein identified it as a mitochondrial aldehyde dehydrogenase, Ald4p. Immunofluorescence microscopy showed that anti-Ald4p antibody specifically stained MFIBs. Freeze-substitution electron microscopy demonstrated that cells that retained MFIBs had electron-dense filamentous structures with a diameter of 10 nm in straight elongated mitochondria. Immunoelectron microscopy showed that Ald4p was localized to the electron-dense filamentous structures in mitochondria. These results together showed that a major component of MFIBs is Ald4p. In addition, we demonstrate that MFIBs are common features that appear in mitochondria of many species of yeast. PMID:24771619

  2. Aldehyde dehydrogenase 2 activation in aged heart improves the autophagy by reducing the carbonyl modification on SIRT1.

    PubMed

    Wu, Bing; Yu, Lu; Wang, Yishi; Wang, Hongtao; Li, Chen; Yin, Yue; Yang, Jingrun; Wang, Zhifa; Zheng, Qiangsun; Ma, Heng

    2016-01-19

    Cardiac aging is characterized by accumulation of damaged proteins and decline of autophagic efficiency. Here, by forestalling SIRT1 carbonylated inactivation in aged heart, we determined the benefits of activation of aldehyde dehydrogenase 2 (ALDH2) on the autophagy. In this study, the ALDH2 KO mice progressively developed age-related heart dysfunction and showed reduction in the life span, which strongly suggests that ALDH2 ablation leads to cardiac aging. What's more, aged hearts displayed a significant decrease ALDH2 activity, resulting in accumulation of 4-HNE-protein adducts and protein carbonyls, impairment in the autophagy flux, and, consequently, deteriorated cardiac function after starvation. Sustained Alda-1 (selective ALDH2 activator) treatment increased cardiac ALDH2 activity and abrogated these effects. Using SIRT1 deficient heterozygous (Sirt1+/-) mice, we found that SIRT1 was necessary for ALDH2 activation-induced autophagy. We further demonstrated that ALDH2 activation attenuated SIRT1 carbonylation and improved SIRT1 activity, thereby increasing the deacetylation of nuclear LC3 and FoxO1. Sequentially, ALDH2 enhanced SIRT1 regulates LC3-Atg7 interaction and FoxO1 increased Rab7 expression, which were both necessary and sufficient for restoring autophagy flux. These results highlight that both accumulation of proteotoxic carbonyl stress linkage with autophagy decline contribute to heart senescence. ALDH2 activation is adequate to improve the autophagy flux by reducing the carbonyl modification on SIRT1, which in turn plays an important role in maintaining cardiac health during aging.

  3. Deficient Expression of Aldehyde Dehydrogenase 1A1 is Consistent With Increased Sensitivity of Gorlin Syndrome Patients to Radiation Carcinogenesis

    PubMed Central

    Wright, Aaron T.; Magnaldo, Thierry; Sontag, Ryan L.; Anderson, Lindsey N.; Sadler, Natalie C.; Piehowski, Paul D.; Gache, Yannick; Weber, Thomas J.

    2016-01-01

    Human phenotypes that are highly susceptible to radiation carcinogenesis have been identified. Sensitive phenotypes often display robust regulation of molecular features that modify biological response, which can facilitate identification of the pathways/networks that contribute to pathophysiological outcomes. Here we interrogate primary dermal fibroblasts isolated from Gorlin syndrome patients (GDFs), who display a pronounced inducible tumorigenic response to radiation, in comparison to normal human dermal fibroblasts (NHDFs). Our approach exploits newly developed thiol reactive probes to define changes in protein thiol profiles in live cell studies, which minimizes artifacts associated with cell lysis. Redox probes revealed deficient expression of an apparent 55 kDa protein thiol in GDFs from independent Gorlin syndrome patients, compared with NHDFs. Proteomics tentatively identified this protein as aldehyde dehydrogenase 1A1 (ALDH1A1), a key enzyme regulating retinoic acid synthesis, and ALDH1A1 protein deficiency in GDFs was confirmed by Western blot. A number of additional protein thiol differences in GDFs were identified, including radiation responsive annexin family members and lamin A/C. Collectively, candidates identified in our study have plausible implications for radiation health effects and cancer susceptibility. PMID:24285572

  4. Impairment of aldehyde dehydrogenase 2 increases accumulation of acetaldehyde-derived DNA damage in the esophagus after ethanol ingestion

    PubMed Central

    Yukawa, Yoshiyuki; Ohashi, Shinya; Amanuma, Yusuke; Nakai, Yukie; Tsurumaki, Mihoko; Kikuchi, Osamu; Miyamoto, Shin’ichi; Oyama, Tsunehiro; Kawamoto, Toshihiro; Chiba, Tsutomu; Matsuda, Tomonari; Muto, Manabu

    2014-01-01

    Ethanol and its metabolite, acetaldehyde, are the definite carcinogens for esophageal squamous cell carcinoma (ESCC), and reduced catalytic activity of aldehyde dehydrogenase 2 (ALDH2), which detoxifies acetaldehyde, increases the risk for ESCC. However, it remains unknown whether the ALDH2 genotype influences the level of acetaldehyde-derived DNA damage in the esophagus after ethanol ingestion. In the present study, we administered ethanol orally or intraperitoneally to Aldh2-knockout and control mice, and we quantified the level of acetaldehyde-derived DNA damage, especially N2-ethylidene-2’-deoxyguanosine (N2-ethylidene-dG), in the esophagus. In the model of oral ethanol administration, the esophageal N2-ethylidene-dG level was significantly higher in Aldh2-knockout mice compared with control mice. Similarly, in the model of intraperitoneal ethanol administration, in which the esophagus is not exposed directly to the alcohol solution, the esophageal N2-ethylidene-dG level was also elevated in Aldh2-knockout mice. This result indicates that circulating ethanol-derived acetaldehyde causes esophageal DNA damage, and that the extent of damage is influenced by knockout of Aldh2. Taken together, our findings strongly suggest the importance of acetaldehyde-derived DNA damage which is induced in the esophagus of individuals with ALDH2 gene impairment. This provides a physiological basis for understanding alcohol-related esophageal carcinogenesis. PMID:24959382

  5. Deficient expression of aldehyde dehydrogenase 1A1 is consistent with increased sensitivity of Gorlin syndrome patients to radiation carcinogenesis.

    PubMed

    Wright, Aaron T; Magnaldo, Thierry; Sontag, Ryan L; Anderson, Lindsey N; Sadler, Natalie C; Piehowski, Paul D; Gache, Yannick; Weber, Thomas J

    2015-06-01

    Human phenotypes that are highly susceptible to radiation carcinogenesis have been identified. Sensitive phenotypes often display robust regulation of molecular features that modify biological response, which can facilitate identification of the pathways/networks that contribute to pathophysiological outcomes. Here we interrogate primary dermal fibroblasts isolated from Gorlin syndrome patients (GDFs), who display a pronounced inducible tumorigenic response to radiation, in comparison to normal human dermal fibroblasts (NHDFs). Our approach exploits newly developed thiol reactive probes to define changes in protein thiol profiles in live cell studies, which minimizes artifacts associated with cell lysis. Redox probes revealed deficient expression of an apparent 55 kDa protein thiol in GDFs from independent Gorlin syndrome patients, compared with NHDFs. Proteomics tentatively identified this protein as aldehyde dehydrogenase 1A1 (ALDH1A1), a key enzyme regulating retinoic acid synthesis, and ALDH1A1 protein deficiency in GDFs was confirmed by Western blot. A number of additional protein thiol differences in GDFs were identified, including radiation responsive annexin family members and lamin A/C. Collectively, candidates identified in our study have plausible implications for radiation health effects and cancer susceptibility.

  6. Implication of an Aldehyde Dehydrogenase Gene and a Phosphinothricin N-Acetyltransferase Gene in the Diversity of Pseudomonas cichorii Virulence

    PubMed Central

    Tanaka, Masayuki; Wali, Ullah Md; Nakayashiki, Hitoshi; Fukuda, Tatsuya; Mizumoto, Hiroyuki; Ohnishi, Kouhei; Kiba, Akinori; Hikichi, Yasufumi

    2011-01-01

    Pseudomonas cichorii harbors the hrp genes. hrp-mutants lose their virulence on eggplant but not on lettuce. A phosphinothricin N-acetyltransferase gene (pat) is located between hrpL and an aldehyde dehydrogenase gene (aldH) in the genome of P. cichorii. Comparison of nucleotide sequences and composition of the genes among pseudomonads suggests a common ancestor of hrp and pat between P. cichorii strains and P. viridiflava strains harboring the single hrp pathogenicity island. In contrast, phylogenetic diversification of aldH corresponded to species diversification amongst pseudomonads. In this study, the involvement of aldH and pat in P. cichorii virulence was analyzed. An aldH-deleted mutant (ΔaldH) and a pat-deleted mutant (Δpat) lost their virulence on eggplant but not on lettuce. P. cichorii expressed both genes in eggplant leaves, independent of HrpL, the transcriptional activator for the hrp. Inoculation into Asteraceae species susceptible to P. cichorii showed that the involvement of hrp, pat and aldH in P. cichorii virulence is independent of each other and has no relationship with the phylogeny of Asteraceae species based on the nucleotide sequences of ndhF and rbcL. It is thus thought that not only the hrp genes but also pat and aldH are implicated in the diversity of P. cichorii virulence on susceptible host plant species. PMID:24704843

  7. The Short-Chain Alcohol Dehydrogenase ABA2 Catalyzes the Conversion of Xanthoxin to Abscisic AldehydeW⃞

    PubMed Central

    González-Guzmán, Miguel; Apostolova, Nadezda; Bellés, José M.; Barrero, José M.; Piqueras, Pedro; Ponce, María R.; Micol, José L.; Serrano, Ramón; Rodríguez, Pedro L.

    2002-01-01

    Mutants able to germinate and perform early growth in medium containing a high NaCl concentration were identified during the course of two independent screenings and named salt resistant (sre) and salobreño (sañ). The sre and sañ mutants also were able to germinate in high-osmoticum medium, indicating that they are osmotolerant in a germination assay. Complementation analyses revealed that sre1-1, sre1-2, sañ3-1, and sañ3-2 were alleles of the abscisic acid (ABA) biosynthesis ABA2 gene. A map-based cloning strategy allowed the identification of the ABA2 gene and molecular characterization of four new aba2 alleles. The ABA2 gene product belongs to the family of short-chain dehydrogenases/reductases, which are known to be NAD- or NADP-dependent oxidoreductases. Recombinant ABA2 protein produced in Escherichia coli exhibits a Km value for xanthoxin of 19 μM and catalyzes in a NAD-dependent manner the conversion of xanthoxin to abscisic aldehyde, as determined by HPLC–mass spectrometry. The ABA2 mRNA is expressed constitutively in all plant organs examined and is not upregulated in response to osmotic stress. The results of this work are discussed in the context of previous genetic and biochemical evidence regarding ABA biosynthesis, confirming the xanthoxin→abscisic aldehyde→ABA transition as the last steps of the major ABA biosynthetic pathway. PMID:12172025

  8. Folate, alcohol, and aldehyde dehydrogenase 2 polymorphism and the risk of oral and pharyngeal cancer in Japanese.

    PubMed

    Matsuo, Keitaro; Rossi, Marta; Negri, Eva; Oze, Isao; Hosono, Satoyo; Ito, Hidemi; Watanabe, Miki; Yatabe, Yasushi; Hasegawa, Yasuhisa; Tanaka, Hideo; Tajima, Kazuo; La Vecchia, Carlo

    2012-03-01

    Folate consumption is inversely associated with the risk of oral and pharyngeal cancer (OPC) and potentially interacts with alcohol drinking in the risk of OPC. Aldehyde dehydrogenase 2 (ALDH2) gene polymorphism is known to interact with alcohol consumption. The aim of this study was to investigate potential interaction between folate, alcohol drinking, and ALDH2 polymorphism in the risk of OPC in a Japanese population. The study group comprised 409 head and neck cancer cases and 1227 age-matched and sex-matched noncancer controls; of these, 251 cases and 759 controls were evaluated for ALDH rs671 polymorphism. Associations were assessed by odds ratios and 95% confidence intervals in multiple logistic regression models. We observed an inverse association between folate consumption and OPC risk. The odds ratio for high folate intake was 0.53 (95% confidence interval: 0.36-0.77) relative to low intake (P trend=0.003). This association was consistent across strata of sex, age, smoking, and ALDH2 genotypes. Interaction between folate consumption, drinking, and ALDH2 genotype was remarkable (three-way interaction, P<0.001). We observed significant interaction among folate, drinking, and ALDH2 genotype in the Japanese population.

  9. Substrate specificity and inhibitor sensitivity of rabbit 20α-hydroxysteroid dehydrogenase.

    PubMed

    Endo, Satoshi; Arai, Yuki; Hara, Akira; Kitade, Yukio; Bunai, Yasuo; El-Kabbani, Ossama; Matsunaga, Toshiyuki

    2013-01-01

    In this study, we examined the substrate specificity and inhibitor sensitivity of rabbit 20α-hydroxysteroid dehydrogenase (AKR1C5), which plays a role in the termination of pregnancy by progesterone inactivation. AKR1C5 moderately reduced the 3-keto group of only 5α-dihydrosteroids with 17β- or 20α/β-hydroxy group among 3-ketosteroids. In contrast, the enzyme reversibly and efficiently catalyzed the reduction of various 17- and 20-ketosteroids, including estrogen precursors (dehydroepiandrosterone, estrone and 5α-androstan-3β-ol-17-one) and tocolytic 5β-pregnane-3,20-dione. In addition to the progesterone inactivation, the formation of estrogens and metabolism of the tocolytic steroid by AKR1C5 may be related to its role in rabbit parturition. AKR1C5 also reduced various non-steroidal carbonyl compounds, including isatin, an antagonist of the C-type natriuretic peptide receptor, and 4-oxo-2-nonenal, suggesting its roles in controlling the bioactive isatin and detoxification of cytotoxic aldehydes. AKR1C5 was potently and competitively inhibited by flavonoids such as kaempferol and quercetin, suggesting that its activity is affected by ingested flavonoids.

  10. Virtual fragment screening for novel inhibitors of 6-phosphogluconate dehydrogenase.

    PubMed

    Ruda, Gian Filippo; Campbell, Gordon; Alibu, Vincent P; Barrett, Michael P; Brenk, Ruth; Gilbert, Ian H

    2010-07-15

    The enzyme 6-phosphogluconate dehydrogenase is a potential drug target for the parasitic protozoan Trypanosoma brucei, the causative organism of human African trypanosomiasis. This enzyme has a polar active site to accommodate the phosphate, hydroxyl and carboxylate groups of the substrate, 6-phosphogluconate. A virtual fragment screen was undertaken of the enzyme to discover starting points for the development of inhibitors which are likely to have appropriate physicochemical properties for an orally bioavailable compound. A virtual screening library was developed, consisting of compounds with functional groups that could mimic the phosphate group of the substrate, but which have a higher pKa. Following docking, hits were clustered and appropriate compounds purchased and assayed against the enzyme. Three fragments were identified that had IC50 values in the low micromolar range and good ligand efficiencies. Based on these initial hits, analogues were procured and further active compounds were identified. Some of the fragments identified represent potential starting points for a medicinal chemistry programme to develop potent drug-like inhibitors of the enzyme. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  11. The bifunctional alcohol and aldehyde dehydrogenase gene, adhE, is necessary for ethanol production in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum.

    PubMed

    Lo, Jonathan; Zheng, Tianyong; Hon, Shuen; Olson, Daniel G; Lynd, Lee R

    2015-04-01

    Thermoanaerobacterium saccharolyticum and Clostridium thermocellum are anaerobic thermophilic bacteria being investigated for their ability to produce biofuels from plant biomass. The bifunctional alcohol and aldehyde dehydrogenase gene, adhE, is present in these bacteria and has been known to be important for ethanol formation in other anaerobic alcohol producers. This study explores the inactivation of the adhE gene in C. thermocellum and T. saccharolyticum. Deletion of adhE reduced ethanol production by >95% in both T. saccharolyticum and C. thermocellum, confirming that adhE is necessary for ethanol formation in both organisms. In both adhE deletion strains, fermentation products shifted from ethanol to lactate production and resulted in lower cell density and longer time to reach maximal cell density. In T. saccharolyticum, the adhE deletion strain lost >85% of alcohol dehydrogenase (ADH) activity. Aldehyde dehydrogenase (ALDH) activity did not appear to be affected, although ALDH activity was low in cell extracts. Adding ubiquinone-0 to the ALDH assay increased activity in the T. saccharolyticum parent strain but did not increase activity in the adhE deletion strain, suggesting that ALDH activity was inhibited. In C. thermocellum, the adhE deletion strain lost >90% of ALDH and ADH activity in cell extracts. The C. thermocellum adhE deletion strain contained a point mutation in the lactate dehydrogenase gene, which appears to deregulate its activation by fructose 1,6-bisphosphate, leading to constitutive activation of lactate dehydrogenase. Thermoanaerobacterium saccharolyticum and Clostridium thermocellum are bacteria that have been investigated for their ability to produce biofuels from plant biomass. They have been engineered to produce higher yields of ethanol, yet questions remain about the enzymes responsible for ethanol formation in these bacteria. The genomes of these bacteria encode multiple predicted aldehyde and alcohol dehydrogenases which could be

  12. Alcohol Dehydrogenase-1B (rs1229984) and Aldehyde Dehydrogenase-2 (rs671) Genotypes Are Strong Determinants of the Serum Triglyceride and Cholesterol Levels of Japanese Alcoholic Men

    PubMed Central

    Yokoyama, Akira; Yokoyama, Tetsuji; Matsui, Toshifumi; Mizukami, Takeshi; Kimura, Mitsuru; Matsushita, Sachio; Higuchi, Susumu; Maruyama, Katsuya

    2015-01-01

    Background Elevated serum triglyceride (TG) and high-density-lipoprotein cholesterol (HDL-C) levels are common in drinkers. The fast-metabolizing alcohol dehydrogenase-1B encoded by the ADH1B*2 allele (vs. ADH1B*1/*1 genotype) and inactive aldehyde dehydrogenase-2 encoded by the ALDH2*2 allele (vs. ALDH2*1/*1 genotype) modify ethanol metabolism and are prevalent (≈90% and ≈40%, respectively) in East Asians. We attempted to evaluate the associations between the ADH1B and ALDH2 genotypes and lipid levels in alcoholics. Methods The population consisted of 1806 Japanese alcoholic men (≥40 years) who had undergone ADH1B and ALDH2 genotyping and whose serum TG, total cholesterol, and HDL-C levels in the fasting state had been measured within 3 days after admission. Results High serum levels of TG (≥150 mg/dl), HDL-C (>80 mg/dl), and low-density-lipoprotein cholesterol (LDL-C calculated by the Friedewald formula ≥140 mg/dl) were observed in 24.3%, 16.8%, and 15.6%, respectively, of the subjects. Diabetes, cirrhosis, smoking, and body mass index (BMI) affected the serum lipid levels. Multivariate analysis revealed that the presence of the ADH1B*2 allele and the active ALDH2*1/*1 genotype increased the odds ratio (OR; 95% confidence interval) for a high TG level (2.22 [1.67–2.94] and 1.39 [0.99–1.96], respectively), and decreased the OR for a high HDL-C level (0.37 [0.28–0.49] and 0.51 [0.37–0.69], respectively). The presence of the ADH1B*2 allele decreased the OR for a high LDL-C level (0.60 [0.45–0.80]). The ADH1B*2 plus ALDH2*1/*1 combination yielded the highest ORs for high TG levels and lowest OR for a high HDL-C level. The genotype effects were more prominent in relation to the higher levels of TG (≥220 mg/dl) and HDL-C (≥100 mg/dl). Conclusions The fast-metabolizing ADH1B and active ALDH2, and especially a combination of the two were strongly associated with higher serum TG levels and lower serum HDL-C levels of alcoholics. The fast

  13. Polymorphisms of alcohol dehydrogenase-1B and aldehyde dehydrogenase-2 and the blood and salivary ethanol and acetaldehyde concentrations of Japanese alcoholic men.

    PubMed

    Yokoyama, Akira; Tsutsumi, Eri; Imazeki, Hiromi; Suwa, Yoshihide; Nakamura, Chizu; Yokoyama, Tetsuji

    2010-07-01

    The effects of genetic polymorphism of aldehyde dehydrogenase-2 (ALDH2) on alcohol metabolism are striking in nonalcoholics, and the effects of genetic polymorphism of alcohol dehydrogenase-1B (ADH1B) are modest at most, whereas genetic polymorphisms of both strongly affect the susceptibility to alcoholism and upper aerodigestive tract (UADT) cancer of drinkers. We evaluated associations between ADH1B/ADH1C/ALDH2 genotypes and the blood and salivary ethanol and acetaldehyde levels of 168 Japanese alcoholic men who came to our hospital for the first time in the morning and had been drinking until the day before. The ethanol levels in their blood and saliva were similar, but the acetaldehyde levels in their saliva were much higher than in their blood, probably because of acetaldehyde production by oral bacteria. Blood and salivary ethanol and acetaldehyde levels were both significantly higher in the subjects with the less active ADH1B*1/*1 genotype than in the ADH1B*2 carriers, but none of the levels differed according to ALDH2 genotype. Significant linkage disequilibrium was detected between the ADH1B and ADH1C genotypes, but ADH1C genotype did not affect the blood or salivary ethanol or acetaldehyde levels. High blood acetaldehyde levels were found even in the active ALDH2*1/*1 alcoholics, which were comparable with the levels of the inactive heterozygous ALDH2*1/*2 alcoholics with less active ADH1B*1/*1. The slope of the increase in blood acetaldehyde level as the blood ethanol level increased was significantly steeper in alcoholics with inactive heterozygous ALDH2*1/*2 plus ADH1B*2 allele than with any other genotype combinations, but the slopes of the increase in salivary acetaldehyde level as the salivary ethanol level increased did not differ between the groups of subjects with any combinations of ALDH2 and ADH1B genotypes. The ADH1B/ALDH2 genotype affected the blood and salivary ethanol and acetaldehyde levels of nonabstinent alcoholics in a different manner

  14. Identification of a suitable and selective inhibitor towards aldehyde oxidase catalyzed reactions.

    PubMed

    Nirogi, Ramakrishna; Kandikere, Vishwottam; Palacharla, Raghava Choudary; Bhyrapuneni, Gopinadh; Kanamarlapudi, Vijaya Bhargava; Ponnamaneni, Ranjith Kumar; Manoharan, Arun Kumar

    2014-03-01

    1. Aldehyde oxidase (AO) is a liver cytosolic molybdoflavoprotein enzyme whose importance in drug metabolism is gaining in the recent. The objective of this work is to find a potent and selective inhibitor for AO activity using phthalazine oxidation as a marker reaction. 2. Among organic solvents tested, it was identified that methanol was not a suitable choice for AO activity even at concentrations less than 0.2% v/v. Acetonitrile and DMSO did not show any effect till 0.5% v/v but thereafter activites tend to decrease. 3. For selectivity, 23 compounds were selected and evaluated for their effects on AO and nine CYP450 enzymes. Among the tested compounds chlorpromazine, estradiol, hydralazine, quetiapine and raloxifene were selected based on their potency of inhibition towards AO activity. 4. Raloxifene was found to be a non-specific inhibitor of all major tested CYP450 enzymes and was excluded as a selective inhibitor for AO. Quetiapine also showed a degree of inhibition towards the major CYP450 tested. Hydralazine used as a specific inhibitor during the past for AO activity demonstrated a stimulation of AO activity at high and low concentrations respectively and the inhibition noted to be time dependent while inhibiting other enzymes like monoamine oxidase. 5. Estradiol showed no inhibition towards the tested CYP450 enzymes and thus proved to be a selective and specific inhibitor for AO activity with an uncompetitive mode of inhibition.

  15. Association of Genetically Determined Aldehyde Dehydrogenase 2 Activity with Diabetic Complications in Relation to Alcohol Consumption in Japanese Patients with Type 2 Diabetes Mellitus: The Fukuoka Diabetes Registry.

    PubMed

    Idewaki, Yasuhiro; Iwase, Masanori; Fujii, Hiroki; Ohkuma, Toshiaki; Ide, Hitoshi; Kaizu, Shinako; Jodai, Tamaki; Kikuchi, Yohei; Hirano, Atsushi; Nakamura, Udai; Kubo, Michiaki; Kitazono, Takanari

    2015-01-01

    Aldehyde dehydrogenase 2 (ALDH2) detoxifies aldehyde produced during ethanol metabolism and oxidative stress. A genetic defect in this enzyme is common in East Asians and determines alcohol consumption behaviors. We investigated the impact of genetically determined ALDH2 activity on diabetic microvascular and macrovascular complications in relation to drinking habits in Japanese patients with type 2 diabetes mellitus. An ALDH2 single-nucleotide polymorphism (rs671) was genotyped in 4,400 patients. Additionally, the relationship of clinical characteristics with ALDH2 activity (ALDH2 *1/*1 active enzyme activity vs. *1/*2 or *2/*2 inactive enzyme activity) and drinking habits (lifetime abstainers vs. former or current drinkers) was investigated cross-sectionally (n = 691 in *1/*1 abstainers, n = 1,315 in abstainers with *2, n = 1,711 in *1/*1 drinkers, n = 683 in drinkers with *2). The multiple logistic regression analysis for diabetic complications was adjusted for age, sex, current smoking habits, leisure-time physical activity, depressive symptoms, diabetes duration, body mass index, hemoglobin A1c, insulin use, high-density lipoprotein cholesterol, systolic blood pressure and renin-angiotensin system inhibitors use. Albuminuria prevalence was significantly lower in the drinkers with *2 than that of other groups (odds ratio [95% confidence interval (CI)]: *1/*1 abstainers as the referent, 0.94 [0.76-1.16] in abstainers with *2, 1.00 [0.80-1.26] in *1/*1 drinkers, 0.71 [0.54-0.93] in drinkers with *2). Retinal photocoagulation prevalence was also lower in drinkers with ALDH2 *2 than that of other groups. In contrast, myocardial infarction was significantly increased in ALDH2 *2 carriers compared with that in ALDH2 *1/*1 abstainers (odds ratio [95% CI]: *1/*1 abstainers as the referent, 2.63 [1.28-6.13] in abstainers with *2, 1.89 [0.89-4.51] in *1/*1 drinkers, 2.35 [1.06-5.79] in drinkers with *2). In summary, patients with type 2 diabetes and ALDH2 *2 displayed a

  16. Association of Genetically Determined Aldehyde Dehydrogenase 2 Activity with Diabetic Complications in Relation to Alcohol Consumption in Japanese Patients with Type 2 Diabetes Mellitus: The Fukuoka Diabetes Registry

    PubMed Central

    Idewaki, Yasuhiro; Iwase, Masanori; Fujii, Hiroki; Ohkuma, Toshiaki; Ide, Hitoshi; Kaizu, Shinako; Jodai, Tamaki; Kikuchi, Yohei; Hirano, Atsushi; Nakamura, Udai; Kubo, Michiaki; Kitazono, Takanari

    2015-01-01

    Aldehyde dehydrogenase 2 (ALDH2) detoxifies aldehyde produced during ethanol metabolism and oxidative stress. A genetic defect in this enzyme is common in East Asians and determines alcohol consumption behaviors. We investigated the impact of genetically determined ALDH2 activity on diabetic microvascular and macrovascular complications in relation to drinking habits in Japanese patients with type 2 diabetes mellitus. An ALDH2 single-nucleotide polymorphism (rs671) was genotyped in 4,400 patients. Additionally, the relationship of clinical characteristics with ALDH2 activity (ALDH2 *1/*1 active enzyme activity vs. *1/*2 or *2/*2 inactive enzyme activity) and drinking habits (lifetime abstainers vs. former or current drinkers) was investigated cross-sectionally (n = 691 in *1/*1 abstainers, n = 1,315 in abstainers with *2, n = 1,711 in *1/*1 drinkers, n = 683 in drinkers with *2). The multiple logistic regression analysis for diabetic complications was adjusted for age, sex, current smoking habits, leisure-time physical activity, depressive symptoms, diabetes duration, body mass index, hemoglobin A1c, insulin use, high-density lipoprotein cholesterol, systolic blood pressure and renin-angiotensin system inhibitors use. Albuminuria prevalence was significantly lower in the drinkers with *2 than that of other groups (odds ratio [95% confidence interval (CI)]: *1/*1 abstainers as the referent, 0.94 [0.76–1.16] in abstainers with *2, 1.00 [0.80–1.26] in *1/*1 drinkers, 0.71 [0.54–0.93] in drinkers with *2). Retinal photocoagulation prevalence was also lower in drinkers with ALDH2 *2 than that of other groups. In contrast, myocardial infarction was significantly increased in ALDH2 *2 carriers compared with that in ALDH2 *1/*1 abstainers (odds ratio [95% CI]: *1/*1 abstainers as the referent, 2.63 [1.28–6.13] in abstainers with *2, 1.89 [0.89–4.51] in *1/*1 drinkers, 2.35 [1.06–5.79] in drinkers with *2). In summary, patients with type 2 diabetes and ALDH2 *2

  17. Fluorescence lifetime analysis and effect of magnesium ions on binding of NADH to human aldehyde dehydrogenase 1.

    PubMed

    Gonnella, Thomas P; Keating, Jennifer M; Kjemhus, Jessica A; Picklo, Matthew J; Biggane, Joseph P

    2013-02-25

    Aldehyde dehydrogenase 1 (ALDH1A1) catalyzes the oxidation of toxic aldehydes to carboxylic acids. Physiologic levels of Mg(2+) ions decrease ALDH1 activity in part by increasing NADH binding affinity to the enzyme. By using time-resolved fluorescence spectroscopy, we have resolved the fluorescent lifetimes (τ) of free NADH in solution (τ=0.4 ns) and two enzyme-bound NADH states (τ=2.0 ns and τ=7.7 ns). We used this technique to investigate the effects of Mg(2+) ions on the ALDH1A1-NADH binding characteristics and enzyme catalysis. From the resolved free and bound NADH fluorescence signatures, the KD values for both NADH conformations in ALDH1A1 ranged from about 24 μM to 1 μM for Mg(2+) ion concentrations of 0-6000 μM, respectively. The rate constants for dissociation of the enzyme-NADH complex ranged from 0.03 s(-1) (6000 μM Mg(2+)) to 0.30s(-1) (0 μM Mg(2+)) as determined by addition of excess NAD(+) to prevent re-association of NADH and resolving the real-time NADH fluorescence signal. During the initial reaction of enzyme with NAD(+) and butyraldehyde, there was an immediate rise in the NADH fluorescence, due to the formation of bound NADH complexes, with a constant steady-state rate of production of free NADH. As the Mg(2+) ion concentration was increased, there was a consistent decrease of the enzyme catalytic turnover from 0.31 s(-1) (0 μM Mg(2+)) to 0.050 s(-1) (6000 μM Mg(2+)) and a distinct shift in steady-state conformational population from one that favors the ALDH1-NADH complex with the shorter fluorescence lifetime (33% excess) in the absence of magnesium ion to one that favors the ALDH1-NADH complex with the longer fluorescence lifetime (13% excess) at 6000 μM Mg(2+). This shift in conformational population at higher Mg(2+) ion concentrations and to lower enzyme activity may be due to longer residence time of the NADH in the ALDH1 pocket. The results from monitoring enzyme catalysis in the absence of magnesium suggests that the ALDH1

  18. Sequence analysis of the oxidase/reductase genes upstream of the Rhodococcus erythropolis aldehyde dehydrogenase gene thcA reveals a gene organisation different from Mycobacterium tuberculosis.

    PubMed

    Nagy, I; De Mot, R

    1999-01-01

    The sequence of the DNA region upstream of the thiocarbamate-inducible aldehyde dehydrogenase gene thcA of Rhodococcus erythropolis NI86/21 was determined. Most of the predicted ORFs are related to various oxidases/reductases, including short-chain oxidases/reductases, GMC oxidoreductases, alpha-hydroxy acid oxidases (subfamily 1 flavin oxidases/dehydrogenases), and subfamily 2 flavin oxidases/dehydrogenases. One ORF is related to enzymes involved in biosynthesis of PQQ or molybdopterin cofactors. In addition, a putative member of the TetR family of regulatory proteins was identified. The substantial sequence divergence from functionally characterized enzymes precludes a reliable prediction about the probable function of these proteins at this stage. In Mycobacterium tuberculosis H37Rv, most of these ORFs have homologs that are also clustered in the genome, but some striking differences in gene organization were observed between Rhodococcus and Mycobacterium.

  19. Aberrant expression of aldehyde dehydrogenase 1A (ALDH1A) subfamily genes in acute lymphoblastic leukaemia is a common feature of T-lineage tumours.

    PubMed

    Longville, Brooke A C; Anderson, Denise; Welch, Mathew D; Kees, Ursula R; Greene, Wayne K

    2015-01-01

    The class 1A aldehyde dehydrogenase (ALDH1A) subfamily of genes encode enzymes that function at the apex of the retinoic acid (RA) signalling pathway. We detected aberrant expression of ALDH1A genes, particularly ALDH1A2, in a majority (72%) of primary paediatric T cell acute lymphoblastic leukaemia (T-ALL) specimens. ALDH1A expression was almost exclusive to T-lineage, but not B-lineage, ALL. To determine whether ALDH1A expression may have relevance to T-ALL cell growth and survival, the effect of inhibiting ALDH1A function was measured on a panel of human ALL cell lines. This revealed that T-ALL proliferation had a higher sensitivity to modulation of ALDH1A activity and RA signalling as compared to ALL cell lines of B-lineage. Consistent with these findings, the genes most highly correlated with ALDH1A2 expression were involved in cell proliferation and apoptosis. Evidence that such genes may be targets of regulation via RA signalling initiated by ALDH1A activity was provided by the TNFRSF10B gene, encoding the apoptotic death receptor TNFRSF10B (also termed TRAIL-R2), which negatively correlated with ALDH1A2 and showed elevated transcription following treatment of T-ALL cell lines with the ALDH1A inhibitor citral (3,7-dimethyl-2,6-octadienal). These data indicate that ALDH1A expression is a common event in T-ALL and supports a role for these enzymes in the pathobiology of this disease.

  20. Association between aldehyde dehydrogenase gene polymorphisms and the phenomenon of field cancerization in patients with head and neck cancer.

    PubMed

    Muto, Manabu; Nakane, Mari; Hitomi, Yoshiaki; Yoshida, Shigeru; Sasaki, Satoshi; Ohtsu, Atsushi; Yoshida, Shigeaki; Ebihara, Satoshi; Esumi, Hiroyasu

    2002-10-01

    Patients with squamous-cell carcinoma in the head and neck (HNSCC) often develop second primary esophageal squamous-cell carcinomas (ESCC). In addition, widespread epithelial oncogenic alterations are also frequently observed in the esophagus and can be made visible as multiple Lugol-voiding lesions (multiple LVL) by Lugol chromoendoscopy. Multiple occurrences of neoplastic change in the upper aerodigestive tract have been explained by the concept of 'field cancerization', usually associated with repeated exposure to carcinogens such as alcohol and cigarette smoke. However, the etiology of second ESCC in HNSCC patients remains unclear and acetaldehyde, the first metabolite of ethanol, has been implicated as the ultimate carcinogen in alcohol-related carcinogenesis. We first investigated the relation between second ESCC and multiple LVL in 78 HNSCC patients. Multiple LVL and second ESCC were observed in 29 (37%) and 21 (27%) patients, respectively. All of the second ESCC were accompanied by multiple LVL. This may indicate that episodes of multiple LVL are precursors for second ESCC. We then examined the association of multiple LVL with the patients' characteristics, including genetic polymorphisms of the alcohol metabolizing enzymes, alcohol dehydrogenase type 3 (ADH3) and aldehyde dehydrogenase type 2 (ALDH2). We also investigated acetaldehyde concentrations in the breath of 52 of the 78 patients. All the patients with multiple LVL were both drinkers and smokers. Multivariable logistic analysis showed that the inactive ALDH2 allele (ALDH2-2) was the strongest contributing factor for the development of multiple LVL (odds ratio 17.6; 95% confidence intervals 4.7-65.3). After alcohol ingestion, acetaldehyde in the breath was elevated to a significantly higher level in all patients with the ALDH2-2 allele than in those without it. The high levels of breath acetaldehyde were significantly modified by the slow-metabolizing ADH3-2 allele. These results reveal strong

  1. Improved Tolerance to Various Abiotic Stresses in Transgenic Sweet Potato (Ipomoea batatas) Expressing Spinach Betaine Aldehyde Dehydrogenase

    PubMed Central

    Fan, Weijuan; Zhang, Min; Zhang, Hongxia; Zhang, Peng

    2012-01-01

    Abiotic stresses are critical delimiters for the increased productivity and cultivation expansion of sweet potato (Ipomoea batatas), a root crop with worldwide importance. The increased production of glycine betaine (GB) improves plant tolerance to various abiotic stresses without strong phenotypic changes, providing a feasible approach to improve stable yield production under unfavorable conditions. The gene encoding betaine aldehyde dehydrogenase (BADH) is involved in the biosynthesis of GB in plants, and the accumulation of GB by the heterologous overexpression of BADH improves abiotic stress tolerance in plants. This study is to improve sweet potato, a GB accumulator, resistant to multiple abiotic stresses by promoted GB biosynthesis. A chloroplastic BADH gene from Spinacia oleracea (SoBADH) was introduced into the sweet potato cultivar Sushu-2 via Agrobacterium-mediated transformation. The overexpression of SoBADH in the transgenic sweet potato improved tolerance to various abiotic stresses, including salt, oxidative stress, and low temperature. The increased BADH activity and GB accumulation in the transgenic plant lines under normal and multiple environmental stresses resulted in increased protection against cell damage through the maintenance of cell membrane integrity, stronger photosynthetic activity, reduced reactive oxygen species (ROS) production, and induction or activation of ROS scavenging by the increased activity of free radical-scavenging enzymes. The increased proline accumulation and systemic upregulation of many ROS-scavenging genes in stress-treated transgenic plants also indicated that GB accumulation might stimulate the ROS-scavenging system and proline biosynthesis via an integrative mechanism. This study demonstrates that the enhancement of GB biosynthesis in sweet potato is an effective and feasible approach to improve its tolerance to multiple abiotic stresses without causing phenotypic defects. This strategy for trait improvement in

  2. Human liver mitochondrial aldehyde dehydrogenase: three-dimensional structure and the restoration of solubility and activity of chimeric forms.

    PubMed Central

    Ni, L.; Zhou, J.; Hurley, T. D.; Weiner, H.

    1999-01-01

    Human liver cytosolic and mitochondrial isozymes of aldehyde dehydrogenase share 70% sequence identity. However, the first 21 residues are not conserved between the human isozymes (15% identity). The three-dimensional structures of the beef mitochondrial and sheep cytosolic forms have virtually identical three-dimensional structures. Here, we solved the structure of the human mitochondrial enzyme and found it to be identical to the beef enzyme. The first 21 residues are found on the surface of the enzyme and make no contact with other subunits in the tetramer. A pair of chimeric enzymes between the human isozymes was made. Each chimera had the first 21 residues from one isozyme and the remaining 479 from the other. When the first 21 residues were from the mitochondrial isozyme, an enzyme with cytosolic-like properties was produced. The other was expressed but was insoluble. It was possible to restore solubility and activity to the chimera that had the first 21 cytosolic residues fused to the mitochondrial ones by making point mutations to residues at the N-terminal end. When residue 19 was changed from tyrosine to a cysteine, the residue found in the mitochondrial form, an active enzyme could be made though the Km for NAD+ was 35 times higher than the native mitochondrial isozyme and the specific activity was reduced by 75%. This residue interacts with residue 203, a nonconserved, nonactive site residue. A mutation of residue 18, which also interacts with 203, restored solubility, but not activity. Mutation to residue 15, which interacts with 104, also restored solubility but not activity. It appears that to have a soluble or active enzyme a favorable interaction must occur between a residue in a surface loop and a residue elsewhere in the molecule even though neither make contact with the active site region of the enzyme. PMID:10631996

  3. Multiple cancers associated with esophageal and oropharyngolaryngeal squamous cell carcinoma and the aldehyde dehydrogenase-2 genotype in male Japanese drinkers.

    PubMed

    Yokoyama, Akira; Watanabe, Hiroshi; Fukuda, Haruhiko; Haneda, Tatsumasa; Kato, Hoichi; Yokoyama, Tetsuji; Muramatsu, Taro; Igaki, Hiroyasu; Tachimori, Yuji

    2002-09-01

    Aldehyde dehydrogenase-2 (ALDH2) is a key enzyme for the elimination of acetaldehyde, an established animal carcinogen generated by alcohol metabolism. In the presence of ALDH2*2, a mutant allele that is prevalent in East Asians, this enzyme is inactive, leading to excessive accumulation of acetaldehyde. Only among Japanese alcoholic patients has the positive association between this inactive form of ALDH2 and multiple-field cancerization in the upper aerodigestive tract been demonstrated. Whether this finding could be extended to multiple-cancer patients in general is of great interest, because the prevalence of esophageal cancer with other organ cancers has increased dramatically during recent decades in Japan. This study compared the ALDH2 genotypes of groups of male Japanese drinkers who had either esophageal squamous cell carcinomas (SCCs) with (n = 26) or without (n = 48) multiplicity or oropharyngolaryngeal SCCs with (n = 17) or without (n = 29) multiplicity. After adjustments for age and drinking and smoking habits, logistic regression analysis showed significantly increased risk for each multiplicity associated with either esophageal or oropharyngolaryngeal SCCs in the presence of the ALDH2*2 allele (odds ratio, 5.26; 95% confidence interval, 1.08-51.06 and odds ratio, 7.36; 95% confidence interval, 1.29-80.70, respectively). This study is the first to strongly link inactive ALDH2 with the multiple cancer susceptibility of male Japanese drinkers with either esophageal or oropharyngolaryngeal cancers. A simple questionnaire about both current and past facial flushing after drinking a glass of beer was highly sensitive (95.6%) in detecting inactive ALDH2 in these patients and may be useful for identifying high-risk patients.

  4. Aldehyde dehydrogenase 2 ameliorates acute cardiac toxicity of ethanol: role of protein phosphatase and forkhead transcription factor.

    PubMed

    Ma, Heng; Li, Ji; Gao, Feng; Ren, Jun

    2009-12-01

    This study was designed to evaluate the role of facilitated detoxification of acetaldehyde, the main metabolic product of ethanol, through systemic overexpression of mitochondrial aldehyde dehydrogenase-2 (ALDH2) on acute ethanol exposure-induced myocardial damage. Binge drinking may exert cardiac toxicity and interfere with heart function, manifested as impaired ventricular contractility, although the underlying mechanism remains poorly defined. ALDH2 transgenic mice were produced using the chicken beta-actin promoter. Wild-type FVB (friend virus B) and ALDH2 mice were challenged with ethanol (3 g/kg, intraperitoneally), and cardiac function was assessed 24 h later using the Langendroff and cardiomyocyte edge-detection systems. Western blot analysis was used to evaluate protein phosphatase 2A and 2C (PP2A and PP2C), phosphorylation of Akt, AMP-activated protein kinase (AMPK), and the transcription factors Foxo3 (Thr32 and Ser413). ALDH2 reduced ethanol-induced elevation in cardiac acetaldehyde levels. Acute ethanol challenge deteriorated myocardial and cardiomyocyte contractile function evidenced by reduction in maximal velocity of pressure development and decline (+/-dP/dt), left ventricular developed pressure, cell shortening, and prolonged relengthening duration, the effects of which were alleviated by ALDH2. Ethanol treatment dampened phosphorylation of Akt and AMPK associated with up-regulated PP2A and PP2C, which was abrogated by ALDH2. ALDH2 significantly attenuated ethanol-induced decrease in Akt- and AMPK-stimulated phosphorylation of Foxo3 at Thr32 and Ser413, respectively. Consistently, ALDH2 rescued ethanol-induced myocardial apoptosis, protein damage, and mitochondrial membrane potential depolarization. Our results suggest that ALDH2 is cardioprotective against acute ethanol toxicity, possibly through inhibition of protein phosphatases, leading to enhanced Akt and AMPK activation, and subsequently, inhibition of Foxo3, apoptosis, and mitochondrial

  5. Head and Neck Squamous Cell Carcinoma Metabolism Draws on Glutaminolysis, and Stemness Is Specifically Regulated by Glutaminolysis via Aldehyde Dehydrogenase.

    PubMed

    Kamarajan, Pachiyappan; Rajendiran, Thekkelnaycke M; Kinchen, Jason; Bermúdez, Mercedes; Danciu, Theodora; Kapila, Yvonne L

    2017-03-03

    Cancer cells use alternate energetic pathways; however, cancer stem cell (CSC) metabolic energetic pathways are unknown. The purpose of this study was to define the metabolic characteristics of head and neck cancer at different points of its pathogenesis with a focus on its CSC compartment. UPLC-MS/MS-profiling and GC-MS-validation studies of human head and neck cancer tissue, saliva, and plasma were used in conjunction with in vitro and in vivo models to carry out this investigation. We identified metabolite biomarker panels that distinguish head and neck cancer from healthy controls, and confirmed involvement of glutamate and glutaminolysis. Glutaminase, which catalyzes glutamate formation from glutamine, and aldehyde dehydrogenase (ALDH), a stemness marker, were highly expressed in primary and metastatic head and neck cancer tissues, tumorspheres, and CSC versus controls. Exogenous glutamine induced stemness via glutaminase, whereas inhibiting glutaminase suppressed stemness in vitro and tumorigenesis in vivo. Head and neck CSC (CD44(hi)/ALDH(hi)) exhibited higher glutaminase, glutamate, and sphere levels than CD44(lo)/ALDH(lo) cells. Glutaminase drove transcriptional and translational ALDH expression, and glutamine directed even CD44(lo)/ALDH(lo) cells toward stemness. Glutaminolysis regulates tumorigenesis and CSC metabolism via ALDH. These findings indicate that glutamate is an important marker of cancer metabolism whose regulation via glutaminase works in concert with ALDH to mediate cancer stemness. Future analyses of glutaminolytic-ALDH driven mechanisms underlying tumorigenic transitions may help in the development of targeted therapies for head and neck cancer and its CSC compartment.

  6. Heterologous expression of betaine aldehyde dehydrogenase gene from Ammopiptanthus nanus confers high salt and heat tolerance to Escherichia coli.

    PubMed

    Yu, Hao-Qiang; Wang, Ying-Ge; Yong, Tai-Ming; She, Yue-Hui; Fu, Feng-Ling; Li, Wan-Chen

    2014-10-01

    Betaine aldehyde dehydrogenase (BADH) catalyzes the synthesis of glycine betaine, a regulator of osmosis, and therefore BADH is considered to play a significant role in response of plants to abiotic stresses. Here, based on the conserved residues of the deduced amino acid sequences of the homologous BADH genes, we cloned the AnBADH gene from the xerophytic leguminous plant Ammopiptanthus nanus by using reverse transcription PCR and rapid amplification of cDNA ends. The full-length cDNA is 1,868 bp long without intron, and contains an open reading frame of 1512 bp, and 3'- and 5'-untranslated regions of 294 and 62 bp. It encodes a 54.71 kDa protein of 503 amino acids. The deduced amino acid sequence shares high homology, conserved amino acid residues and sequence motifs crucial for the function with the BADHs in other leguminous species. The sequence of the open reading frame was used to construct a prokaryotic expression vector pET32a-AnBADH, and transform Escherichia coli. The transformants expressed the heterologous AnBADH gene under the induction of isopropyl β-D-thiogalactopyranoside, and demonstrated significant enhancement of salt and heat tolerance under the stress conditions of 700 mmol L(-1) NaCl and 55°C high temperature. This result suggests that the AnBADH gene might play a crucial role in adaption of A. nanus to the abiotic stresses, and have the potential to be applied to transgenic operations of commercially important crops for improvement of abiotic tolerance.

  7. Umbilical cord blood-derived aldehyde dehydrogenase-expressing progenitor cells promote recovery from acute ischemic injury.

    PubMed

    Putman, David M; Liu, Kevin Y; Broughton, Heather C; Bell, Gillian I; Hess, David A

    2012-10-01

    Umbilical cord blood (UCB) represents a readily available source of hematopoietic and endothelial precursors at early ontogeny. Understanding the proangiogenic functions of these somatic progenitor subtypes after transplantation is integral to the development of improved cell-based therapies to treat ischemic diseases. We used fluorescence-activated cell sorting to purify a rare (<0.5%) population of UCB cells with high aldehyde dehydrogenase (ALDH(hi) ) activity, a conserved stem/progenitor cell function. ALDH(hi) cells were depleted of mature monocytes and T- and B-lymphocytes and were enriched for early myeloid (CD33) and stem cell-associated (CD34, CD133, and CD117) phenotypes. Although these cells were primarily hematopoietic in origin, UCB ALDH(hi) cells demonstrated a proangiogenic transcription profile and were highly enriched for both multipotent myeloid and endothelial colony-forming cells in vitro. Coculture of ALDH(hi) cells in hanging transwells promoted the survival of human umbilical vein endothelial cells (HUVEC) under growth factor-free and serum-free conditions. On growth factor depleted matrigel, ALDH(hi) cells significantly increased tube-like cord formation by HUVEC. After induction of acute unilateral hind limb ischemia by femoral artery ligation, transplantation of ALDH(hi) cells significantly enhanced the recovery of perfusion in ischemic limbs. Despite transient engraftment in the ischemic hind limb, early recruitment of ALDH(hi) cells into ischemic muscle tissue correlated with increased murine von Willebrand factor blood vessel and CD31+ capillary densities. Thus, UCB ALDH(hi) cells represent a readily available population of proangiogenic progenitors that promote vascular regeneration. This work provides preclinical justification for the development of therapeutic strategies to treat ischemic diseases using UCB-derived ALDH(hi) mixed progenitor cells. Copyright © 2012 AlphaMed Press.

  8. Carboxymethylated liver alcohol dehydrogenase: kinetic and thermodynamic characterization of reactions with substrates and inhibitors.

    PubMed

    Dahl, K H; Dunn, M F

    1984-12-18

    Liver alcohol dehydrogenase (LADH) carboxymethylated at Cys-46 (CMLADH) forms two different ternary complexes with 4-trans-(N,N-dimethylamino)cinnamaldehyde (DACA). The complex with reduced nicotinamide adenine dinucleotide (NADH) is characterized by a 38-nm red shift of the long-wavelength pi, pi* transition to 436 nm, while the complex with oxidized nicotinamide adenine dinucleotide (NAD+) is characterized by a 60-nm red shift to 458 nm. CMLADH also forms a ternary complex with NAD+ and the Z isomer of 4-trans-(N,N-dimethylamino)cinnamaldoxime in which the absorption of the oxime (lambda max = 354 nm) is red shifted 80 nm to 434 nm. Pyrazole and 4-methylpyrazole are weak competitive inhibitors of ligand binding to the substrate site of native LADH. These inhibitors were found to form ternary complexes with CMLADH and NADH which are more stable than the corresponding complexes with the native enzyme. The transient reductions of the aldehydes DACA and p-nitrobenzaldehyde (NBZA) were studied under single-turnover conditions. Carboxymethylation decreases the DACA reduction rate 80-fold and renders the process essentially independent of pH over the region 5-9, whereas this process depends on a pKa of 6.0 in the native enzyme. At pH 7.0, the rate constant for NBZA reduction also is decreased at least 80-fold to a value of 7.7 +/- 0.3 s-1. Since primary kinetic isotope effects are observed when NADH is substituted with (4R)-4-deuterio-NADH (kH/kD = 3.0 for DACA and kH/kD = 2.3 for NBZA), the rate-limiting step for both aldehydes involves hydride transfer. The altered pH dependence is concluded to be due to an increase in the pK value of the zinc-coordinated DACA-alcohol in the ternary complex with NAD+ by more than 3 units. This perturbation is brought about by the close proximity of the negatively charged carboxymethyl carboxylate.

  9. Stimulation of reductive glycerol metabolism by overexpression of an aldehyde dehydrogenase in a recombinant Klebsiella pneumoniae strain defective in the oxidative pathway.

    PubMed

    Luo, Lian Hua; Seo, Jeong-Woo; Oh, Baek-Rock; Seo, Pil-Soo; Heo, Sun-Yeon; Hong, Won-Kyung; Kim, Dae-Hyuk; Kim, Chul Ho

    2011-08-01

    Previously, we constructed a glycerol oxidative pathway-deficient mutant strain of Klebsiella pneumoniae by inactivation of glycerol dehydrogenase (dhaD) to eliminate by-product synthesis during production of 1,3-propanediol (1,3-PD) from glycerol. Although by-product formation was successfully blocked in the resultant strain, the yield of 1,3-PD was not enhanced, probably because dhaD disruption resulted in insufficient regeneration of the cofactor NADH essential for the activity of 1,3-PD oxidoreductase (DhaT). To improve cofactor regeneration, in the present study we overexpressed an NAD(+)-dependent aldehyde dehydrogenase in the recombinant strain. To this end, an aldehyde dehydrogenase AldHk homologous to E. coli AldH but with NAD(+)-dependent propionaldehyde dehydrogenase activity was identified in K. pneumoniae. Functional analysis revealed that the substrate specificity of AldHk embraced various aldehydes including propionaldehyde, and that NAD(+) was preferred over NADP(+) as a cofactor. Overexpression of AldHk in the glycerol oxidative pathway-deficient mutant AK/pVOTHk resulted in a 3.6-fold increase (0.57 g l(-1) to 2.07 g l(-1)) in the production of 3-hydroxypropionic acid (3-HP), and a 1.1-fold enhancement (8.43 g l(-1) to 9.65 g l(-1)) of 1,3-PD synthesis, when glycerol was provided as the carbon source, compared to the levels synthesized by the control strain (AK/pVOT). Batch fermentation using AK/pVOTHk showed a significant increase (to 70%, w/w) in conversion of glycerol to the reductive metabolites, 1,3-PD and 3-HP, with no production of by-products except acetate.

  10. Effect of 15-hydroxyprostaglandin dehydrogenase inhibitor on wound healing.

    PubMed

    Seo, Seung Yong; Han, Song-Iy; Bae, Chun Sik; Cho, Hoon; Lim, Sung Chul

    2015-06-01

    PGE2 is an important mediator of wound healing. It is degraded and inactivated by 15-hydroxyprostaglandin dehydrogenase (15-PGDH). Various growth factors, type IV collagen, TIMP-2 and PGE2 are important mediators of inflammation involving wound healing. Overproduction of TGF-β and suppression of PGE2 are found in excessive wound scarring. If we make the condition downregulating growth factors and upregulating PGE2, the wound will have a positive effect which results in little scar formation after healing. TD88 is a 15-PGDH inhibitor based on thiazolinedione structure. We evaluated the effect of TD88 on wound healing. In 10 guinea pigs (4 control and 6 experimental groups), we made four 1cm diameter-sized circular skin defects on each back. TD88 and vehicle were applicated on the wound twice a day for 4 days in the experimental and control groups, respectively. Tissue samples were harvested for qPCR and histomorphometric analyses on the 2nd and 4th day after treatment. Histomorphometric analysis showed significant reepithelization in the experimental group. qPCR analysis showed significant decrease of PDGF, CTGF and TIMP-2, but significant increase of type IV collagen in the experimental group. Taken together TD88 could be a good effector on wound healing, especially in the aspects of prevention of scarring. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Green tea catechins: inhibitors of glycerol-3-phosphate dehydrogenase.

    PubMed

    Kao, Chung-Cheng; Wu, Bo-Tsung; Tsuei, Yi-Wei; Shih, Li-Jane; Kuo, Yu-Liang; Kao, Yung-Hsi

    2010-05-01

    Green tea catechins, especially (-)-epigallocatechin-3-gallate (EGCG), are known to regulate obesity and fat accumulation. We performed a kinetic analysis in a cell-free system to determine the mode of inhibition of glycerol-3-phosphate dehydrogenase (GPDH; EC 1.1.1.8) by EGCG. GPDH catalyzes the beta-nicotinamide adenine dinucleotide (NADH)-dependent reduction of dihydroxyacetone phosphate (DHAP) to yield glycerol-3-phosphate, which serves as one of the major precursors of triacylglycerols. We found that EGCG dose-dependently inhibited GPDH activity at a concentration of approximately 20 muM for 50 % inhibition. The IC (50) values of other green tea catechins, such as (-)-epicatechin, (-)-epicatechin-3-gallate, and (-)-epigallocatechin, were all above 100 microM. This suggests a catechin type-dependent effect. Based on double-reciprocal plots of the kinetic data, EGCG was a noncompetitive inhibitor of the GPDH substrates, NADH and DHAP, with respective inhibition constants (Ki) of 18 and 31 microM. Results of this study possibly support previous studies that EGCG mediates fat content. Georg Thieme Verlag KG Stuttgart. New York.

  12. Metabolism of the novel IMP dehydrogenase inhibitor benzamide riboside.

    PubMed

    Jäger, Walter; Salamon, Alexandra; Szekeres, Thomas

    2002-04-01

    Benzamide riboside (BR) is a novel anticancer agent exhibiting pronounced activity against several human tumor cell lines via the inhibition of inosine 5'-monophosphate dehydrogenase (IMPDH) that catalyzes the formation of xanthine 5'-monophosphate from inosine 5'-monophosphate and nicotinamide adenine dinucleotide, thereby restricting the biosynthesis of guanylates. Phosphorylation of BR to its 5'-monophosphate derivative appears to be ubiquitous in most cells catalyzed by the enzymes, adenosine kinase, nicotinamide nucleoside kinase and 5' nucleotidase. BR 5'-monophosphate is then converted to the active metabolite benzamide adenine dinucleotide (BAD) by NMN adenylyltransferase, the rate-limiting enzyme in the biosynthesis of NAD. As BAD is more potent in the inhibition of IMPDH than BR and BR 5'-monophosphate, cytotoxicity of BR is closely connected with intercellular metabolism to BAD. However, intracellular BAD level is also affected by BADase activity, a phosphodiesterase which hydrolyzes BAD to BR-5'-monophosphate and AMP. A recent study demonstrates enzymatic deamination of BR to non-cytotoxic benzene carboxylic acid (BR-COOH) as the main hepatic BR biotransformation product in rat liver. As the IMPDH inhibitors tiazofurin and ribavirin exhibit predominant accumulation and biotransformation in liver, hepatic metabolism may be an important factor also for BR activation and inactivation and should be considered in human liver during cancer therapy when BR is used as a single drug or in combination with other anticancer agents.

  13. Quinazolinethiones and quinazolinediones, novel inhibitors of inosine monophosphate dehydrogenase: synthesis and initial structure-activity relationships.

    PubMed

    Buckley, George M; Davies, Natasha; Dyke, Hazel J; Gilbert, Philip J; Hannah, Duncan R; Haughan, Alan F; Hunt, Caroline A; Pitt, William R; Profit, Rachael H; Ray, Nicholas C; Richard, Marianna D; Sharpe, Andrew; Taylor, Alicia J; Whitworth, Justine M; Williams, Sophie C

    2005-02-01

    The development of a series of novel quinazolinethiones and quinazolinediones as inhibitors of inosine monophosphate dehydrogenase (IMPDH) is described. The synthesis, in vitro inhibitory values for IMPDH II and in vitro inhibitory value for PBMC proliferation are discussed.

  14. Structural insights into xenobiotic and inhibitor binding to human aldehyde oxidase.

    PubMed

    Coelho, Catarina; Foti, Alessandro; Hartmann, Tobias; Santos-Silva, Teresa; Leimkühler, Silke; Romão, Maria João

    2015-10-01

    Aldehyde oxidase (AOX) is a xanthine oxidase (XO)-related enzyme with emerging importance due to its role in the metabolism of drugs and xenobiotics. We report the first crystal structures of human AOX1, substrate free (2.6-Å resolution) and in complex with the substrate phthalazine and the inhibitor thioridazine (2.7-Å resolution). Analysis of the protein active site combined with steady-state kinetic studies highlight the unique features, including binding and substrate orientation at the active site, that characterize human AOX1 as an important drug-metabolizing enzyme. Structural analysis of the complex with the noncompetitive inhibitor thioridazine revealed a new, unexpected and fully occupied inhibitor-binding site that is structurally conserved among mammalian AOXs and XO. The new structural insights into the catalytic and inhibition mechanisms of human AOX that we now report will be of great value for the rational analysis of clinical drug interactions involving inhibition of AOX1 and for the prediction and design of AOX-stable putative drugs.

  15. The oxidative fermentation of ethanol in Gluconacetobacter diazotrophicus is a two-step pathway catalyzed by a single enzyme: alcohol-aldehyde Dehydrogenase (ADHa).

    PubMed

    Gómez-Manzo, Saúl; Escamilla, José E; González-Valdez, Abigail; López-Velázquez, Gabriel; Vanoye-Carlo, América; Marcial-Quino, Jaime; de la Mora-de la Mora, Ignacio; Garcia-Torres, Itzhel; Enríquez-Flores, Sergio; Contreras-Zentella, Martha Lucinda; Arreguín-Espinosa, Roberto; Kroneck, Peter M H; Sosa-Torres, Martha Elena

    2015-01-07

    Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH) and the aldehyde dehydrogenase (ALDH). We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa) of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2-C6) and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde.

  16. The Oxidative Fermentation of Ethanol in Gluconacetobacter diazotrophicus Is a Two-Step Pathway Catalyzed by a Single Enzyme: Alcohol-Aldehyde Dehydrogenase (ADHa)

    PubMed Central

    Gómez-Manzo, Saúl; Escamilla, José E.; González-Valdez, Abigail; López-Velázquez, Gabriel; Vanoye-Carlo, América; Marcial-Quino, Jaime; de la Mora-de la Mora, Ignacio; Garcia-Torres, Itzhel; Enríquez-Flores, Sergio; Contreras-Zentella, Martha Lucinda; Arreguín-Espinosa, Roberto; Kroneck, Peter M. H.; Sosa-Torres, Martha Elena

    2015-01-01

    Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH) and the aldehyde dehydrogenase (ALDH). We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa) of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2–C6) and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde. PMID:25574602

  17. A novel protective mechanism for mitochondrial aldehyde dehydrogenase (ALDH2) in type i diabetes-induced cardiac dysfunction: role of AMPK-regulated autophagy.

    PubMed

    Guo, Yuli; Yu, Wenjun; Sun, Dongdong; Wang, Jiaxing; Li, Congye; Zhang, Rongqing; Babcock, Sara A; Li, Yan; Liu, Min; Ma, Meijuan; Shen, Mingzhi; Zeng, Chao; Li, Na; He, Wei; Zou, Qian; Zhang, Yingmei; Wang, Haichang

    2015-02-01

    Mitochondrial aldehyde dehydrogenase (ALDH2) is known to offer myocardial protection against stress conditions including ischemia-reperfusion injury, alcoholism and diabetes mellitus although the precise mechanism is unclear. This study was designed to evaluate the effect of ALDH2 on diabetes-induced myocardial injury with a focus on autophagy. Wild-type FVB and ALDH2 transgenic mice were challenged with streptozotozin (STZ, 200mg/kg, i.p.) for 3months to induce experimental diabetic cardiomyopathy. Diabetes triggered cardiac remodeling and contractile dysfunction as evidenced by cardiac hypertrophy, decreased cell shortening and prolonged relengthening duration, the effects of which were mitigated by ALDH2. Lectin staining displayed that diabetes promoted cardiac hypertrophy, the effect of which was alleviated by ALDH2. Western blot analysis revealed dampened autophagy protein markers including LC3B ratio and Atg7 along with upregulated p62 following experimental diabetes, the effect of which was reconciled by ALDH2. Phosphorylation level of AMPK was decreased and its downstream signaling molecule FOXO3a was upregulated in both diabetic cardiac tissue and in H9C2 cells with high glucose exposure. All these effect were partly abolished by ALDH2 overexpression and ALDH2 agonist Alda1. High glucose challenge dampened autophagy in H9C2 cells as evidenced by enhanced p62 levels and decreased levels of Atg7 and LC3B, the effect of which was alleviated by the ALDH2 activator Alda-1. High glucose-induced cell death and apoptosis were reversed by Alda-1. The autophagy inhibitor 3-MA and the AMPK inhibitor compound C mitigated Alda-1-offered beneficial effect whereas the autophagy inducer rapamycin mimicked or exacerbated high glucose-induced cell injury. Moreover, compound C nullified Alda-1-induced protection against STZ-induced changes in autophagy and function. Our results suggested that ALDH2 protects against diabetes-induced myocardial dysfunction possibly through an

  18. Expression of the Vibrio cholerae gene encoding aldehyde dehydrogenase is under control of ToxR, the cholera toxin transcriptional activator.

    PubMed Central

    Parsot, C; Mekalanos, J J

    1991-01-01

    The toxR gene of Vibrio cholerae encodes a transcriptional activator required for the expression of the cholera toxin genes (ctxAB) and more than 15 other genes encoding secreted or membrane proteins. The latter group includes virulence genes involved in the biogenesis of the TCP pilus, the accessory colonization factor, and such ToxR-activated genes as tagA, mutations in which cause no detectable virulence defect in the suckling mouse model. To analyze the regulation of expression and the structure of tagA, we have cloned and sequenced about 2 kb of DNA upstream from a tagA::TnphoA fusion. While the portion of the tagA gene product examined presented no extensive similarity to any known protein, the amino acid sequence deduced from an open reading frame (designated aldA) located upstream from and in opposite orientation to tagA was highly similar to the sequences of eukaryotic aldehyde dehydrogenases. An assay of aldehyde dehydrogenase activity in extracts of a wild-type V. cholerae strainand an aldA mutant confirmed that aldA encodes an aldehyde dehydrogenase. Expression of the aldA gene was studied together with that of tagA in both V. cholerae and Escherichia coli. The expression of both tagA and aldA was environmentally regulated and dependent on a functional toxR gene in V. cholerae, but neither promoter was activated by ToxR in E. coli, suggesting that expression of tagA and aldA requires an additional transcriptional activator besides ToxR. The aldA gene is the first example of a gene encoding a cytoplasmic protein that is under the control of ToxR, and this suggests that metabolic enzymes may constitute novel members of virulence regulons in bacteria. Images PMID:1902210

  19. Aldehyde dehydrogenase and estrogen receptor define a hierarchy of cellular differentiation in the normal human mammary epithelium

    PubMed Central

    2014-01-01

    Introduction Although estrogen and progesterone play a key role in normal mammary development and in breast cancer, the potential for proliferation and lineage differentiation as well as origin of cells that express the estrogen receptor (ER) in normal breast epithelium are not known. Some evidence suggests that normal human mammary stem/progenitor cells are ER–, but the identity of these cells and the cellular hierarchy of breast epithelium are still subjects of controversy. It is likely that elucidation of these aspects will bring insight into the cellular origin of breast cancer subtypes. Methods We used fluorescence-activated cell sorting of primary human mammary epithelial cells along with in vitro and in vivo functional assays to examine the hierarchic relation between cells with aldehyde dehydrogenase enzymatic activity (ALDH+ cells) and ER+ cells in the normal human breast epithelium. We assessed the proliferation and lineage differentiation potential of these cells in vitro and in vivo. A gene reporter assay was used to separate live ER+ and ER– mammary epithelial cells. With shRNA-mediated knockdown, we investigated the role of ALDH isoforms in the functionality of mammary epithelial progenitor cells. Results We describe a cellular hierarchy in the normal human mammary gland in which ER–/ALDH+ cells with functional properties of stem/progenitor cells generate ER+ progenitor cells, which in turn give rise to cells of luminal lineage. We show that the ALDH1A1 isoform, through its function in the retinoic acid metabolism, affects the proliferation and/or early differentiation of stem/progenitor cells and is important for branching morphogenesis. Conclusions This study presents direct evidence that ER+ cells are generated by ER–/ALDH+ stem/progenitor cells. We also show that ER+ cells are able to generate cell progeny of luminal lineage in vitro and in vivo. Loss of ALDH1A1 function impairs this process, as well as branching morphogenesis and

  20. Aryl hydrocarbon receptor-associated genes in rat liver: regional coinduction of aldehyde dehydrogenase 3 and glutathione transferase Ya.

    PubMed

    Lindros, K O; Oinonen, T; Kettunen, E; Sippel, H; Muro-Lupori, C; Koivusalo, M

    1998-02-15

    The tumor-associated aldehyde dehydrogenase 3 (ALDH3) and the glutathione transferase (GST)Ya form are coded by members of the Ah (aryl hydrocarbon) battery group of genes activated in the liver by polycyclic hydrocarbons such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The physiological role of the Ah receptor (AHR), its gene-activating mechanism and its endogenous ligands are still poorly clarified. We had previously observed that 3-methylcholanthrene (3MC) and beta-naphthoflavone (betaNF) induced the AHR-associated CYP1A1/1A2 pair in different liver regions, an effect not explained by the acinar distribution of the AHR protein. Here, we investigated AHR-associated regional induction by comparing the expression patterns of ALDH3 and GSTYa. Analysis of samples from periportal and perivenous cell lysates from 3MC-treated animals revealed that ALDH3 mRNA, protein and benzaldehyde-NADP associated activity were all confined to the perivenous region. In contrast, such regio-specific induction was not seen after beta-NF induction. Immunohistochemically, a peculiar mono- or oligocellular induction pattern of ALDH3 was seen, consistently surrounding terminal hepatic veins after 3MC but mainly in the midzonal region after betaNF. A ligand-specific difference in regional induction of GSTYa1 mRNA was also observed: The constitutive perivenous dominance was preserved after 3MC while induction by betaNF was mainly periportal. A 3MC-betaNF difference was also seen by immunohistochemistry and at the GSTYa protein level, in contrast to that of the AHR-unassociated GSTYb protein. However, experiments with hepatocytes isolated from the periportal or perivenous region to replicate these inducer-specific induction responses in vitro were unsuccessful. These data demonstrate that the different acinar induction patterns by 3MC and betaNF previously observed for CYP1A1 and CYP1A2 are seen also for two other Ah battery genes, GSTYa1 and ALDH3, but in a modified, gene-specific form. We

  1. Malignant phyllodes tumors display mesenchymal stem cell features and aldehyde dehydrogenase/disialoganglioside identify their tumor stem cells.

    PubMed

    Lin, Jin-Jin; Huang, Chiun-Sheng; Yu, John; Liao, Guo-Shiou; Lien, Huang-Chun; Hung, Jung-Tung; Lin, Ruey-Jen; Chou, Fen-Pi; Yeh, Kun-Tu; Yu, Alice L

    2014-03-26

    Although breast phyllodes tumors are rare, there is no effective therapy other than surgery. Little is known about their tumor biology. A malignant phyllodes tumor contains heterologous stromal elements, and can transform into rhabdomyosarcoma, liposarcoma and osteosarcoma. These versatile properties prompted us to explore their possible relationship to mesenchymal stem cells (MSCs) and to search for the presence of cancer stem cells (CSCs) in phyllodes tumors. Paraffin sections of malignant phyllodes tumors were examined for various markers by immunohistochemical staining. Xenografts of human primary phyllodes tumors were established by injecting freshly isolated tumor cells into the mammary fat pad of non-obese diabetic-severe combined immunodeficient (NOD-SCID) mice. To search for CSCs, xenografted tumor cells were sorted into various subpopulations by flow cytometry and examined for their in vitro mammosphere forming capacity, in vivo tumorigenicity in NOD-SCID mice and their ability to undergo differentiation. Immunohistochemical analysis revealed the expression of the following 10 markers: CD44, CD29, CD106, CD166, CD105, CD90, disialoganglioside (GD2), CD117, Aldehyde dehydrogenase 1 (ALDH), and Oct-4, and 7 clinically relevant markers (CD10, CD34, p53, p63, Ki-67, Bcl-2, vimentin, and Globo H) in all 51 malignant phyllodes tumors examined, albeit to different extents. Four xenografts were successfully established from human primary phyllodes tumors. In vitro, ALDH+ cells sorted from xenografts displayed approximately 10-fold greater mammosphere-forming capacity than ALDH- cells. GD2+ cells showed a 3.9-fold greater capacity than GD2- cells. ALDH+/GD2+cells displayed 12.8-fold greater mammosphere forming ability than ALDH-/GD2- cells. In vivo, the tumor-initiating frequency of ALDH+/GD2+ cells were up to 33-fold higher than that of ALDH+ cells, with as few as 50 ALDH+/GD2+ cells being sufficient for engraftment. Moreover, we provided the first evidence for

  2. Malignant phyllodes tumors display mesenchymal stem cell features and aldehyde dehydrogenase/disialoganglioside identify their tumor stem cells

    PubMed Central

    2014-01-01

    Introduction Although breast phyllodes tumors are rare, there is no effective therapy other than surgery. Little is known about their tumor biology. A malignant phyllodes tumor contains heterologous stromal elements, and can transform into rhabdomyosarcoma, liposarcoma and osteosarcoma. These versatile properties prompted us to explore their possible relationship to mesenchymal stem cells (MSCs) and to search for the presence of cancer stem cells (CSCs) in phyllodes tumors. Methods Paraffin sections of malignant phyllodes tumors were examined for various markers by immunohistochemical staining. Xenografts of human primary phyllodes tumors were established by injecting freshly isolated tumor cells into the mammary fat pad of non-obese diabetic-severe combined immunodeficient (NOD-SCID) mice. To search for CSCs, xenografted tumor cells were sorted into various subpopulations by flow cytometry and examined for their in vitro mammosphere forming capacity, in vivo tumorigenicity in NOD-SCID mice and their ability to undergo differentiation. Results Immunohistochemical analysis revealed the expression of the following 10 markers: CD44, CD29, CD106, CD166, CD105, CD90, disialoganglioside (GD2), CD117, Aldehyde dehydrogenase 1 (ALDH), and Oct-4, and 7 clinically relevant markers (CD10, CD34, p53, p63, Ki-67, Bcl-2, vimentin, and Globo H) in all 51 malignant phyllodes tumors examined, albeit to different extents. Four xenografts were successfully established from human primary phyllodes tumors. In vitro, ALDH+ cells sorted from xenografts displayed approximately 10-fold greater mammosphere-forming capacity than ALDH- cells. GD2+ cells showed a 3.9-fold greater capacity than GD2- cells. ALDH+/GD2+cells displayed 12.8-fold greater mammosphere forming ability than ALDH-/GD2- cells. In vivo, the tumor-initiating frequency of ALDH+/GD2+ cells were up to 33-fold higher than that of ALDH+ cells, with as few as 50 ALDH+/GD2+ cells being sufficient for engraftment. Moreover, we

  3. Increased Expression of Aldehyde Dehydrogenase 2 Reduces Renal Cell Apoptosis During Ischemia/Reperfusion Injury After Hypothermic Machine Perfusion.

    PubMed

    Zhong, Zibiao; Hu, Qianchao; Fu, Zhen; Wang, Ren; Xiong, Yan; Zhang, Yang; Liu, Zhongzhong; Wang, Yanfeng; Ye, Qifa

    2016-06-01

    Hypothermic machine perfusion (MP) can reduce graft's injury after kidney transplantation; however, the mechanism has not been elucidated. In the past decade, many studies showed that aldehyde dehydrogenase 2 (ALDH2) is a protease which can inhibit cell apoptosis. Therefore, this study aims to explore whether ALDH2 takes part in reducing organ damage after MP. Eighteen healthy male New Zealand rabbits (12 weeks old, weight 3.0 ± 0.3 kg) were randomly divided into three groups: normal group, MP group, and cold storage (CS) group (n = 6). The left kidney of rabbits underwent warm ischemia for 35 min through clamping the left renal pedicle and then reperfusion for 1 h. Left kidneys were preserved by MP or CS (4°C for 4 h) in vivo followed by the right nephrectomy and 24-h reperfusion, and then the specimens and blood were collected. Finally, concentration of urine creatinine (Cr), blood urea nitrogen (BUN), and 4-HNE were tested. Renal apoptosis was detected by TUNEL staining, and the expression of ALDH2, cleaved-caspase 3, bcl-2/ bax, MAPK in renal tissue was detected by immunohistochemistry or Western blot; 24 h after surgery, the concentration of Cr in MP group was 355 ± 71μmol/L, in CS group was 511 ± 44 μmol/L (P < 0.05), while the BUN was 15.02 ± 2.34 mmol/L in MP group, 22.64 ± 3.58 mmol/L in CS group (P < 0.05). The rate of apoptosis and expression of cleaved caspase-3, p-P38, p-ERK, and p-JNK in MP group was significantly lower than that in CS group (P < 0.05), while expression of ALDH2 and bcl-2/bax in MP group was significantly higher than that in CS group (P < 0.05); expression of cleaved caspase-3 in both MP and CS group significantly increased as compared with that in normal group (P < 0.05). In conclusion, increased expression of ALDH2 can reduce the renal cell apoptosis through inhibiting MAPK pathway during ischemia/reperfusion injury (IRI) after hypothermic MP.

  4. Expansion of Umbilical Cord Blood Aldehyde Dehydrogenase Expressing Cells Generates Myeloid Progenitor Cells that Stimulate Limb Revascularization.

    PubMed

    Putman, David M; Cooper, Tyler T; Sherman, Stephen E; Seneviratne, Ayesh K; Hewitt, Mark; Bell, Gillian I; Hess, David A

    2017-07-01

    Uncompromised by chronic disease-related comorbidities, human umbilical cord blood (UCB) progenitor cells with high aldehyde dehydrogenase activity (ALDH(hi) cells) stimulate blood vessel regeneration after intra-muscular transplantation. However, implementation of cellular therapies using UCB ALDH(hi) cells for critical limb ischemia, the most severe form of severe peripheral artery disease, is limited by the rarity (<0.5%) of these cells. Our goal was to generate a clinically-translatable, allogeneic cell population for vessel regenerative therapies, via ex vivo expansion of UCB ALDH(hi) cells without loss of pro-angiogenic potency. Purified UCB ALDH(hi) cells were expanded >18-fold over 6-days under serum-free conditions. Consistent with the concept that ALDH-activity is decreased as progenitor cells differentiate, only 15.1% ± 1.3% of progeny maintained high ALDH-activity after culture. However, compared to fresh UCB cells, expansion increased the total number of ALDH(hi) cells (2.7-fold), CD34(+) /CD133(+) cells (2.8-fold), and hematopoietic colony forming cells (7.7-fold). Remarkably, injection of expanded progeny accelerated recovery of perfusion and improved limb usage in immunodeficient mice with femoral artery ligation-induced limb ischemia. At 7 or 28 days post-transplantation, mice transplanted with expanded ALDH(hi) cells showed augmented endothelial cell proliferation and increased capillary density compared to controls. Expanded cells maintained pro-angiogenic mRNA expression and secreted angiogenesis-associated growth factors, chemokines, and matrix modifying proteins. Coculture with expanded cells augmented human microvascular endothelial cell survival and tubule formation under serum-starved, growth factor-reduced conditions. Expanded UCB-derived ALDH(hi) cells represent an alternative to autologous bone marrow as an accessible source of pro-angiogenic hematopoietic progenitor cells for the refinement of vascular regeneration

  5. Inhibition of xanthine oxidase by the aldehyde oxidase inhibitor raloxifene: implications for identifying molybdopterin nitrite reductases.

    PubMed

    Weidert, E R; Schoenborn, S O; Cantu-Medellin, N; Choughule, K V; Jones, J P; Kelley, E E

    2014-02-15

    Sources of nitric oxide alternative to nitric oxide synthases are gaining significant traction as crucial mediators of vessel function under hypoxic inflammatory conditions. For example, capacity to catalyze the one electron reduction of nitrite (NO2-) to ·NO has been reported for hemoglobin, myoglobin and molybdopterin-containing enzymes including xanthine oxidoreductase (XOR) and aldehyde oxidase (AO). For XOR and AO, use of selective inhibition strategies is therefore crucial when attempting to assign relative contributions to nitrite-mediated ·NO formation in cells and tissue. To this end, XOR inhibition has been accomplished with application of classic pyrazolopyrimidine-based inhibitors allo/oxypurinol or the newly FDA-approved XOR-specific inhibitor, Uloric® (febuxostat). Likewise, raloxifene, an estrogen receptor antagonist, has been identified as a potent (Ki=1.0 nM) inhibitor of AO. Herein, we characterize the inhibition kinetics of raloxifene for XOR and describe the resultant effects on inhibiting XO-catalyzed ·NO formation. Exposure of purified XO to raloxifene (PBS, pH 7.4) resulted in a dose-dependent (12.5-100 μM) inhibition of xanthine oxidation to uric acid. Dixon plot analysis revealed a competitive inhibition process with a Ki=13 μM. This inhibitory process was more effective under acidic pH; similar to values encountered under hypoxic/inflammatory conditions. In addition, raloxifene also inhibited anoxic XO-catalyzed reduction of NO2- to NO (EC50=64 μM). In contrast to having no effect on XO-catalyzed uric acid production, the AO inhibitor menadione demonstrated potent inhibition of XO-catalyzed NO2- reduction (EC50=60 nM); somewhat similar to the XO-specific inhibitor, febuxostat (EC50=4 nM). Importantly, febuxostat was found to be a very poor inhibitor of human AO (EC50=613 μM) suggesting its usefulness for validating XO-dependent contributions to NO2- reduction in biological systems. Combined, these data indicate care should be taken

  6. In vivo ethanol elimination in man, monkey and rat: A lack of relationship between the ethanol metabolism and the hepatic activities of alcohol and aldehyde dehydrogenases

    SciTech Connect

    Zorzano, A. ); Herrera, E. )

    1990-01-01

    The in vivo ethanol elimination in human subjects, monkeys and rats was investigated after an oral ethanol dosage. After 0.4 g. ethanol/kg of body weight, ethanol elimination was much slower in human subjects than in monkeys. In order to detect a rise in monkey plasma ethanol concentrations as early as observed in human subjects, ethanol had to be administered at a dose of 3 g/kg body weight. Ethanol metabolism in rats was also much faster than in human subjects. However, human liver showed higher alcohol dehydrogenase activity and higher low Km aldehyde dehydrogenase activity than rat liver. Thus, our data suggest a lack of relationship between hepatic ethanol-metabolizing activities and the in vivo ethanol elimination rate.

  7. Pepper aldehyde dehydrogenase CaALDH1 interacts with Xanthomonas effector AvrBsT and promotes effector-triggered cell death and defence responses

    PubMed Central

    Kim, Nak Hyun; Hwang, Byung Kook

    2015-01-01

    Xanthomonas type III effector AvrBsT induces hypersensitive cell death and defence responses in pepper (Capsicum annuum) and Nicotiana benthamiana. Little is known about the host factors that interact with AvrBsT. Here, we identified pepper aldehyde dehydrogenase 1 (CaALDH1) as an AvrBsT-interacting protein. Bimolecular fluorescence complementation and co-immunoprecipitation assays confirmed the interaction between CaALDH1 and AvrBsT in planta. CaALDH1:smGFP fluorescence was detected in the cytoplasm. CaALDH1 expression in pepper was rapidly and strongly induced by avirulent Xanthomonas campestris pv. vesicatoria (Xcv) Ds1 (avrBsT) infection. Transient co-expression of CaALDH1 with avrBsT significantly enhanced avrBsT-triggered cell death in N. benthamiana leaves. Aldehyde dehydrogenase activity was higher in leaves transiently expressing CaALDH1, suggesting that CaALDH1 acts as a cell death enhancer, independently of AvrBsT. CaALDH1 silencing disrupted phenolic compound accumulation, H2O2 production, defence response gene expression, and cell death during avirulent Xcv Ds1 (avrBsT) infection. Transgenic Arabidopsis thaliana overexpressing CaALDH1 exhibited enhanced defence response to Pseudomonas syringae pv. tomato and Hyaloperonospora arabidopsidis infection. These results indicate that cytoplasmic CaALDH1 interacts with AvrBsT and promotes plant cell death and defence responses. PMID:25873668

  8. Pepper aldehyde dehydrogenase CaALDH1 interacts with Xanthomonas effector AvrBsT and promotes effector-triggered cell death and defence responses.

    PubMed

    Kim, Nak Hyun; Hwang, Byung Kook

    2015-06-01

    Xanthomonas type III effector AvrBsT induces hypersensitive cell death and defence responses in pepper (Capsicum annuum) and Nicotiana benthamiana. Little is known about the host factors that interact with AvrBsT. Here, we identified pepper aldehyde dehydrogenase 1 (CaALDH1) as an AvrBsT-interacting protein. Bimolecular fluorescence complementation and co-immunoprecipitation assays confirmed the interaction between CaALDH1 and AvrBsT in planta. CaALDH1:smGFP fluorescence was detected in the cytoplasm. CaALDH1 expression in pepper was rapidly and strongly induced by avirulent Xanthomonas campestris pv. vesicatoria (Xcv) Ds1 (avrBsT) infection. Transient co-expression of CaALDH1 with avrBsT significantly enhanced avrBsT-triggered cell death in N. benthamiana leaves. Aldehyde dehydrogenase activity was higher in leaves transiently expressing CaALDH1, suggesting that CaALDH1 acts as a cell death enhancer, independently of AvrBsT. CaALDH1 silencing disrupted phenolic compound accumulation, H2O2 production, defence response gene expression, and cell death during avirulent Xcv Ds1 (avrBsT) infection. Transgenic Arabidopsis thaliana overexpressing CaALDH1 exhibited enhanced defence response to Pseudomonas syringae pv. tomato and Hyaloperonospora arabidopsidis infection. These results indicate that cytoplasmic CaALDH1 interacts with AvrBsT and promotes plant cell death and defence responses.

  9. Detection of activity and mass spectrometric identification of mouse liver carboxylesterase and aldehyde dehydrogenase separated by non-denaturing two-dimensional electrophoresis after extraction with detergents.

    PubMed

    Shimazaki, Youji; Manabe, Takashi

    2005-05-20

    To examine the activities and identity of enzymes associated with organelles such as microsomes and mitochondria, proteins from mouse liver were extracted using the non-ionic detergents Nonidet P-40 (NP-40), polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene isooctylphenyl ester (Triton X), n-octyl beta-D-glucoside (octyl glycoside) or anionic detergent sodium dodecylsulfate (SDS) after the removal of cytosolic proteins. The proteins extracted by detergents were separated by non-denaturing two-dimensional electrophoresis (2-DE). The activities of esterase and aldehyde dehydrogenase were retained by non-denaturing 2-DE after treatment with each non-ionic detergent, but the activities were reduced or lost when the proteins were extracted with more than 0.5% SDS. For proteomic analysis of the organelle-associated proteins in mouse liver, proteins were separated by non-denaturing 2-DE and were identified using electrospray ionization tandem mass spectrometry (ESI-MS/MS) after the proteins were solubilized by octyl glycoside, NP-40 and 0.1% SDS. Several organelle-associated proteins such as carboxylesterase, aldehyde dehydrogenase, glucose regulated protein and HSP60 were identified. These results indicate that the activities and identity of detergent-soluble enzymes can be examined by this non-denaturing 2-DE and mass spectrometry.

  10. The disulfiram metabolites S-methyl-N,N-diethyldithiocarbamoyl sulfoxide and S-methyl-N,N-diethylthiocarbamoyl sulfone irreversibly inactivate betaine aldehyde dehydrogenase from Pseudomonas aeruginosa, both in vitro and in situ, and arrest bacterial growth.

    PubMed

    Zaldívar-Machorro, Víctor J; López-Ortiz, Manuel; Demare, Patricia; Regla, Ignacio; Muñoz-Clares, Rosario A

    2011-02-01

    Betaine aldehyde dehydrogenase from the human opportunistic pathogen Pseudomonas aeruginosa (PaBADH) catalyzes the irreversible, NAD(P)(+)-dependent oxidation of betaine aldehyde, producing glycine betaine, an osmoprotectant. PaBADH participates in the catabolism of choline and likely in the defense against the osmotic and oxidative stresses to which the bacterium is exposed when infecting human tissues. Given that choline or choline precursors are abundant in infected tissues, PaBADH is a potential drug target because its inhibition will lead to the build up of the toxic betaine aldehyde inside bacterial cells. We tested the thiol reagents, disulfiram (DSF) and five DSF metabolites-diethyldithiocarbamic acid (DDC), S-methyl-N,N-diethyldithiocarbamoyl sulfoxide (MeDDTC-SO) and sulfone (MeDDTC-SO(2)), and S-methyl-N,N-diethylthiocarbamoyl sulfoxide (MeDTC-SO) and sulfone (MeDTC-SO(2))-as inhibitors of PaBADH and P. aeruginosa growth. As in vitro PaBADH inhibitors, their order of potency was: MeDDTC-SO(2)>DSF>MeDTC-SO(2)>MeDDTC-SO>MeDTC-SO. DDC did not inactivate the enzyme. PaBADH inactivation by DSF metabolites (i) was not affected by NAD(P)(+), (ii) could not be reverted by dithiothreitol, and (iii) did not affect the quaternary structure of the enzyme. Of the DSF metabolites tested, MeDTC-SO(2) and MeDDTC-SO produced significant in situ PaBADH inactivation and arrest of P. aeruginosa growth in choline containing media, in which the expression of PaBADH is induced. They had no effect in media lacking choline, indicating that PaBADH is their main intracellular target, and that arrest of growth is due to accumulation of betaine aldehyde. The in vitro and in situ kinetics of enzyme inactivation by these two compounds were very similar, indicating no restriction on their uptake by the cells. MeDDTC-SO(2) and DSF have no inhibitory effects in situ, probably because their high reactivity towards intracellular nonessential thiols causes their depletion. Our results

  11. In vivo measurement of aldehyde dehydrogenase-2 activity in rat liver ethanol model using dynamic MRSI of hyperpolarized [1-(13) C]pyruvate.

    PubMed

    Josan, Sonal; Xu, Tao; Yen, Yi-Fen; Hurd, Ralph; Ferreira, Julio; Chen, Che-Hong; Mochly-Rosen, Daria; Pfefferbaum, Adolf; Mayer, Dirk; Spielman, Daniel

    2013-06-01

    To date, measurements of the activity of aldehyde dehydrogenase-2 (ALDH2), a critical mitochondrial enzyme for the elimination of certain cytotoxic aldehydes in the body and a promising target for drug development, have been largely limited to in vitro methods. Recent advancements in MRS of hyperpolarized (13) C-labeled substrates have provided a method to detect and image in vivo metabolic pathways with signal-to-noise ratio gains greater than 10 000-fold over conventional MRS techniques. However aldehydes, because of their toxicity and short T1 relaxation times, are generally poor targets for such (13) C-labeled studies. In this work, we show that dynamic MRSI of hyperpolarized [1-(13) C]pyruvate and its conversion to [1-(13) C]lactate can provide an indirect in vivo measurement of ALDH2 activity via the concentration of NADH (nicotinamide adenine dinucleotide, reduced form), a co-factor common to both the reduction of pyruvate to lactate and the oxidation of acetaldehyde to acetate. Results from a rat liver ethanol model (n = 9) show that changes in (13) C-lactate labeling following the bolus injection of hyperpolarized pyruvate are highly correlated with changes in ALDH2 activity (R(2) = 0.76).

  12. Cynomolgus monkey as a surrogate for human aldehyde oxidase metabolism of the EGFR inhibitor BIBX1382.

    PubMed

    Hutzler, J Matthew; Cerny, Matthew A; Yang, Young-Sun; Asher, Constance; Wong, Diane; Frederick, Kosea; Gilpin, Kyle

    2014-10-01

    BIBX1382 was an epidermal growth factor receptor inhibitor under clinical investigation for treatment of cancer. This candidate possessed an attractive preclinical absorption, distribution, metabolism, and excretion profile, yet failed in clinical studies due in part to poor oral exposure, resulting from extensive metabolism by aldehyde oxidase (AO). In vitro metabolism studies were performed in liver cytosol and cryopreserved hepatocytes from multiple species. In addition, a pharmacokinetic study was performed in cynomolgus monkey for comparison with the reported human pharmacokinetics of BIBX1382. Estimated hepatic clearance of BIBX1382 in rhesus (42 ml/min per kg) and cynomolgus monkey (43 ml/min per kg) liver cytosol was comparable to human (≥93% of liver blood flow). Metabolite identification after incubation of BIBX1382 in liver cytosol fortified with the AO inhibitor raloxifene confirmed that AO is involved in the formation of the predominant metabolite (BIBU1476, M1) in cynomolgus monkey. After intravenous and oral administration of BIBX1382 to cynomolgus monkeys, high plasma clearance (118 ml/min per kg) and low oral exposure (C(max) = 12.7 nM and 6% oral bioavailability) was observed, with the exposure of M1 exceeding BIBX1382 after oral dosing. This pharmacokinetic profile compared favorably with the human clinical data of BIBX1382 (plasma clearance 25-55 ml/min per kg and 5% oral bioavailability). Thus, it appears that cynomolgus monkey represents a suitable surrogate for the observed human AO metabolism of BIBX1382. To circumvent clinical failures due to uncharacterized metabolism by AO, in vitro studies in the appropriate subcellular fraction, followed by pharmacokinetic and toxicokinetic studies in the appropriately characterized surrogate species should be conducted for substrates of AO.

  13. Histamine H4-Receptors Inhibit Mast Cell Renin Release in Ischemia/Reperfusion via Protein Kinase Cε-Dependent Aldehyde Dehydrogenase Type-2 Activation

    PubMed Central

    Aldi, Silvia; Takano, Ken-ichi; Tomita, Kengo; Koda, Kenichiro; Chan, Noel Y.-K.; Marino, Alice; Salazar-Rodriguez, Mariselis; Thurmond, Robin L.

    2014-01-01

    Renin released by ischemia/reperfusion (I/R) from cardiac mast cells (MCs) activates a local renin-angiotensin system (RAS) causing arrhythmic dysfunction. Ischemic preconditioning (IPC) inhibits MC renin release and consequent activation of this local RAS. We postulated that MC histamine H4-receptors (H4Rs), being Gαi/o-coupled, might activate a protein kinase C isotype–ε (PKCε)–aldehyde dehydrogenase type-2 (ALDH2) cascade, ultimately eliminating MC-degranulating and renin-releasing effects of aldehydes formed in I/R and associated arrhythmias. We tested this hypothesis in ex vivo hearts, human mastocytoma cells, and bone marrow–derived MCs from wild-type and H4R knockout mice. We found that activation of MC H4Rs mimics the cardioprotective anti-RAS effects of IPC and that protection depends on the sequential activation of PKCε and ALDH2 in MCs, reducing aldehyde-induced MC degranulation and renin release and alleviating reperfusion arrhythmias. These cardioprotective effects are mimicked by selective H4R agonists and disappear when H4Rs are pharmacologically blocked or genetically deleted. Our results uncover a novel cardioprotective pathway in I/R, whereby activation of H4Rs on the MC membrane, possibly by MC-derived histamine, leads sequentially to PKCε and ALDH2 activation, reduction of toxic aldehyde-induced MC renin release, prevention of RAS activation, reduction of norepinephrine release, and ultimately to alleviation of reperfusion arrhythmias. This newly discovered protective pathway suggests that MC H4Rs may represent a new pharmacologic and therapeutic target for the direct alleviation of RAS-induced cardiac dysfunctions, including ischemic heart disease and congestive heart failure. PMID:24696042

  14. Red Algal Bromophenols as Glucose 6-Phosphate Dehydrogenase Inhibitors

    PubMed Central

    Mikami, Daisuke; Kurihara, Hideyuki; Kim, Sang Moo; Takahashi, Koretaro

    2013-01-01

    Five bromophenols isolated from three Rhodomelaceae algae (Laurencia nipponica, Polysiphonia morrowii, Odonthalia corymbifera) showed inhibitory effects against glucose 6-phosphate dehydrogenase (G6PD). Among them, the symmetric bromophenol dimer (5) showed the highest inhibitory activity against G6PD. PMID:24152564

  15. Structure and mechanism of action of the hydroxy-aryl-aldehyde class of IRE1 endoribonuclease inhibitors

    SciTech Connect

    Sanches, Mario; Duffy, Nicole M.; Talukdar, Manisha; Thevakumaran, Nero; Chiovitti, David; Canny, Marella D.; Lee, Kenneth; Kurinov, Igor; Uehling, David; Al-awar, Rima; Poda, Gennadiy; Prakesch, Michael; Wilson, Brian; Tam, Victor; Schweitzer, Colleen; Toro, Andras; Lucas, Julie L.; Vuga, Danka; Lehmann, Lynn; Durocher, Daniel; Zeng, Qingping; Patterson, John B.; Sicheri, Frank

    2014-10-24

    Endoplasmic reticulum (ER) stress activates the unfolded protein response and its dysfunction is linked to multiple diseases. The stress transducer IRE1α is a transmembrane kinase endoribonuclease (RNase) that cleaves mRNA substrates to re-establish ER homeostasis. Aromatic ring systems containing hydroxy–aldehyde moieties, termed hydroxy–aryl–aldehydes (HAA), selectively inhibit IRE1α RNase and thus represent a novel chemical series for therapeutic development. We solved crystal structures of murine IRE1α in complex with three HAA inhibitors. HAA inhibitors engage a shallow pocket at the RNase-active site through pi-stacking interactions with His910 and Phe889, an essential Schiff base with Lys907 and a hydrogen bond with Tyr892. Structure–activity studies and mutational analysis of contact residues define the optimal chemical space of inhibitors and validate the inhibitor-binding site. These studies lay the foundation for understanding both the biochemical and cellular functions of IRE1α using small molecule inhibitors and suggest new avenues for inhibitor design.

  16. Use of competitive dead-end inhibitors to determine the chemical mechanism of action of yeast alcohol dehydrogenase.

    PubMed

    Leskovac, V; Trivić, S; Anderson, B M

    1998-01-01

    In this work, we have postulated a comprehensive and unified chemical mechanism of action for yeast alcohol dehydrogenase (EC 1.1.1.1, constitutive, cytoplasmic), isolated from Saccharomyces cerevisiae. The chemical mechanism of yeast enzyme is based on the integrity of the proton relay system: His-51....NAD+....Thr-48....R.CH2OH(H2O)....Zn++, stretching from His-51 on the surface of enzyme to the active site zinc atom in the substrate-binding site of enzyme. Further, it is based on extensive studies of steady-state kinetic properties of enzyme which were published recently. In this study, we have reported the pH-dependence of dissociation constants for several competitive dead-end inhibitors of yeast enzyme froin their binary complexes with enzyme, or their ternary complexes with enzyme and NAD+ or NADH; inhibitors include: pyrazole, acetamide, sodium azide, 2-fluoroethanol, and 2,2,2-trifluorethanol. The unified mechanism describes the structures of four dissociation forms of apoenzyme, two forms of the binary complex E.NAD+, three forms of the ternary complex E.NAD+.alcohol, two forms of the ternary complex E.NADH.aldehyde and three binary complexes E.NADH. Appropriate pKa values have been ascribed to protonation forms of most of the above mentioned complexes of yeast enzyme with coenzymes and substrates.

  17. Ergot alkaloid biosynthesis in Aspergillus fumigatus: conversion of chanoclavine-I to chanoclavine-I aldehyde catalyzed by a short-chain alcohol dehydrogenase FgaDH.

    PubMed

    Wallwey, Christiane; Matuschek, Marco; Li, Shu-Ming

    2010-02-01

    Ergot alkaloids are toxins and important pharmaceuticals which are produced biotechnologically on an industrial scale. A putative gene fgaDH has been identified in the biosynthetic gene cluster of fumigaclavine C, an ergot alkaloid of the clavine-type. The deduced gene product FgaDH comprises 261 amino acids with a molecular mass of about 27.8 kDa and contains the conserved motifs of classical short-chain dehydrogenases/reductases (SDRs), but shares no worth mentioning sequence similarity with SDRs and other known proteins. The coding region of fgaDH consisting of two exons was amplified by PCR from a cDNA library of Aspergillus fumigatus, cloned into pQE60 and overexpressed in E. coli. The soluble tetrameric His(6)-FgaDH was purified to apparent homogeneity and characterized biochemically. It has been shown that FgaDH catalyzes the oxidation of chanoclavine-I in the presence of NAD(+) resulting in the formation of chanoclavine-I aldehyde, which was unequivocally identified by NMR and MS analyzes. Therefore, FgaDH functions as a chanoclavine-I dehydrogenase and represents a new group of short-chain dehydrogenases. K (M) values for chanoclavine-I and NAD(+) were determined at 0.27 and 1.1 mM, respectively. The turnover number was 0.38 s(-1).

  18. Aromatase, estrone sulfatase, and 17β-hydroxysteroid dehydrogenase: structure-function studies and inhibitor development.

    PubMed

    Hong, Yanyan; Chen, Shiuan

    2011-07-04

    Aromatase, estrone sulfatase, and 17β-hydroxysteroid dehydrogenase type 1 are involved in the key steps of 17β-estradiol biosynthesis. Structure-function studies of aromatase, estrone sulfatase and 17β-hydroxysteroid dehydrogenase type 1 are important to evaluate the molecular basis of the interaction between these enzymes and their inhibitors. Selective and potent inhibitors of the three enzymes have been developed as antiproliferative agents in hormone-dependent breast carcinoma. New treatment strategies for hormone-dependent breast cancer are discussed.

  19. Benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II from Acinetobacter calcoaceticus. Substrate specificities and inhibition studies.

    PubMed Central

    MacKintosh, R W; Fewson, C A

    1988-01-01

    The apparent Km and maximum velocity values of benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II from Acinetobacter calcoaceticus were determined for a range of alcohols and aldehydes and the corresponding turnover numbers and specificity constants were calculated. Benzyl alcohol was the most effective alcohol substrate for benzyl alcohol dehydrogenase. Perillyl alcohol was the second most effective substrate, and was the only non-aromatic alcohol oxidized. The other substrates of benzyl alcohol dehydrogenase were all aromatic in nature, with para-substituted derivatives of benzyl alcohol being better substrates than other derivatives. Coniferyl alcohol and cinnamyl alcohol were also substrates. Benzaldehyde was much the most effective substrate for benzaldehyde dehydrogenase II. Benzaldehydes with a single small substituent group in the meta or para position were better substrates than any other benzaldehyde derivatives. Benzaldehyde dehydrogenase II could also oxidize the aliphatic aldehydes hexan-1-al and octan-1-al, although poorly. Benzaldehyde dehydrogenase II was substrate-inhibited by benzaldehyde when the assay concentration exceeded approx. 10 microM. Benzaldehyde dehydrogenase II, but not benzyl alcohol dehydrogenase, exhibited esterase activity with 4-nitrophenyl acetate as substrate. Both benzyl alcohol dehydrogenase and benzaldehyde dehydrogenase II were inhibited by the thiol-blocking reagents iodoacetate, iodoacetamide, 4-chloromercuribenzoate and N-ethylmaleimide. Benzyl alcohol or benzaldehyde respectively protected against these inhibitions. NAD+ also gave some protection. Neither benzyl alcohol dehydrogenase nor benzaldehyde dehydrogenase II was inhibited by the metal-ion-chelating agents EDTA, 2,2'-bipyridyl, pyrazole or 2-phenanthroline. Neither enzyme was inhibited by a range of plausible metabolic inhibitors such as mandelate, phenylglyoxylate, benzoate, succinate, acetyl-CoA, ATP or ADP. Benzaldehyde dehydrogenase II was

  20. 3,4-Dihydroxyphenylacetic acid is a potential aldehyde dehydrogenase inducer in murine hepatoma Hepa1c1c7 cells.

    PubMed

    Liu, Yujia; Kurita, Ayuki; Nakashima, Sayaka; Zhu, Beiwei; Munemasa, Shintaro; Nakamura, Toshiyuki; Murata, Yoshiyuki; Nakamura, Yoshimasa

    2017-10-01

    3,4-Dihydroxyphenylacetic acid (DOPAC) is one of the major colonic microflora-produced catabolites of quercetin glycosides, such as quercetin 4'-glucoside derived from onion. Here, we investigated whether DOPAC modulates the aldehyde dehydrogenase (ALDH) activity and protects the cells from the acetaldehyde-induced cytotoxicity in vitro. DOPAC was shown to enhance not only the total ALDH activity, but also the gene expression of ALDH1A1, ALDH2 and ALDH3A1 in a concentration-dependent manner. DOPAC simultaneously stimulated the nuclear translocation of NFE2-related factor 2 and aryl hydrocarbon receptor. The pretreatment of DOPAC completely protected the cells from the acetaldehyde-induced cytotoxicity. The present study suggested that DOPAC acts as a potential ALDH inducer to prevent the alcohol-induced abnormal reaction.

  1. Isolation of human umbilical cord blood aldehyde dehydrogenase-expressing progenitor cells that modulate vascular regenerative functions in vitro and in vivo.

    PubMed

    Putman, David M; Hess, David A

    2013-01-01

    This unit describes the isolation and application of human umbilical cord blood progenitor cells to modulate vascular regenerative functions using in vitro co-culture systems and in vivo transplantation models. Using aldehyde dehydrogenase as a marker of stem cell function, blood-derived progenitors can be efficiently purified form human umbilical cord blood using flow cytometry. We describe in vitro approaches to measure cell-mediated effects on the survival, proliferation, and tube-forming function of endothelial cells using growth-rate assays and Matrigel tube-forming assays. Additionally, we provide a detailed protocol for inducing acute unilateral hindlimb ischemia in immune-deficient mice to assess progenitor cell-modulated effects on vascular regeneration by tracking the recovery of blood flow using noninvasive laser Doppler perfusion imaging. Collectively, we present combined in vitro and in vivo transplantation strategies for the pre-clinical assessment of human progenitor cell-based therapies to treat ischemic disease.

  2. Interaction between osmotic and oxidative stress in diabetic precataractous lens: studies with a sorbitol dehydrogenase inhibitor.

    PubMed

    Obrosova, I G; Fathallah, L; Lang, H J

    1999-12-15

    Both sorbitol accumulation-linked osmotic stress and "pseudohypoxia" [increase in NADH/NAD+, similar to that in hypoxic tissues, and attributed to increased sorbitol dehydrogenase (1-iditol:NAD+ 5-oxidoreductase; EC 1.1.1.14; SDH) activity] have been invoked among the mechanisms underlying oxidative injury in target tissues for diabetic complications. We used the specific SDH inhibitor SDI-157 [2-methyl-4(4-N,N-dimethylaminosulfonyl-1-piperazino)pyrimid ine] to evaluate the role of osmotic stress versus "pseudohypoxia" in oxidative stress occurring in diabetic precataractous lens. Control and diabetic rats were treated with or without SDI-157 (100 mg/kg/day for 3 weeks). Lens malondialdehyde (MDA) plus 4-hydroxyalkenals (4-HA), MDA, GSH, and ascorbate levels, as well as the GSSG/GSH ratios, were similar in SDI-treated and untreated control rats, thus indicating that SDI-157 was not a prooxidant. Intralenticular osmotic stress, manifested by sorbitol levels, was more severe in SDI-treated diabetic rats (38.2+/-6.8 vs 21.2+/-3.5 micromol/g in untreated diabetic and 0.758+/-0.222 micromol/g in control rats, P<0.01 for both), while the decrease in the free cytosolic NAD+/NADH ratio was partially prevented (120+/-16 vs 88+/-11 in untreated diabetic rats and 143+/-13 in controls, P<0.01 for both). GSH and ascorbate levels were decreased, while MDA plus 4-HA and MDA levels were increased in diabetic rats versus controls; both antioxidant depletion and lipid aldehyde accumulation were exacerbated by SDI treatment. Superoxide dismutase (superoxide:superoxide oxidoreductase; EC 1.15.1.1), GSSG reductase (NAD[P]H:oxidized-glutathione oxidoreductase; EC 1.6.4.2), GSH transferase (glutathione S-transferase; EC 2.5.1.18), GSH peroxidase (glutathione:hydrogen-peroxide oxidoreductase; EC 1.11.1.9), and cytoplasmic NADH oxidase activities were increased in diabetic rats versus controls, and all the enzymes but GSH peroxidase were up-regulated further by SDI. In conclusion, sorbitol

  3. Characterization of the Saccharomyces cerevisiae YMR318C (ADH6) gene product as a broad specificity NADPH-dependent alcohol dehydrogenase: relevance in aldehyde reduction.

    PubMed Central

    Larroy, Carol; Fernández, M Rosario; González, Eva; Parés, Xavier; Biosca, Josep A

    2002-01-01

    YMR318C represents an open reading frame from Saccharomyces cerevisiae with unknown function. It possesses a conserved sequence motif, the zinc-containing alcohol dehydrogenase (ADH) signature, specific to the medium-chain zinc-containing ADHs. In the present study, the YMR318C gene product has been purified to homogeneity from overexpressing yeast cells, and found to be a homodimeric ADH, composed of 40 kDa subunits and with a pI of 5.0-5.4. The enzyme was strictly specific for NADPH and was active with a wide variety of substrates, including aliphatic (linear and branched-chain) and aromatic primary alcohols and aldehydes. Aldehydes were processed with a 50-fold higher catalytic efficiency than that for the corresponding alcohols. The highest k(cat)/K(m) values were found with pentanal>veratraldehyde > hexanal > 3-methylbutanal >cinnamaldehyde. Taking into consideration the substrate specificity and sequence characteristics of the YMR318C gene product, we have proposed this gene to be called ADH6. The disruption of ADH6 was not lethal for the yeast under laboratory conditions. Although S. cerevisiae is considered a non lignin-degrading organism, the catalytic activity of ADHVI can direct veratraldehyde and anisaldehyde, arising from the oxidation of lignocellulose by fungal lignin peroxidases, to the lignin biodegradation pathway. ADHVI is the only S. cerevisiae enzyme able to significantly reduce veratraldehyde in vivo, and its overexpression allowed yeast to grow under toxic concentrations of this aldehyde. The enzyme may also be involved in the synthesis of fusel alcohols. To our knowledge this is the first NADPH-dependent medium-chain ADH to be characterized in S. cerevisiae. PMID:11742541

  4. Characterization of the Saccharomyces cerevisiae YMR318C (ADH6) gene product as a broad specificity NADPH-dependent alcohol dehydrogenase: relevance in aldehyde reduction.

    PubMed

    Larroy, Carol; Fernández, M Rosario; González, Eva; Parés, Xavier; Biosca, Josep A

    2002-01-01

    YMR318C represents an open reading frame from Saccharomyces cerevisiae with unknown function. It possesses a conserved sequence motif, the zinc-containing alcohol dehydrogenase (ADH) signature, specific to the medium-chain zinc-containing ADHs. In the present study, the YMR318C gene product has been purified to homogeneity from overexpressing yeast cells, and found to be a homodimeric ADH, composed of 40 kDa subunits and with a pI of 5.0-5.4. The enzyme was strictly specific for NADPH and was active with a wide variety of substrates, including aliphatic (linear and branched-chain) and aromatic primary alcohols and aldehydes. Aldehydes were processed with a 50-fold higher catalytic efficiency than that for the corresponding alcohols. The highest k(cat)/K(m) values were found with pentanal>veratraldehyde > hexanal > 3-methylbutanal >cinnamaldehyde. Taking into consideration the substrate specificity and sequence characteristics of the YMR318C gene product, we have proposed this gene to be called ADH6. The disruption of ADH6 was not lethal for the yeast under laboratory conditions. Although S. cerevisiae is considered a non lignin-degrading organism, the catalytic activity of ADHVI can direct veratraldehyde and anisaldehyde, arising from the oxidation of lignocellulose by fungal lignin peroxidases, to the lignin biodegradation pathway. ADHVI is the only S. cerevisiae enzyme able to significantly reduce veratraldehyde in vivo, and its overexpression allowed yeast to grow under toxic concentrations of this aldehyde. The enzyme may also be involved in the synthesis of fusel alcohols. To our knowledge this is the first NADPH-dependent medium-chain ADH to be characterized in S. cerevisiae.

  5. Over-expression of different aldehyde dehydrogenase genes in Arabidopsis thaliana confers tolerance to abiotic stress and protects plants against lipid peroxidation and oxidative stress.

    PubMed

    Kotchoni, Simeon O; Kuhns, Christine; Ditzer, Andrea; Kirch, Hans-Hubert; Bartels, Dorothea

    2006-06-01

    Aldehyde dehydrogenases (ALDHs) play a major role in the detoxification processes of aldehydes generated in plants when exposed to abiotic stress. In previous studies, we have shown that the Arabidopsis thaliana ALDH3I1 gene is transcriptionally activated by abiotic stress, and over-expression of the ALDH3I1 gene confers stress tolerance in transgenic plants. The A. thaliana genome contains 14 ALDH genes expressed in different sub-cellular compartments and are presumably involved in different reactions. The purpose of this study was to compare the potential of a cytoplasmic and a chloroplastic stress-inducible ALDH in conferring stress tolerance under different conditions. We demonstrated that constitutive or stress-inducible expression of both the chloroplastic ALDH3I1 and the cytoplasmic ALDH7B4 confers tolerance to osmotic and oxidative stress. Stress tolerance in transgenic plants is accompanied by a reduction of H2O2 and malondialdehyde (MDA) derived from cellular lipid peroxidation. Involvement of ALDHs in stress tolerance was corroborated by the analysis of ALDH3I1 and ALDH7B4 T-DNA knockout (KO) mutants. Both mutant lines exhibited higher sensitivity to dehydration and salt than wild-type (WT) plants. The results indicate that ALDH3I1 and ALDH7B4 not only function as aldehyde-detoxifying enzymes, but also as efficient reactive oxygen species (ROS) scavengers and lipid peroxidation-inhibiting enzymes. The potential of ALDHs to interfere with H2O2 was also shown for recombinant bacterial proteins.

  6. Cell Active Hydroxylactam Inhibitors of Human Lactate Dehydrogenase with Oral Bioavailability in Mice.

    PubMed

    Purkey, Hans E; Robarge, Kirk; Chen, Jinhua; Chen, Zhongguo; Corson, Laura B; Ding, Charles Z; DiPasquale, Antonio G; Dragovich, Peter S; Eigenbrot, Charles; Evangelista, Marie; Fauber, Benjamin P; Gao, Zhenting; Ge, Hongxiu; Hitz, Anna; Ho, Qunh; Labadie, Sharada S; Lai, Kwong Wah; Liu, Wenfeng; Liu, Yajing; Li, Chiho; Ma, Shuguang; Malek, Shiva; O'Brien, Thomas; Pang, Jodie; Peterson, David; Salphati, Laurent; Sideris, Steve; Ultsch, Mark; Wei, BinQing; Yen, Ivana; Yue, Qin; Zhang, Huihui; Zhou, Aihe

    2016-10-13

    A series of trisubstituted hydroxylactams was identified as potent enzymatic and cellular inhibitors of human lactate dehydrogenase A. Utilizing structure-based design and physical property optimization, multiple inhibitors were discovered with <10 μM lactate IC50 in a MiaPaca2 cell line. Optimization of the series led to 29, a potent cell active molecule (MiaPaca2 IC50 = 0.67 μM) that also possessed good exposure when dosed orally to mice.

  7. Significant improvement of stress tolerance in tobacco plants by overexpressing a stress-responsive aldehyde dehydrogenase gene from maize (Zea mays).

    PubMed

    Huang, Weizao; Ma, Xinrong; Wang, Qilin; Gao, Yongfeng; Xue, Ying; Niu, Xiangli; Yu, Guirong; Liu, Yongsheng

    2008-11-01

    Aldehyde dehydrogenases (ALDHs) play a central role in detoxification processes of aldehydes generated in plants when exposed to the stressed conditions. In order to identify genes required for the stresses responses in the grass crop Zea mays, an ALDH (ZmALDH22A1) gene was isolated and characterized. ZmALDH22A1 belongs to the family ALDH22 that is currently known only in plants. The ZmALDH22A1 encodes a protein of 593 amino acids that shares high identity with the orthologs from Saccharum officinarum (95%), Oryza sativa (89%), Triticum aestivum (87%) and Arabidopsis thaliana (77%), respectively. Real-time PCR analysis indicates that ZmALDH22A1 is expressed differentially in different tissues. Various elevated levels of ZmALDH22A1 expression have been detected when the seedling roots exposed to abiotic stresses including dehydration, high salinity and abscisic acid (ABA). Tomato stable transformation of construct expressing the ZmALDH22A1 signal peptide fused with yellow fluorescent protein (YFP) driven by the CaMV35S-promoter reveals that the fusion protein is targeted to plastid. Transgenic tobacco plants overexpressing ZmALDH22A1 shows elevated stresses tolerance. Stresses tolerance in transgenic plants is accompanied by a reduction of malondialdehyde (MDA) derived from cellular lipid peroxidation.

  8. Human Salivary Aldehyde Dehydrogenase: Purification, Kinetic Characterization and Effect of Ethanol, Hydrogen Peroxide and Sodium Dodecyl Sulfate on the Activity of the Enzyme.

    PubMed

    Alam, Md Fazle; Laskar, Amaj Ahmed; Choudhary, Hadi Hasan; Younus, Hina

    2016-09-01

    Human salivary aldehyde dehydrogenase (hsALDH) enzyme appears to be the first line of defense in the body against exogenous toxic aldehydes. However till date much work has not been done on this important member of the ALDH family. In this study, we have purified hsALDH to homogeneity by diethylaminoethyl-cellulose (DEAE-cellulose) ion-exchange chromatography in a single step. The molecular mass of the homodimeric enzyme was determined to be approximately 108 kDa. Four aromatic substrates; benzaldehyde, cinnamaldehyde, 2-naphthaldehyde and 6-methoxy-2-naphthaldehyde were used for determining the activity of pure hsALDH. K m values for these substrates were calculated to be 147.7, 5.31, 0.71 and 3.31 μM, respectively. The best substrates were found to be cinnamaldehyde and 2-naphthaldehyde since they exhibited high V max /K m values. 6-methoxy-2-naphthaldehyde substrate was used for further kinetic characterization of pure hsALDH. The pH and temperature optima of hsALDH were measured to be pH 8 and 45 °C, respectively. The pure enzyme is highly unstable at high temperatures. Ethanol, hydrogen peroxide and SDS activate hsALDH, therefore it is safe and beneficial to include them in mouthwashes and toothpastes in low concentrations.

  9. Characterization of the molecular mechanisms underlying increased ischemic damage in the aldehyde dehydrogenase 2 genetic polymorphism using a human induced pluripotent stem cell model system

    PubMed Central

    Ebert, Antje D.; Kodo, Kazuki; Liang, Ping; Wu, Haodi; Huber, Bruno C.; Riegler, Johannes; Churko, Jared; Lee, Jaecheol; de Almeida, Patricia; Lan, Feng; Diecke, Sebastian; Burridge, Paul W.; Gold, Joseph D.; Mochly-Rosen, Daria; Wu, Joseph C.

    2014-01-01

    Nearly 8% of the human population carries an inactivating point mutation in the gene that encodes the cardioprotective enzyme aldehyde dehydrogenase 2 (ALDH2). This genetic polymorphism (ALDH2*2) is linked to more severe outcomes from ischemic heart damage and an increased risk of coronary artery disease (CAD), but the underlying molecular bases are unknown. We investigated the ALDH2*2 mechanisms in a human model system of induced pluripotent stem cell–derived cardiomyocytes (iPSC-CMs) generated from individuals carrying the most common heterozygous form of the ALDH2*2 genotype. We showed that the ALDH2*2 mutation gave rise to elevated amounts of reactive oxygen species and toxic aldehydes, thereby inducing cell cycle arrest and activation of apoptotic signaling pathways, especially during ischemic injury. We established that ALDH2 controls cell survival decisions by modulating oxidative stress levels and that this regulatory circuitry was dysfunctional in the loss-of-function ALDH2*2 genotype, causing up-regulation of apoptosis in cardiomyocytes after ischemic insult. These results reveal a new function for the metabolic enzyme ALDH2 in modulation of cell survival decisions. Insight into the molecular mechanisms that mediate ALDH2*2-related increased ischemic damage is important for the development of specific diagnostic methods and improved risk management of CAD and may lead to patient-specific cardiac therapies. PMID:25253673

  10. Association between the aldehyde dehydrogenase 2*2 allele and smoking-related chronic airway obstruction in a Japanese general population: a pilot study.

    PubMed

    Morita, Kazunori; Masuda, Natsuki; Oniki, Kentaro; Saruwatari, Junji; Kajiwara, Ayami; Otake, Koji; Ogata, Yasuhiro; Nakagawa, Kazuko

    2015-07-16

    Aldehyde dehydrogenase 2 (ALDH2) detoxifies exogenous and endogenous toxic aldehydes; however, its protective effect against cigarette smoke in airways is unknown. We therefore examined whether the inactive ALDH2*2 allele is associated with smoking-related chronic airway obstruction. We conducted a cross-sectional study including 684 Japanese participants in a health screening program, and a retrospective longitudinal study in the elderly subgroup. The risks of airway obstruction in the ever-smokers with the ALDH2*1/*2 and *2/*2 genotypes were two and three times higher, respectively, than in the never-smokers with the ALDH2*1/*1 genotype. Moreover, the combined effect of smoking and the ALDH2*2 allele was prominent in the asthmatic subjects. In a longitudinal association analysis, the combination of the ALDH2 genotype and pack-years of smoking synergistically increased the risk of airway obstruction. The number of pack-years of smoking at baseline was identified to be a significant predictor of airway obstruction only in the ALDH2*2 allele carriers. In addition, the ALDH2*2 allele was also associated with the incidence of smoking-related airway obstruction, in the Cox proportional hazards model. This pilot study demonstrated for the first time a significant gene-environment interaction between the ALDH2*2 allele and cumulative exposure to cigarette smoke on the risk of airway obstruction.

  11. Combined effects of current-smoking and the aldehyde dehydrogenase 2*2 allele on the risk of myocardial infarction in Japanese patients.

    PubMed

    Morita, Kazunori; Miyazaki, Hiroko; Saruwatari, Junji; Oniki, Kentaro; Kumagae, Naoki; Tanaka, Takahiro; Kajiwara, Ayami; Otake, Koji; Ogata, Yasuhiro; Arima, Yuichiro; Hokimoto, Seiji; Ogawa, Hisao; Nakagawa, Kazuko

    2015-01-05

    Aldehyde dehydrogenase 2 (ALDH2) detoxifies toxic aldehydes, e.g. acetaldehyde in cigarette smoke; however, the interactive effects between smoking status and the ALDH2 genotype on coronary artery disease (CAD) have not been reported. We investigated the effects of smoking status and the ALDH2 genotype, and assessed their interactive and combined effects on the risk of myocardial infarction (MI) or stable angina (SA), including 221 MI and 175 SA subjects and 473 age- and sex-matched controls without CAD. Current-smoking and the ALDH2*2 allele additively increased the risk of MI (adjusted odds ratio 4.54, 95% confidence interval 2.25-9.15), although this combination was not associated with the risk of SA. This combination also increased the peak creatine kinase (CK) level synergistically in the acute MI (AMI) subjects. Moreover, current-smoking was found to be a significant risk factor for an increased peak CK level in the ALDH2*2 allele carriers (B 2220.2IU/L, p=0.008), but not the non-carriers. Additionally, a synergistic effect of this combination on the triglycerides levels was also found in the AMI subjects. These preliminary findings suggest that the combination of current-smoking and the inactive ALDH2*2 allele may increase the risk of MI additively and the infarct size synergistically.

  12. Synthesis and Evaluation of Macrocyclic Peptide Aldehydes as Potent and Selective Inhibitors of the 20S Proteasome.

    PubMed

    Wilson, David L; Meininger, Isabel; Strater, Zack; Steiner, Stephanie; Tomlin, Frederick; Wu, Julia; Jamali, Haya; Krappmann, Daniel; Götz, Marion G

    2016-03-10

    This research explores the first design and synthesis of macrocyclic peptide aldehydes as potent inhibitors of the 20S proteasome. Two novel macrocyclic peptide aldehydes based on the ring-size of the macrocyclic natural product TMC-95 were prepared and evaluated as inhibitors of the 20S proteasome. Both compounds inhibited in the low nanomolar range and proved to be selective for the proteasome over other serine and cysteine proteases, particularly when compared to linear analogues with similar amino acid sequences. In HeLa cells, both macrocycles efficiently inhibited activation of nuclear factor-κB (NF-κB) transcription factor by blocking proteasomal degradation of the inhibitor protein IκBα after cytokine stimulation. Due to their covalent mechanism of binding these compounds represent a 1000-fold increase in inhibitory potency over previously reported noncovalently binding TMC-95 analogues. Molecular modeling of the macrocyclic peptides confirms the preference of the large S3 pocket for large, hydrophobic residues and the ability to exploit this to improve selectivity of proteasome inhibitors.

  13. Synthesis and Evaluation of Macrocyclic Peptide Aldehydes as Potent and Selective Inhibitors of the 20S Proteasome

    PubMed Central

    2016-01-01

    This research explores the first design and synthesis of macrocyclic peptide aldehydes as potent inhibitors of the 20S proteasome. Two novel macrocyclic peptide aldehydes based on the ring-size of the macrocyclic natural product TMC-95 were prepared and evaluated as inhibitors of the 20S proteasome. Both compounds inhibited in the low nanomolar range and proved to be selective for the proteasome over other serine and cysteine proteases, particularly when compared to linear analogues with similar amino acid sequences. In HeLa cells, both macrocycles efficiently inhibited activation of nuclear factor-κB (NF-κB) transcription factor by blocking proteasomal degradation of the inhibitor protein IκBα after cytokine stimulation. Due to their covalent mechanism of binding these compounds represent a 1000-fold increase in inhibitory potency over previously reported noncovalently binding TMC-95 analogues. Molecular modeling of the macrocyclic peptides confirms the preference of the large S3 pocket for large, hydrophobic residues and the ability to exploit this to improve selectivity of proteasome inhibitors. PMID:26985310

  14. Enzymatic oxidation of phthalazine with guinea pig liver aldehyde oxidase and liver slices: inhibition by isovanillin.

    PubMed

    Panoutsopoulos, Georgios I; Beedham, Christine

    2004-01-01

    The enzymes aldehyde oxidase and xanthine oxidase catalyze the oxidation of a wide range of N-heterocycles and aldehydes. These enzymes are widely known for their role in the metabolism of N-heterocyclic xenobiotics where they provide a protective barrier by aiding in the detoxification of ingested nitrogen-containing heterocycles. Isovanillin has been shown to inhibit the metabolism of aromatic aldehydes by aldehyde oxidase, but its inhibition towards the heterocyclic compounds has not been studied. The present investigation examines the oxidation of phthalazine in the absence and in the presence of the inhibitor isovanillin by partially purified aldehyde oxidase from guinea pig liver. In addition, the interaction of phthalazine with freshly prepared guinea pig liver slices, both in the absence and presence of specific inhibitors of several liver oxidizing enzymes, was investigated. ldehyde oxidase rapidly converted phthalazine into 1-phthalazinone, which was completely inhibited in the presence of isovanillin (a specific inhibitor of aldehyde oxidase). In freshly prepared liver slices, phthalazine was also rapidly converted to 1-phthalazinone. The formation of 1-phthalazinone was completely inhibited by isovanillin, whereas disulfiram (a specific inhibitor of aldehyde dehydrogenase) only inhibited 1-phthalazinone formation by 24% and allopurinol (a specific inhibitor of xanthine oxidase) had little effect. Therefore, isovanillin has been proved as an inhibitor of the metabolism of heterocyclic substrates, such as phthalazine, by guinea pig liver aldehyde oxidase, since it had not been tested before. Thus it would appear from the inhibitor results that aldehyde oxidase is the predominant enzyme in the oxidation of phthalazine to 1-phthalazinone in freshly prepared guinea pig liver slices, whereas xanthine oxidase only contributes to a small extent and aldehyde dehydrogenase does not take any part.

  15. Exploring the evolutionary route of the acquisition of betaine aldehyde dehydrogenase activity by plant ALDH10 enzymes: implications for the synthesis of the osmoprotectant glycine betaine

    PubMed Central

    2014-01-01

    Background Plant ALDH10 enzymes are aminoaldehyde dehydrogenases (AMADHs) that oxidize different ω-amino or trimethylammonium aldehydes, but only some of them have betaine aldehyde dehydrogenase (BADH) activity and produce the osmoprotectant glycine betaine (GB). The latter enzymes possess alanine or cysteine at position 441 (numbering of the spinach enzyme, SoBADH), while those ALDH10s that cannot oxidize betaine aldehyde (BAL) have isoleucine at this position. Only the plants that contain A441- or C441-type ALDH10 isoenzymes accumulate GB in response to osmotic stress. In this work we explored the evolutionary history of the acquisition of BAL specificity by plant ALDH10s. Results We performed extensive phylogenetic analyses and constructed and characterized, kinetically and structurally, four SoBADH variants that simulate the parsimonious intermediates in the evolutionary pathway from I441-type to A441- or C441-type enzymes. All mutants had a correct folding, average thermal stabilities and similar activity with aminopropionaldehyde, but whereas A441S and A441T exhibited significant activity with BAL, A441V and A441F did not. The kinetics of the mutants were consistent with their predicted structural features obtained by modeling, and confirmed the importance of position 441 for BAL specificity. The acquisition of BADH activity could have happened through any of these intermediates without detriment of the original function or protein stability. Phylogenetic studies showed that this event occurred independently several times during angiosperms evolution when an ALDH10 gene duplicate changed the critical Ile residue for Ala or Cys in two consecutive single mutations. ALDH10 isoenzymes frequently group in two clades within a plant family: one includes peroxisomal I441-type, the other peroxisomal and non-peroxisomal I441-, A441- or C441-type. Interestingly, high GB-accumulators plants have non-peroxisomal A441- or C441-type isoenzymes, while low-GB accumulators

  16. Novel and potent 17beta-hydroxysteroid dehydrogenase type 1 inhibitors.

    PubMed

    Lawrence, Harshani R; Vicker, Nigel; Allan, Gillian M; Smith, Andrew; Mahon, Mary F; Tutill, Helena J; Purohit, Atul; Reed, Michael J; Potter, Barry V L

    2005-04-21

    Structure-based drug design using the crystal structure of human 17beta-hydroxysteroid dehydrogenase type 1 (17beta-HSD1) led to the discovery of novel, selective, and the most potent inhibitors of 17beta-HSD1 reported to date. Compounds 1 and 2 contain a side chain with an m-pyridylmethyl-amide functionality extended from the 16beta position of a steroid scaffold. A mode of binding is proposed for these inhibitors, and 2 is a steroid-based 17beta-HSD1 inhibitor with the potential for further development.

  17. Selective Inhibitors of 17beta-Hydroxysteroid Dehydrogenase

    DTIC Science & Technology

    2003-07-01

    this project is to develop selective inhibitors of human Type I 17 Beta-HSD as "lead compounds" for stmcture-based drug design . The crystal structure...of human Type I 17 Beta-HSD is available to aid in structure-based drug design . The concept that the Rossmann fold may represent a useful drug target is a new concept in drug design .

  18. A novel cofactor-binding mode in bacterial IMP dehydrogenases explains inhibitor selectivity

    DOE PAGES

    Makowska-Grzyska, Magdalena; Kim, Youngchang; Maltseva, Natalia; ...

    2015-01-09

    The steadily rising frequency of emerging diseases and antibiotic resistance creates an urgent need for new drugs and targets. Inosine 5'-monophosphate dehydrogenase (IMP dehydrogenase or IMPDH) is a promising target for the development of new antimicrobial agents. IMPDH catalyzes the oxidation of IMP to XMP with the concomitant reduction of NAD+, which is the pivotal step in the biosynthesis of guanine nucleotides. Potent inhibitors of bacterial IMPDHs have been identified that bind in a structurally distinct pocket that is absent in eukaryotic IMPDHs. The physiological role of this pocket was not understood. Here, we report the structures of complexes withmore » different classes of inhibitors of Bacillus anthracis, Campylobacter jejuni, and Clostridium perfringens IMPDHs. These structures in combination with inhibition studies provide important insights into the interactions that modulate selectivity and potency. We also present two structures of the Vibrio cholerae IMPDH in complex with IMP/NAD+ and XMP/NAD+. In both structures, the cofactor assumes a dramatically different conformation than reported previously for eukaryotic IMPDHs and other dehydrogenases, with the major change observed for the position of the NAD+ adenosine moiety. More importantly, this new NAD+-binding site involves the same pocket that is utilized by the inhibitors. Thus, the bacterial IMPDH-specific NAD+-binding mode helps to rationalize the conformation adopted by several classes of prokaryotic IMPDH inhibitors. As a result, these findings offer a potential strategy for further ligand optimization.« less

  19. A novel cofactor-binding mode in bacterial IMP dehydrogenases explains inhibitor selectivity.

    PubMed

    Makowska-Grzyska, Magdalena; Kim, Youngchang; Maltseva, Natalia; Osipiuk, Jerzy; Gu, Minyi; Zhang, Minjia; Mandapati, Kavitha; Gollapalli, Deviprasad R; Gorla, Suresh Kumar; Hedstrom, Lizbeth; Joachimiak, Andrzej

    2015-02-27

    The steadily rising frequency of emerging diseases and antibiotic resistance creates an urgent need for new drugs and targets. Inosine 5'-monophosphate dehydrogenase (IMP dehydrogenase or IMPDH) is a promising target for the development of new antimicrobial agents. IMPDH catalyzes the oxidation of IMP to XMP with the concomitant reduction of NAD(+), which is the pivotal step in the biosynthesis of guanine nucleotides. Potent inhibitors of bacterial IMPDHs have been identified that bind in a structurally distinct pocket that is absent in eukaryotic IMPDHs. The physiological role of this pocket was not understood. Here, we report the structures of complexes with different classes of inhibitors of Bacillus anthracis, Campylobacter jejuni, and Clostridium perfringens IMPDHs. These structures in combination with inhibition studies provide important insights into the interactions that modulate selectivity and potency. We also present two structures of the Vibrio cholerae IMPDH in complex with IMP/NAD(+) and XMP/NAD(+). In both structures, the cofactor assumes a dramatically different conformation than reported previously for eukaryotic IMPDHs and other dehydrogenases, with the major change observed for the position of the NAD(+) adenosine moiety. More importantly, this new NAD(+)-binding site involves the same pocket that is utilized by the inhibitors. Thus, the bacterial IMPDH-specific NAD(+)-binding mode helps to rationalize the conformation adopted by several classes of prokaryotic IMPDH inhibitors. These findings offer a potential strategy for further ligand optimization. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. A Novel Cofactor-binding Mode in Bacterial IMP Dehydrogenases Explains Inhibitor Selectivity*

    PubMed Central

    Makowska-Grzyska, Magdalena; Kim, Youngchang; Maltseva, Natalia; Osipiuk, Jerzy; Gu, Minyi; Zhang, Minjia; Mandapati, Kavitha; Gollapalli, Deviprasad R.; Gorla, Suresh Kumar; Hedstrom, Lizbeth; Joachimiak, Andrzej

    2015-01-01

    The steadily rising frequency of emerging diseases and antibiotic resistance creates an urgent need for new drugs and targets. Inosine 5′-monophosphate dehydrogenase (IMP dehydrogenase or IMPDH) is a promising target for the development of new antimicrobial agents. IMPDH catalyzes the oxidation of IMP to XMP with the concomitant reduction of NAD+, which is the pivotal step in the biosynthesis of guanine nucleotides. Potent inhibitors of bacterial IMPDHs have been identified that bind in a structurally distinct pocket that is absent in eukaryotic IMPDHs. The physiological role of this pocket was not understood. Here, we report the structures of complexes with different classes of inhibitors of Bacillus anthracis, Campylobacter jejuni, and Clostridium perfringens IMPDHs. These structures in combination with inhibition studies provide important insights into the interactions that modulate selectivity and potency. We also present two structures of the Vibrio cholerae IMPDH in complex with IMP/NAD+ and XMP/NAD+. In both structures, the cofactor assumes a dramatically different conformation than reported previously for eukaryotic IMPDHs and other dehydrogenases, with the major change observed for the position of the NAD+ adenosine moiety. More importantly, this new NAD+-binding site involves the same pocket that is utilized by the inhibitors. Thus, the bacterial IMPDH-specific NAD+-binding mode helps to rationalize the conformation adopted by several classes of prokaryotic IMPDH inhibitors. These findings offer a potential strategy for further ligand optimization. PMID:25572472

  1. Structural, Biochemical, and Computational Studies Reveal the Mechanism of Selective Aldehyde Dehydrogenase 1A1 Inhibition by Cytotoxic Duocarmycin Analogues.

    PubMed

    Koch, Maximilian F; Harteis, Sabrina; Blank, Iris D; Pestel, Galina; Tietze, Lutz F; Ochsenfeld, Christian; Schneider, Sabine; Sieber, Stephan A

    2015-11-09

    Analogues of the natural product duocarmycin bearing an indole moiety were shown to bind aldehyde dehydrogenase 1A1 (ALDH1A1) in addition to DNA, while derivatives without the indole solely addressed the ALDH1A1 protein. The molecular mechanism of selective ALDH1A1 inhibition by duocarmycin analogues was unraveled through cocrystallization, mutational studies, and molecular dynamics simulations. The structure of the complex shows the compound embedded in a hydrophobic pocket, where it is stabilized by several crucial π-stacking and van der Waals interactions. This binding mode positions the cyclopropyl electrophile for nucleophilic attack by the noncatalytic residue Cys302, thereby resulting in covalent attachment, steric occlusion of the active site, and inhibition of catalysis. The selectivity of duocarmycin analogues for ALDH1A1 is unique, since only minor alterations in the sequence of closely related protein isoforms restrict compound accessibility. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Mitochondrial aldehyde dehydrogenase 2 plays protective roles in heart failure after myocardial infarction via suppression of the cytosolic JNK/p53 pathway in mice.

    PubMed

    Sun, Aijun; Zou, Yunzeng; Wang, Ping; Xu, Danling; Gong, Hui; Wang, Shijun; Qin, Yingjie; Zhang, Peng; Chen, Yunqin; Harada, Mutsuo; Isse, Toyoshi; Kawamoto, Toshihiro; Fan, Huizhi; Yang, Pengyuan; Akazawa, Hiroshi; Nagai, Toshio; Takano, Hiroyuki; Ping, Peipei; Komuro, Issei; Ge, Junbo

    2014-09-18

    Increasing evidence suggests a critical role for mitochondrial aldehyde dehydrogenase 2 (ALDH2) in protection against cardiac injuries; however, the downstream cytosolic actions of this enzyme are largely undefined. Proteomic analysis identified a significant downregulation of mitochondrial ALDH2 in the heart of a rat heart failure model after myocardial infarction. The mechanistic insights underlying ALDH2 action were elucidated using murine models overexpressing ALDH2 or its mutant or with the ablation of the ALDH2 gene (ALDH2 knockout) and neonatal cardiomyocytes undergoing altered expression and activity of ALDH2. Left ventricle dilation and dysfunction and cardiomyocyte death after myocardial infarction were exacerbated in ALDH2-knockout or ALDH2 mutant-overexpressing mice but were significantly attenuated in ALDH2-overexpressing mice. Using an anoxia model of cardiomyocytes with deficiency in ALDH2 activities, we observed prominent cardiomyocyte apoptosis and increased accumulation of the reactive aldehyde 4-hydroxy-2-nonenal (4-HNE). We subsequently examined the impacts of mitochondrial ALDH2 and 4-HNE on the relevant cytosolic protective pathways. Our data documented 4-HNE-stimulated p53 upregulation via the phosphorylation of JNK, accompanying increased cardiomyocyte apoptosis that was attenuated by inhibition of p53. Importantly, elevation of 4-HNE also triggered a reduction of the cytosolic HSP70, further corroborating cytosolic action of the 4-HNE instigated by downregulation of mitochondrial ALDH2. Downregulation of ALDH2 in the mitochondria induced an elevation of 4-HNE, leading to cardiomyocyte apoptosis by subsequent inhibition of HSP70, phosphorylation of JNK, and activation of p53. This chain of molecular events took place in both the mitochondria and the cytosol, contributing to the mechanism underlying heart failure. © 2014 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

  3. Cloning and molecular characterization of the betaine aldehyde dehydrogenase involved in the biosynthesis of glycine betaine in white shrimp (Litopenaeus vannamei).

    PubMed

    Delgado-Gaytán, María F; Rosas-Rodríguez, Jesús A; Yepiz-Plascencia, Gloria; Figueroa-Soto, Ciria G; Valenzuela-Soto, Elisa M

    2017-02-15

    The enzyme betaine aldehyde dehydrogenase (BADH) catalyzes the irreversible oxidation of betaine aldehyde to glycine betaine (GB), a very efficient osmolyte accumulated during osmotic stress. In this study, we determined the nucleotide sequence of the cDNA for the BADH from the white shrimp Litopenaeus vannamei (LvBADH). The cDNA was 1882 bp long, with a complete open reading frame of 1524 bp, encoding 507 amino acids with a predicted molecular mass of 54.15 kDa and a pI of 5.4. The predicted LvBADH amino acid sequence shares a high degree of identity with marine invertebrate BADHs. Catalytic residues (C-298, E-264 and N-167) and the decapeptide VTLELGGKSP involved in nucleotide binding and highly conserved in BADHs were identified in the amino acid sequence. Phylogenetic analyses classified LvBADH in a clade that includes ALDH9 sequences from marine invertebrates. Molecular modeling of LvBADH revealed that the protein has amino acid residues and sequence motifs essential for the function of the ALDH9 family of enzymes. LvBADH modeling showed three potential monovalent cation binding sites, one site is located in an intra-subunit cavity; other in an inter-subunit cavity and a third in a central-cavity of the protein. The results show that LvBADH shares a high degree of identity with BADH sequences from marine invertebrates and enzymes that belong to the ALDH9 family. Our findings suggest that the LvBADH has molecular mechanisms of regulation similar to those of other BADHs belonging to the ALDH9 family, and that BADH might be playing a role in the osmoregulation capacity of L. vannamei.

  4. Molecular characterization and organ-specific expression of the gene that encodes betaine aldehyde dehydrogenase from the white shrimp Litopenaeus vannamei in response to osmotic stress.

    PubMed

    Delgado-Gaytán, María F; Hernández-Palomares, Magally L E; Soñanez-Organis, José G; Muhlia-Almazán, Adriana; Sánchez-Paz, Arturo; Stephens-Camacho, Norma A; Valenzuela-Soto, Elisa M; Rosas-Rodríguez, Jesús A

    2015-11-01

    Crustaceans overcome osmotic disturbances by regulating their intracellular concentration of ions and osmolytes. Glycine betaine (GB), an osmolyte accumulated in response to hyperosmotic stress, is synthesized by betaine aldehyde dehydrogenase (BADH EC 1.2.1.8) through the oxidation of betaine aldehyde. A partial BADH cDNA sequence from the white shrimp Litopenaeus vannamei was obtained and its organ-specific expression during osmotic stress (low and high salinity) was evaluated. The partial BADH cDNA sequence (LvBADH) is 1103bp long and encodes an open reading frame for 217 protein residues. The amino acid sequence of LvBADH is related to that of other BADHs, TMABA-DH and ALDH9 from invertebrate and vertebrate homologues, and includes the essential domains of their function and regulation. LvBADH activity and mRNA expression were detected in the gills, hepatopancreas and muscle with the highest levels in the hepatopancreas. LvBADH mRNA expression increased 2-3-fold in the hepatopancreas and gills after 7days of osmotic variation (25 and 40ppt). In contrast, LvBADH mRNA expression in muscle decreased 4-fold and 15-fold after 7days at low and high salinity, respectively. The results indicate that LvBADH is ubiquitously expressed, but its levels are organ-specific and regulated by osmotic stress, and that LvBADH is involved in the cellular response of crustaceans to variations in environmental salinity. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. A wide host-range metagenomic library from a waste water treatment plant yields a novel alcohol/aldehyde dehydrogenase.

    PubMed

    Wexler, Margaret; Bond, Philip L; Richardson, David J; Johnston, Andrew W B

    2005-12-01

    Using DNA obtained from the metagenome of an anaerobic digestor in a waste water treatment plant, we constructed a gene library cloned in the wide host-range cosmid pLAFR3. One cosmid enabled Rhizobium leguminosarum to grow on ethanol as sole carbon and energy source, this being due to the presence of a gene, termed adhEMeta. The AdhEMeta protein most closely resembles the AdhE alcohol dehydrogenase of Clostridium acetobutylicum, where it catalyses the formation of ethanol and butanol in a two-step reductive process. However, cloned adhEMeta did not confer ethanol utilization ability to Escherichia coli or to Pseudomonas aeruginosa, even though it was transcribed in both these hosts. Further, cell-free extracts of E. coli and R. leguminosarum containing cloned adhEMeta had butanol and ethanol dehydrogenase activities when assayed in vitro. In contrast to the well-studied AdhE proteins of C. acetobutylicum and E. coli, the enzyme specified by adhEMeta is not inactivated by oxygen and it enables alcohol to be catabolized. Cloned adhEMeta did, however, confer one phenotype to E. coli. AdhE- mutants of E. coli fail to ferment glucose and introduction of adhEMeta restored the growth of such mutants when grown under fermentative conditions. These observations show that the use of wide host-range vectors enhances the efficacy with which metagenomic libraries can be screened for genes that confer novel functions.

  6. Identification of New Structural Fragments for the Design of Lactate Dehydrogenase A Inhibitors.

    PubMed

    Nilov, D K; Kulikov, A V; Prokhorova, E A; Švedas, V K

    2016-01-01

    Human lactate dehydrogenase A plays an important role in the glucose metabolism of tumor cells and constitutes an attractive target for chemotherapy. Molecular fragments able to bind in the active site of this enzyme and form hydrogen bonds with the Arg168 guanidinium group, as well as additional interactions with the loop 96-111 in the closed conformation, have been identified by virtual screening of sulfonates and experimental testing of their inhibitory effect. The sulfo group can occupy a similar position as the carboxyl group of the substrate and its structural analogs, whereas the benzothiazole group attached via a linker can be located in the coenzyme (NADH) binding site. Thus, the value of merging individual structural elements of the inhibitor by a linker was demonstrated and ways of further structural modification for the design of more effective inhibitors of lactate dehydrogenase A were established.

  7. Identification of New Structural Fragments for the Design of Lactate Dehydrogenase A Inhibitors

    PubMed Central

    Nilov, D.K.; Kulikov, A.V.; Prokhorova, E.A.; Švedas, V.K.

    2016-01-01

    Human lactate dehydrogenase A plays an important role in the glucose metabolism of tumor cells and constitutes an attractive target for chemotherapy. Molecular fragments able to bind in the active site of this enzyme and form hydrogen bonds with the Arg168 guanidinium group, as well as additional interactions with the loop 96–111 in the closed conformation, have been identified by virtual screening of sulfonates and experimental testing of their inhibitory effect. The sulfo group can occupy a similar position as the carboxyl group of the substrate and its structural analogs, whereas the benzothiazole group attached via a linker can be located in the coenzyme (NADH) binding site. Thus, the value of merging individual structural elements of the inhibitor by a linker was demonstrated and ways of further structural modification for the design of more effective inhibitors of lactate dehydrogenase A were established. PMID:27795851

  8. Structure-guided Development of Specific Pyruvate Dehydrogenase Kinase Inhibitors Targeting the ATP-binding Pocket*

    PubMed Central

    Tso, Shih-Chia; Qi, Xiangbing; Gui, Wen-Jun; Wu, Cheng-Yang; Chuang, Jacinta L.; Wernstedt-Asterholm, Ingrid; Morlock, Lorraine K.; Owens, Kyle R.; Scherer, Philipp E.; Williams, Noelle S.; Tambar, Uttam K.; Wynn, R. Max; Chuang, David T.

    2014-01-01

    Pyruvate dehydrogenase kinase isoforms (PDKs 1–4) negatively regulate activity of the mitochondrial pyruvate dehydrogenase complex by reversible phosphorylation. PDK isoforms are up-regulated in obesity, diabetes, heart failure, and cancer and are potential therapeutic targets for these important human diseases. Here, we employed a structure-guided design to convert a known Hsp90 inhibitor to a series of highly specific PDK inhibitors, based on structural conservation in the ATP-binding pocket. The key step involved the substitution of a carbonyl group in the parent compound with a sulfonyl in the PDK inhibitors. The final compound of this series, 2-[(2,4-dihydroxyphenyl)sulfonyl]isoindoline-4,6-diol, designated PS10, inhibits all four PDK isoforms with IC50 = 0.8 μm for PDK2. The administration of PS10 (70 mg/kg) to diet-induced obese mice significantly augments pyruvate dehydrogenase complex activity with reduced phosphorylation in different tissues. Prolonged PS10 treatments result in improved glucose tolerance and notably lessened hepatic steatosis in the mouse model. The results support the pharmacological approach of targeting PDK to control both glucose and fat levels in obesity and type 2 diabetes. PMID:24356970

  9. Inhibition of human alcohol and aldehyde dehydrogenases by aspirin and salicylate: assessment of the effects on first-pass metabolism of ethanol.

    PubMed

    Lee, Shou-Lun; Lee, Yung-Pin; Wu, Min-Li; Chi, Yu-Chou; Liu, Chiu-Ming; Lai, Ching-Long; Yin, Shih-Jiun

    2015-05-01

    Previous studies have reported that aspirin significantly reduced the first-pass metabolism (FPM) of ethanol in humans thereby increasing adverse effects of alcohol. The underlying causes, however, remain poorly understood. Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), principal enzymes responsible for metabolism of ethanol, are complex enzyme families that exhibit functional polymorphisms among ethnic groups and distinct tissue distributions. We investigated the inhibition profiles by aspirin and its major metabolite salicylate of ethanol oxidation by recombinant human ADH1A, ADH1B1, ADH1B2, ADH1B3, ADH1C1, ADH1C2, ADH2, and ADH4, and acetaldehyde oxidation by ALDH1A1 and ALDH2, at pH 7.5 and 0.5 mM NAD(+). Competitive inhibition pattern was found to be a predominant type among the ADHs and ALDHs studied, although noncompetitive and uncompetitive inhibitions were also detected in a few cases. The inhibition constants of salicylate for the ADHs and ALDHs were considerably lower than that of aspirin with the exception of ADH1A that can be ascribed to a substitution of Ala-93 at the bottom of substrate pocket as revealed by molecular docking experiments. Kinetic inhibition equation-based simulations show at higher therapeutic levels of blood plasma salicylate (1.5 mM) that the decrease of activities at 2-10 mM ethanol for ADH1A/ADH2 and ADH1B2/ADH1B3 are predicted to be 75-86% and 31-52%, respectively, and that the activity decline for ALDH1A1 and ALDH2 at 10-50 μM acetaldehyde to be 62-73%. Our findings suggest that salicylate may substantially inhibit hepatic FPM of alcohol at both the ADH and ALDH steps when concurrent intaking aspirin.

  10. Alcohol and aldehyde dehydrogenase polymorphisms and a new strategy for prevention and screening for cancer in the upper aerodigestive tract in East Asians.

    PubMed

    Yokoyama, Akira; Omori, Tai; Yokoyama, Tetsuji

    2010-01-01

    The ethanol in alcoholic beverages and the acetaldehyde associated with alcohol consumption are Group 1 human carcinogens (WHO, International Agency for Research on Cancer). The combination of alcohol consumption, tobacco smoking, the inactive heterozygous aldehyde dehydrogenase-2 genotype (ALDH2*1/*2) and the less-active homozygous alcohol dehydrogenase-1B genotype (ADH1B*1/*1) increases the risk of squamous cell carcinoma (SCC) in the upper aerodigestive tract (UADT) in a multiplicative fashion in East Asians. In addition to being exposed to locally high levels of ethanol, the UADT is exposed to a very high concentration of acetaldehyde from a variety of sources, including that as an ingredient of alcoholic beverages per se and that found in tobacco smoke; acetaldehyde is also produced by salivary microorganisms and mucosal enzymes and is present as blood acetaldehyde. The inefficient degradation of acetaldehyde by weakly expressed ALDH2 in the UADT may be cri! tical to the local accumulation of acetaldehyde, especially in ALDH2*1/*2 carriers. ADH1B*1/*1 carriers tend to experience less intense alcohol flushing and are highly susceptible to heavy drinking and alcoholism. Heavy drinking by persons with the less-active ADH1B*1/*1 leads to longer exposure of the UADT to salivary ethanol and acetaldehyde. The ALDH2*1/*2 genotype is a very strong predictor of synchronous and metachronous multiple SCCs in the UADT. High red cell mean corpuscular volume (MCV), esophageal dysplasia, and melanosis in the UADT, all of which are frequently found in ALDH2*1/*2 drinkers, are useful for identifying high-risk individuals. We invented a simple flushing questionnaire that enables prediction of the ALDH2 phenotype. New health appraisal models that include ALDH2 genotype, the simple flushing questionnaire, or MCV are powerful tools for devising a new strategy for prevention and screening for UADT cancer in East Asians.

  11. Genetic polymorphisms of alcohol dehydrogense-1B and aldehyde dehydrogenase-2, alcohol flushing, mean corpuscular volume, and aerodigestive tract neoplasia in Japanese drinkers.

    PubMed

    Yokoyama, Akira; Mizukami, Takeshi; Yokoyama, Tetsuji

    2015-01-01

    Genetic polymorphisms of alcohol dehydrogenase-1B (ADH1B) and aldehyde dehydrogenase-2 (ALDH2) modulate exposure levels to ethanol/acetaldehyde. Endoscopic screening of 6,014 Japanese alcoholics yielded high detection rates of esophageal squamous cell carcinoma (SCC; 4.1%) and head and neck SCC (1.0%). The risks of upper aerodigestive tract SCC/dysplasia, especially of multiple SCC/dysplasia, were increased in a multiplicative fashion by the presence of a combination of slow-metabolizing ADH1B*1/*1 and inactive heterozygous ALDH2*1/*2 because of prolonged exposure to higher concentrations of ethanol/acetaldehyde. A questionnaire asking about current and past facial flushing after drinking a glass (≈180 mL) of beer is a reliable tool for detecting the presence of inactive ALDH2. We invented a health-risk appraisal (HRA) model including the flushing questionnaire and drinking, smoking, and dietary habits. Esophageal SCC was detected at a high rate by endoscopic mass-screening in high HRA score persons. A total of 5.0% of 4,879 alcoholics had a history of (4.0%) or newly diagnosed (1.0%) gastric cancer. Their high frequency of a history of gastric cancer is partly explained by gastrectomy being a risk factor for alcoholism because of altered ethanol metabolism, e.g., by blood ethanol level overshooting. The combination of H. pylori-associated atrophic gastritis and ALDH2*1/*2 showed the greatest risk of gastric cancer in alcoholics. High detection rates of advanced colorectal adenoma/carcinoma were found in alcoholics, 15.7% of 744 immunochemical fecal occult blood test (IFOBT)-negative alcoholics and 31.5% of the 393 IFOBT-positive alcoholics. Macrocytosis with an MCV≥106 fl increased the risk of neoplasia in the entire aerodigestive tract of alcoholics, suggesting that poor nutrition as well as ethanol/acetaldehyde exposure plays an important role in neoplasia.

  12. The Structural Basis of Cryptosporidium-Specific IMP Dehydrogenase Inhibitor Selectivity

    SciTech Connect

    MacPherson, Iain S.; Kirubakaran, Sivapriya; Gorla, Suresh Kumar; Riera, Thomas V.; D’Aquino, J. Alejandro; Zhang, Minjia; Cuny, Gregory D.; Hedstrom, Lizbeth

    2010-03-29

    Cryptosporidium parvum is a potential biowarfare agent, an important AIDS pathogen, and a major cause of diarrhea and malnutrition. No vaccines or effective drug treatment exist to combat Cryptosporidium infection. This parasite relies on inosine 5{prime}-monophosphate dehydrogenase (IMPDH) to obtain guanine nucleotides, and inhibition of this enzyme blocks parasite proliferation. Here, we report the first crystal structures of CpIMPDH. These structures reveal the structural basis of inhibitor selectivity and suggest a strategy for further optimization. Using this information, we have synthesized low-nanomolar inhibitors that display 10{sup 3} selectivity for the parasite enzyme over human IMPDH2.

  13. Interconvertible geometric isomers of Plasmodium falciparum dihydroorotate dehydrogenase inhibitors exhibit multiple binding modes.

    PubMed

    McConkey, Glenn A; Bedingfield, Paul T P; Burrell, David R; Chambers, Nicholas C; Cunningham, Fraser; Prior, Timothy J; Fishwick, Colin W G; Boa, Andrew N

    2017-08-15

    Two new tricyclic β-aminoacrylate derivatives (2e and 3e) have been found to be inhibitors of Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) with Ki 0.037 and 0.15μM respectively. (1)H and (13)C NMR spectroscopic data show that these compounds undergo ready cis-trans isomerisation at room temperature in polar solvents. In silico docking studies indicate that for both molecules there is neither conformation nor double bond configuration which bind preferentially to PfDHODH. This flexibility is favourable for inhibitors of this channel that require extensive positioning to reach their binding site. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Engineering the isobutanol biosynthetic pathway in Escherichia coli by comparison of three aldehyde reductase/alcohol dehydrogenase genes.

    PubMed

    Atsumi, Shota; Wu, Tung-Yun; Eckl, Eva-Maria; Hawkins, Sarah D; Buelter, Thomas; Liao, James C

    2010-01-01

    Biofuels synthesized from renewable resources are of increasing interest because of global energy and environmental problems. We have previously demonstrated production of higher alcohols from Escherichia coli using a 2-keto acid-based pathway. Here, we have compared the effect of various alcohol dehydrogenases (ADH) for the last step of the isobutanol production. E. coli has the yqhD gene which encodes a broad-range ADH. Isobutanol production significantly decreased with the deletion of yqhD, suggesting that the yqhD gene on the genome contributed to isobutanol production. The adh genes of two bacteria and one yeast were also compared in E. coli harboring the isobutanol synthesis pathway. Overexpression of yqhD or adhA in E. coli showed better production than ADH2, a result confirmed by activity measurements with isobutyraldehyde.

  15. Alcohol and aldehyde dehydrogenase from Saccharomyces cerevisiae: specific activity and influence on the production of acetic acid, ethanol and higher alcohols in the first 48 h of fermentation of grape must.

    PubMed

    Millán, C; Mauricio, J C; Ortega, J M

    1990-01-01

    The changes in the specific activity of alcohol dehydrogenase (ADH-I and ADH-II) and aldehyde dehydrogenases [AIDH-NADP+ and AIDH-NAD(P)+] from Saccharomyces cerevisiae during the first 48 h of fermentation of grape must were investigated. The biosynthesis of ADH-I and AIDH-NADP+ took place basically during the adaptation of the yeasts to the must (first 4 h), while that of ADH-II occurred immediately after exponential growth (after 12 h). From the products produced by the yeast, only the specific rate of production of ethanol was found to be directly related to the specific activity of ADH-I.

  16. Novel inhibitors of 17beta-hydroxysteroid dehydrogenase type 1: templates for design.

    PubMed

    Allan, Gillian M; Vicker, Nigel; Lawrence, Harshani R; Tutill, Helena J; Day, Joanna M; Huchet, Marion; Ferrandis, Eric; Reed, Michael J; Purohit, Atul; Potter, Barry V L

    2008-04-15

    The 17beta-hydroxysteroid dehydrogenases (17beta-HSDs) catalyze the interconversion between the oxidized and reduced forms of androgens and estrogens at the 17 position. The 17beta-HSD type 1 enzyme (17beta-HSD1) catalyzes the reduction of estrone (E1) to estradiol and is expressed in malignant breast cells. Inhibitors of this enzyme thus have potential as treatments for hormone dependent breast cancer. Syntheses and biological evaluation of novel non-steroidal inhibitors designed to mimic the E1 template are reported using information from potent steroidal inhibitors. Of the templates investigated biphenyl ethanone was promising and led to inhibitors with IC(50) values in the low micromolar range.

  17. Cloning of the Membrane-Bound Aldehyde Dehydrogenase Gene of Acetobacter polyoxogenes and Improvement of Acetic Acid Production by Use of the Cloned Gene

    PubMed Central

    Fukaya, Masahiro; Tayama, Kenji; Tamaki, Toshimi; Tagami, Haruko; Okumura, Hajime; Kawamura, Yoshiya; Beppu, Teruhiko

    1989-01-01

    A genomic clone bank of Acetobacter polyoxogenes NBI1028 constructed in Escherichia coli by use of the expression vector pUC18 was screened with antibody raised against membrane-bound aldehyde dehydrogenase (ALDH; 75 kilodaltons [kDa]) from A. polyoxogenes NBI1028. A clone that synthesized a 41-kDa protein cross-reactive with anti-ALDH antibody was isolated. For cloning of the full-length ALDH structural gene, a cosmid gene bank was screened by Southern blot hybridization with the cloned DNA as a probe, and subcloning from the positive cosmid clone was performed with shuttle vector pMV24. Plasmid pAL25, containing the full-length ALDH structural gene, was isolated and expressed in both E. coli and Acetobacter aceti to produce a fused protein (78 kDa) with a short NH2-terminal β-galactosidase peptide. pAL25 conferred ALDH production on a mutant of A. aceti lacking the enzyme activity. Transformation of A. aceti subsp. xylinum NBI2099 with pAL25 caused 2- and 1.4-fold increases in the production rate and in the maximum concentration of acetic acid in submerged fermentation, respectively. Images PMID:16347820

  18. Double negative feedback loop between reprogramming factor LIN28 and microRNA let-7 regulates aldehyde dehydrogenase 1-positive cancer stem cells

    PubMed Central

    Yang, Xiaojun; Lin, Xiaojuan; Zhong, Xiaomin; Kaur, Sippy; Li, Ning; Liang, Shun; Lassus, Heini; Wang, Liping; Katsaros, Dionyssios; Montone, Kathleen; Zhao, Xia; Zhang, Youcheng; Bützow, Ralf; Coukos, George; Zhang, Lin

    2010-01-01

    A relatively rare aldehyde dehydrogenase 1 (ALDH1) positive “stem cell-like” subpopulation of tumor cells has the unique ability to initiate and perpetuate tumor growth; moreover it is highly resistant to chemotherapy and significantly associated with poor clinical outcomes. The development of more effective therapies for cancer requires targeting of this cell population. Using cDNA microarray analysis, we identified that the expression of the C. elegans lin-28 homolog (LIN28) was positively correlated with the percentage of ALDH1+ tumor cells; this was further validated in an independent set of tissue arrays (n=197). Both lose-of-function and gain-of-function studies demonstrated that LIN28 plays a critical role in the maintenance of ALDH1+ tumor cells. In addition, we found that there is a double negative feedback loop between LIN28 and let-7 in tumor cells, and that let-7 negatively regulates ALDH1+ tumor cells. Finally, we report that a LIN28/let-7 loop modulates self renewal and differentiation of mammary gland epithelial progenitor cells. Our data provide evidence that cancer stem cells may arise through a “reprogramming-like” mechanism. A rebalancing of the LIN28/let-7 regulatory loop could be a novel therapeutic strategy to target ALDH1+ cancer stem cells. PMID:21045151

  19. Pharmacological recruitment of aldehyde dehydrogenase 3A1 (ALDH3A1) to assist ALDH2 in acetaldehyde and ethanol metabolism in vivo

    PubMed Central

    Chen, Che-Hong; Cruz, Leslie A.; Mochly-Rosen, Daria

    2015-01-01

    Correcting a genetic mutation that leads to a loss of function has been a challenge. One such mutation is in aldehyde dehydrogenase 2 (ALDH2), denoted ALDH2*2. This mutation is present in ∼0.6 billion East Asians and results in accumulation of toxic acetaldehyde after consumption of ethanol. To temporarily increase metabolism of acetaldehyde in vivo, we describe an approach in which a pharmacologic agent recruited another ALDH to metabolize acetaldehyde. We focused on ALDH3A1, which is enriched in the upper aerodigestive track, and identified Alda-89 as a small molecule that enables ALDH3A1 to metabolize acetaldehyde. When given together with the ALDH2-specific activator, Alda-1, Alda-89 reduced acetaldehyde-induced behavioral impairment by causing a rapid reduction in blood ethanol and acetaldehyde levels after acute ethanol intoxication in both wild-type and ALDH2-deficient, ALDH2*1/*2, heterozygotic knock-in mice. The use of a pharmacologic agent to recruit an enzyme to metabolize a substrate that it usually does not metabolize may represent a novel means to temporarily increase elimination of toxic agents in vivo. PMID:25713355

  20. Pharmacological recruitment of aldehyde dehydrogenase 3A1 (ALDH3A1) to assist ALDH2 in acetaldehyde and ethanol metabolism in vivo.

    PubMed

    Chen, Che-Hong; Cruz, Leslie A; Mochly-Rosen, Daria

    2015-03-10

    Correcting a genetic mutation that leads to a loss of function has been a challenge. One such mutation is in aldehyde dehydrogenase 2 (ALDH2), denoted ALDH2*2. This mutation is present in ∼ 0.6 billion East Asians and results in accumulation of toxic acetaldehyde after consumption of ethanol. To temporarily increase metabolism of acetaldehyde in vivo, we describe an approach in which a pharmacologic agent recruited another ALDH to metabolize acetaldehyde. We focused on ALDH3A1, which is enriched in the upper aerodigestive track, and identified Alda-89 as a small molecule that enables ALDH3A1 to metabolize acetaldehyde. When given together with the ALDH2-specific activator, Alda-1, Alda-89 reduced acetaldehyde-induced behavioral impairment by causing a rapid reduction in blood ethanol and acetaldehyde levels after acute ethanol intoxication in both wild-type and ALDH2-deficient, ALDH2*1/*2, heterozygotic knock-in mice. The use of a pharmacologic agent to recruit an enzyme to metabolize a substrate that it usually does not metabolize may represent a novel means to temporarily increase elimination of toxic agents in vivo.

  1. Expression of aldehyde dehydrogenase family 1 member A1 and high mobility group box 1 in oropharyngeal squamous cell carcinoma in association with survival time

    PubMed Central

    Qian, Xu; Coordes, Annekatrin; Kaufmann, Andreas M.; Albers, Andreas E.

    2016-01-01

    Despite the development of novel multimodal treatment combinations in advanced oropharyngeal squamous cell carcinoma (OSCC), outcomes remain poor. The identification of specifically validated biomarkers is required to understand the underlying molecular mechanisms, to evaluate treatment efficiency and to develop novel therapeutic targets. The present study, therefore, examined the presence of aldehyde dehydrogenase family 1 member A1 (ALDH1A1) and high mobility group box 1 (HMGB1) expression in primary OSCC and analyzed the impact on survival time. In 59 patients with OSCC, the expression of ALDH1A1, p16 and HMGB1, and their clinicopathological data were analyzed. HMGB1 positivity was significantly increased in patients with T1-2 stage disease compared with T3-4 stage disease (P<0.001), whereas ALDH1A1 positivity was not. ALDH1A1+ tumors showed significantly lower differentiation than ALDH1A1− tumors (P=0.018). Multivariate analysis showed that ALDH1A1 positivity (P=0.041) and nodal status (N2-3) (P=0.036) predicted a poor prognosis. In this patient cohort, ALDH1A1 and nodal status were identified as independent predictors of a shorter overall survival time. The study results, therefore, provide evidence of the prognostic value of ALDH1A1 as a marker for cancer stem cells and nodal status in OSCC patients. PMID:27900016

  2. Expression of aldehyde dehydrogenase family 1 member A1 and high mobility group box 1 in oropharyngeal squamous cell carcinoma in association with survival time.

    PubMed

    Qian, Xu; Coordes, Annekatrin; Kaufmann, Andreas M; Albers, Andreas E

    2016-11-01

    Despite the development of novel multimodal treatment combinations in advanced oropharyngeal squamous cell carcinoma (OSCC), outcomes remain poor. The identification of specifically validated biomarkers is required to understand the underlying molecular mechanisms, to evaluate treatment efficiency and to develop novel therapeutic targets. The present study, therefore, examined the presence of aldehyde dehydrogenase family 1 member A1 (ALDH1A1) and high mobility group box 1 (HMGB1) expression in primary OSCC and analyzed the impact on survival time. In 59 patients with OSCC, the expression of ALDH1A1, p16 and HMGB1, and their clinicopathological data were analyzed. HMGB1 positivity was significantly increased in patients with T1-2 stage disease compared with T3-4 stage disease (P<0.001), whereas ALDH1A1 positivity was not. ALDH1A1(+) tumors showed significantly lower differentiation than ALDH1A1(-) tumors (P=0.018). Multivariate analysis showed that ALDH1A1 positivity (P=0.041) and nodal status (N2-3) (P=0.036) predicted a poor prognosis. In this patient cohort, ALDH1A1 and nodal status were identified as independent predictors of a shorter overall survival time. The study results, therefore, provide evidence of the prognostic value of ALDH1A1 as a marker for cancer stem cells and nodal status in OSCC patients.

  3. Enhancement of 1,3-propanediol production by expression of pyruvate decarboxylase and aldehyde dehydrogenase from Zymomonas mobilis in the acetolactate-synthase-deficient mutant of Klebsiella pneumoniae.

    PubMed

    Lee, Sung-Mok; Hong, Won-Kyung; Heo, Sun-Yeon; Park, Jang Min; Jung, You Ree; Oh, Baek-Rock; Joe, Min-Ho; Seo, Jeong-Woo; Kim, Chul Ho

    2014-08-01

    The acetolactate synthase (als)-deficient mutant of Klebsiella pneumoniae fails to produce 1,3-propanediol (1,3-PD) or 2,3-butanediol (2,3-BD), and is defective in glycerol metabolism. In an effort to recover production of the industrially valuable 1,3-PD, we introduced the Zymomonas mobilis pyruvate decarboxylase (pdc) and aldehyde dehydrogenase (aldB) genes into the als-deficient mutant to activate the conversion of pyruvate to ethanol. Heterologous expression of pdc and aldB efficiently recovered glycerol metabolism in the 2,3-BD synthesis-defective mutant, enhancing the production of 1,3-PD by preventing the accumulation of pyruvate. Production of 1,3-PD in the pdc- and aldB-expressing als-deficient mutant was further enhanced by increasing the aeration rate. This system uses metabolic engineering to produce 1,3-PD while minimizing the generation of 2,3-BD, offering a breakthrough for the industrial production of 1,3-PD from crude glycerol.

  4. Metabolism of trans, trans-muconaldehyde, a cytotoxic metabolite of benzene, in mouse liver by alcohol dehydrogenase Adh1 and aldehyde reductase AKR1A4

    SciTech Connect

    Short, Duncan M.; Lyon, Robert; Watson, David G.; Barski, Oleg A.; McGarvie, Gail; Ellis, Elizabeth M. . E-mail: Elizabeth.ellis@strath.ac.uk

    2006-01-15

    The reductive metabolism of trans, trans-muconaldehyde, a cytotoxic metabolite of benzene, was studied in mouse liver. Using an HPLC-based stopped assay, the primary reduced metabolite was identified as 6-hydroxy-trans, trans-2,4-hexadienal (OH/CHO) and the secondary metabolite as 1,6-dihydroxy-trans, trans-2,4-hexadiene (OH/OH). The main enzymes responsible for the highest levels of reductase activity towards trans, trans-muconaldehyde were purified from mouse liver soluble fraction first by Q-sepharose chromatography followed by either blue or red dye affinity chromatography. In mouse liver, trans, trans-muconaldehyde is predominantly reduced by an NADH-dependent enzyme, which was identified as alcohol dehydrogenase (Adh1). Kinetic constants obtained for trans, trans-muconaldehyde with the native Adh1 enzyme showed a V {sub max} of 2141 {+-} 500 nmol/min/mg and a K {sub m} of 11 {+-} 4 {mu}M. This enzyme was inhibited by pyrazole with a K {sub I} of 3.1 {+-} 0.57 {mu}M. Other fractions were found to contain muconaldehyde reductase activity independent of Adh1, and one enzyme was identified as the NADPH-dependent aldehyde reductase AKR1A4. This showed a V {sub max} of 115 nmol/min/mg and a K {sub m} of 15 {+-} 2 {mu}M and was not inhibited by pyrazole.

  5. Tryptophan in Alcoholism Treatment I:  Kynurenine Metabolites Inhibit the Rat Liver Mitochondrial Low Km Aldehyde Dehydrogenase Activity, Elevate Blood Acetaldehyde Concentration and Induce Aversion to Alcohol

    PubMed Central

    Badawy, Abdulla A.-B.; Bano, Samina; Steptoe, Alex

    2011-01-01

    Aims: The aims were to provide proofs of mechanism and principle by establishing the ability of kynurenine metabolites to inhibit the liver mitochondrial low Km aldehyde dehydrogenase (ALDH) activity after administration and in vivo, and to induce aversion to alcohol. Methods: Kynurenic acid (KA), 3-hydroxykynurenine (3-HK) and 3-hydroxyanthranilic acid (3-HAA) were administered to normal male Wistar rats and ALDH activity was determined both in vitro in liver homogenates and in vivo (by measuring blood acetaldehyde following ethanol administration). Alcohol consumption was studied in an aversion model in rats and in alcohol-preferring C57 mice. Results: ALDH activity was significantly inhibited by all three metabolites by doses as small as 1 mg/kg body wt. Blood acetaldehyde accumulation after ethanol administration was strongly elevated by KA and 3-HK and to a lesser extent by 3-HAA. All three metabolites induced aversion to alcohol in rats and decreased alcohol preference in mice. Conclusions: The above kynurenine metabolites of tryptophan induce aversion to alcohol by inhibiting ALDH activity. An intellectual property covering the use of 3-HK and 3-HAA and derivatives thereof in the treatment of alcoholism by aversion awaits further development. PMID:21896552

  6. (13) C magnetic resonance spectroscopic imaging of hyperpolarized [1-(13) C, U-(2) H5 ] ethanol oxidation can be used to assess aldehyde dehydrogenase activity in vivo.

    PubMed

    Dzien, Piotr; Kettunen, Mikko I; Marco-Rius, Irene; Serrao, Eva M; Rodrigues, Tiago B; Larkin, Timothy J; Timm, Kerstin N; Brindle, Kevin M

    2015-05-01

    Aldehyde dehydrogenase (ALDH2) is an emerging drug target for the treatment of heart disease, cocaine and alcohol dependence, and conditions caused by genetic polymorphisms in ALDH2. Noninvasive measurement of ALDH2 activity in vivo could inform the development of these drugs and accelerate their translation to the clinic. [1-(13) C, U-(2) H5 ] ethanol was hyperpolarized using dynamic nuclear polarization, injected into mice and its oxidation in the liver monitored using (13) C MR spectroscopy and spectroscopic imaging. Oxidation of [1-(13) C, U-(2) H5 ] ethanol to [1-(13) C] acetate was observed. Saturation of the acetaldehyde resonance, which was below the level of detection in vivo, demonstrated that acetate was produced via acetaldehyde. Irreversible inhibition of ALDH2 activity with disulfiram resulted in a proportional decrease in the amplitude of the acetate resonance. (13) C magnetic resonance spectroscopy measurements of hyperpolarized [1-(13) C, U-(2) H5 ] ethanol oxidation allow real-time assessment of ALDH2 activity in liver in vivo. © 2014 The Authors. Magnetic Resonance in Medicine Published by Wiley Periodicals, Inc. on behalf of International Society of Medicine in Resonance.

  7. [Overexpression of Spinacia oleracea betaine aldehyde dehydrogenase (SoBADH) gene confers the salt and cold tolerant in Gossypium hirsutum L].

    PubMed

    Luo, Xiaoli; Xiao, Juanli; Wang, Zhian; Zhang, Anhong; Tian, Yingchuan; Wu, Jiahe

    2008-08-01

    The open reading frame of Spinacia oleracea Betaine Aldehyde Dehydrogenase (SoBADH) was retrieved from Spinacia oleracea and inserted into the Agrobacterium tumefaciens binary vector pBin438, which was driven by CaMV35S promoter, and produced the new binary vector pBSB. A. tumefaciens LBA4404 carrying this plasmid was used in genetic transformation of plants. Forty-five primary transgenic plants were detected by PCR and verified by the Southern blotting from 65 regenerated plants, of which 27 transgenic plants had only one copy of T-DNA. The Northern blotting and Western blotting analysis indicated that the SoBADH gene had been transcribed mRNA and expression protein in the transgenic cotton lines. The testing of SoBADH activity of transgenic plant leaves showed that the enzyme activity was much higher than that of the non-transgenic cotton. The growth of transgenic plants was well under the salinity and freezing stress, whereas the non-transgenic plant grew poorly and even died. Challenging with salinity, the height and fresh weight of transgenic plants was higher compared with those of non-transgenic plants. Under the freezing stress, the relative conductivity of leaf electrolyte leakage of the transgenic cotton lines was lower than that of non-transgenic plants. These results demonstrated that the SoBADH gene could over express in the exogenous plants, and could be used in genetic engineering for cotton stress resistance.

  8. JWH-018 ω-OH, a shared hydroxy metabolite of the two synthetic cannabinoids JWH-018 and AM-2201, undergoes oxidation by alcohol dehydrogenase and aldehyde dehydrogenase enzymes in vitro forming the carboxylic acid metabolite.

    PubMed

    Holm, Niels Bjerre; Noble, Carolina; Linnet, Kristian

    2016-09-30

    Synthetic cannabinoids are new psychoactive substances (NPS) acting as agonists at the cannabinoid receptors. The aminoalkylindole-type synthetic cannabinoid naphthalen-1-yl-(1-pentylindol-3-yl)methanone (JWH-018) was among the first to appear on the illicit drug market and its metabolism has been extensively investigated. The N-pentyl side chain is a major site of human cytochrome P450 (CYP)-mediated oxidative metabolism, and the ω-carboxylic acid metabolite appears to be a major in vivo human urinary metabolite. This metabolite is, however, not formed to any significant extent in human liver microsomal (HLM) incubations raising the possibility that the discrepancy is due to involvement of cytosolic enzymes. Here we demonstrate in incubations with human liver cytosol (HLC), that JWH-018 ω-OH, but not the JWH-018 parent compound, is a substrate for nicotinamide adenine dinucleotide (NAD(+))-dependent alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) enzymes. The sole end-product identified in HLC was the JWH-018 ω-COOH metabolite, while trapping tests with methoxyamine proved the presence of the aldehyde intermediate. ADH/ALDH and UDP-glucuronosyl-transferases (UGT) enzymes may therefore both act on the JWH-018 ω-OH substrate. Finally, we note that for [1-(5-fluoropentyl)indol-3-yl]-naphthalen-1-yl-methanone (AM-2201), the ω-fluorinated analog of JWH-018, a high amount of JWH-018 ω-OH was formed in HLM incubated without NADPH, suggesting that the oxidative defluorination is efficiently catalyzed by non-CYP enzyme(s). The pathway presented here may therefore be especially important for N-(5-fluoropentyl) substituted synthetic cannabinoids, because the oxidative defluorination can occur even if the CYP-mediated metabolism preferentially takes place on other parts of the molecule than the N-alkyl side chain. Controlled clinical studies in humans are ultimately required to demonstrate the in vivo importance of the oxidation pathway presented here

  9. Susceptibility to lung cancer and genetic polymorphisms in the alcohol metabolite-related enzymes alcohol dehydrogenase 3, aldehyde dehydrogenase 2, and cytochrome P450 2E1 in the Japanese population.

    PubMed

    Minegishi, Yuji; Tsukino, Hiromasa; Muto, Manabu; Goto, Koichi; Gemma, Akihiko; Tsugane, Shoichiro; Kudoh, Shoji; Nishiwaki, Yutaka; Esumi, Hiroyasu

    2007-07-15

    It is believed that acetaldehyde plays an important role in alcohol-related carcinogenesis; although current epidemiologic studies have provided inconsistent findings on the association between alcohol consumption and the risk of lung cancer. To clarify the hypothesis that genetic polymorphisms in alcohol-metabolizing enzymes may influence susceptibility to lung cancer, the authors conducted a hospital-based case-control study and examined genetic polymorphisms in the alcohol dehydrogenase 3, aldehyde dehydrogenase 2 (ALDH(2)), and cytochrome P450 2E1 genes in 505 patients with histologically confirmed lung cancer and in a group of 256 noncancer controls who provided complete cigarette and alcohol consumption histories. Genotyping was conducted by polymerase chain reaction-restriction fragment-length polymorphism assay. A significant association was noted between alcohol consumption and lung cancer risk. Thus, using the median value for the controls as the cut-off point, the odds ratios (OR) for light and heavy drinkers were 1.76 and 1.95, respectively (P for trend = .012), compared with nondrinkers. In addition, there was a significant trend toward increased risk of lung cancer in drinkers with ALDH(2) variant alleles (P for trend <.0001). The adjusted OR for heavy drinkers was 6.15 compared with nondrinkers. Regarding associations between histologic type and genotypes, the ALDH(2) variant allele was significantly less common in patients who had adenocarcinoma compared with controls. The current observations suggested a positive association between alcohol consumption and the risk of lung cancer: Drinking may increase the risk, especially among individuals who have the variant ALDH(2) alleles.

  10. Interaction of the membrane-bound succinate dehydrogenase with substrate and competitive inhibitors.

    PubMed

    Kotlyar, A B; Vinogradov, A D

    1984-01-18

    The protective effect of dicarboxylates on the active-site-directed inhibition of the membrane-bound succinate dehydrogenase by N-ethylmaleimide, steady-state kinetics methods for Ki and Ks determinations, and equilibrium studies were employed to quantitate the relative affinities of succinate, fumarate, malonate and oxaloacetate to the reduced and oxidized species of the enzyme. A more than 10-fold difference in the relative affinities of the reduced and oxidized succinate dehydrogenase to succinate, fumarate and oxaloacetate is found, whereas the reactivity of the active-site sulphydryl group does not depend on the redox state of the enzyme. The redox-state-dependent changes in the affinity of the membrane-bound succinate dehydrogenase to oxaloacetate can be quantitatively accounted for by a 10-fold increase in the rate of dissociation of the enzyme-inhibitor complex which occurs upon reduction of the enzyme. The data obtained give no support for either the existence of a sulphydryl group other than the active-site one important for the catalysis or for the presence of a separate dicarboxylate-specific regulatory site in the succinate dehydrogenase molecule.

  11. Computational optimization of AG18051 inhibitor for amyloid-beta binding alcohol dehydrogenase enzyme

    NASA Astrophysics Data System (ADS)

    Marques, Alexandra T.; Antunes, Agostinho; Fernandes, Pedro A.; Ramos, Maria J.

    Amyloid-beta (Abeta) binding alcohol dehydrogenase (ABAD) is a multifunctional enzyme involved in maintaining the homeostasis. The enzyme can also mediate some diseases, including genetic diseases, Alzheimer's disease, and possibly some prostate cancers. Potent inhibitors of ABAD might facilitate a better clarification of the functions of the enzyme under normal and pathogenic conditions and might also be used for therapeutic intervention in disease conditions mediated by the enzyme. The AG18051 is the only presently available inhibitor of ABAD. It binds in the active-site cavity of the enzyme and reacts with the NAD+ cofactor to form a covalent adduct. In this work, we use computational methods to perform a rational optimization of the AG18051 inhibitor, through the introduction of chemical substitutions directed to improve the affinity of the inhibitor to the enzyme. The molecular mechanics-Poisson-Boltzmann surface area methodology was used to predict the relative free binding energy of the different modified inhibitor-NAD-enzyme complexes. We show that it is possible to increase significantly the affinity of the inhibitor to the enzyme with small modifications, without changing the overall structure and ADME (absorption, distribution, metabolism, and excretion) properties of the original inhibitor.

  12. A novel cofactor-binding mode in bacterial IMP dehydrogenases explains inhibitor selectivity

    SciTech Connect

    Makowska-Grzyska, Magdalena; Kim, Youngchang; Maltseva, Natalia; Osipiuk, Jerzy; Gu, Minyi; Zhang, Minjia; Mandapati, Kavitha; Gollapalli, Deviprasad R.; Gorla, Suresh Kumar; Hedstrom, Lizbeth; Joachimiak, Andrzej

    2015-01-09

    The steadily rising frequency of emerging diseases and antibiotic resistance creates an urgent need for new drugs and targets. Inosine 5'-monophosphate dehydrogenase (IMP dehydrogenase or IMPDH) is a promising target for the development of new antimicrobial agents. IMPDH catalyzes the oxidation of IMP to XMP with the concomitant reduction of NAD+, which is the pivotal step in the biosynthesis of guanine nucleotides. Potent inhibitors of bacterial IMPDHs have been identified that bind in a structurally distinct pocket that is absent in eukaryotic IMPDHs. The physiological role of this pocket was not understood. Here, we report the structures of complexes with different classes of inhibitors of Bacillus anthracis, Campylobacter jejuni, and Clostridium perfringens IMPDHs. These structures in combination with inhibition studies provide important insights into the interactions that modulate selectivity and potency. We also present two structures of the Vibrio cholerae IMPDH in complex with IMP/NAD+ and XMP/NAD+. In both structures, the cofactor assumes a dramatically different conformation than reported previously for eukaryotic IMPDHs and other dehydrogenases, with the major change observed for the position of the NAD+ adenosine moiety. More importantly, this new NAD+-binding site involves the same pocket that is utilized by the inhibitors. Thus, the bacterial IMPDH-specific NAD+-binding mode helps to rationalize the conformation adopted by several classes of prokaryotic IMPDH inhibitors. As a result, these findings offer a potential strategy for further ligand optimization.

  13. Immobilization of L-arabinitol dehydrogenase on aldehyde-functionalized silicon oxide nanoparticles for L-xylulose production.

    PubMed

    Singh, Raushan Kumar; Tiwari, Manish Kumar; Singh, Ranjitha; Haw, Jung-Rim; Lee, Jung-Kul

    2014-02-01

    L-Xylulose is a potential starting material for therapeutics. However, its translation into clinical practice has been hampered by its inherently low bioavailability. In addition, the high cost associated with the production of L-xylulose is a major factor hindering its rapid deployment beyond the laboratory. In the current study, L-arabinitol 4-dehydrogenase from Hypocrea jecorina (HjLAD), which catalyzes the conversion of L-arabinitol into L-xylulose, was immobilized onto various carriers, and the immobilized enzymes were characterized. HjLAD covalently immobilized onto silicon oxide nanoparticles showed the highest immobilization efficiency (94.7 %). This report presents a comparative characterization of free and immobilized HjLAD, including its thermostability and kinetic parameters. The thermostability of HjLAD immobilized on silicon oxide nanoparticles was more than 14.2-fold higher than free HjLAD; the t1/2 of HjLAD at 25 °C was enhanced from 190 min (free) to 45 h (immobilized). In addition, the immobilized HjLAD retained 94 % of its initial activity after 10 cycles. When the immobilized HjLAD was used to catalyze the biotransformation of L-arabinitol to L-xylulose, 66 % conversion and a productivity of 7.9 g · h(-1) · L(-1) were achieved. The enhanced thermostability and reusability of HjLAD suggest that immobilization of HjLAD onto silicon oxide nanoparticles has the potential for use in the industrial production of rare sugars.

  14. Discovery of Adamantyl Heterocyclic Ketones as Potent 11β-Hydroxysteroid Dehydrogenase Type 1 Inhibitors

    PubMed Central

    Su, Xiangdong; Vicker, Nigel; Thomas, Mark P; Pradaux-Caggiano, Fabienne; Halem, Heather; Culler, Michael D; Potter, Barry V L

    2011-01-01

    11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) plays a key role in converting intracellular cortisone to physiologically active cortisol, which is implicated in the development of several phenotypes of metabolic syndrome. Inhibition of 11β-HSD1 activity with selective inhibitors has beneficial effects on various conditions, including diabetes, dyslipidemia and obesity, and therefore constitutes a promising strategy to discover novel therapies for metabolic and cardiovascular diseases. A series of novel adamantyl heterocyclic ketones provides potent and selective inhibitors of human 11β-HSD1. Lead compounds display low nanomolar inhibition against human and mouse 11β-HSD1 and are selective with no activity against 11β-HSD2 and 17β-HSD1. Selected potent 11β-HSD1 inhibitors show moderate metabolic stability upon incubation with human liver microsomes and weak inhibition of human CYP450 enzymes. PMID:21608132

  15. The role of fluorine in stabilizing the bioactive conformation of dihydroorotate dehydrogenase inhibitors.

    PubMed

    Bonomo, Silvia; Tosco, Paolo; Giorgis, Marta; Lolli, Marco; Fruttero, Roberta

    2013-03-01

    Dihydroorotate dehydrogenase (DHODH) is an important drug target due to its prominent role in pyrimidine biosynthesis. Leflunomide and brequinar are two well-known DHODH inhibitors, which bind to the enzyme in the same pocket with different binding modes. We have recently realized a series of new inhibitors based on the 4-hydroxy-1,2,5-oxadiazole ring, whose activity profile was found to be closely dependent on the degree of fluorine substitution at the phenyl ring adjacent to the oxadiazole moiety; a positive influence of fluorine on the DHODH inhibitory potency was observed previously [Baumgartner et al. (2006) J Med Chem 49:1239-1247]. Potential energy surface scans showed that fluorine plays an important role in stabilizing the bioactive conformations; additionally, fluorine influences the balance between leflunomide-like and brequinar-like binding modes. These findings may serve as a guide to design more potent DHODH inhibitors.

  16. Discovery of adamantyl heterocyclic ketones as potent 11β-hydroxysteroid dehydrogenase type 1 inhibitors.

    PubMed

    Su, Xiangdong; Vicker, Nigel; Thomas, Mark P; Pradaux-Caggiano, Fabienne; Halem, Heather; Culler, Michael D; Potter, Barry V L

    2011-08-01

    11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) plays a key role in converting intracellular cortisone to physiologically active cortisol, which is implicated in the development of several phenotypes of metabolic syndrome. Inhibition of 11β-HSD1 activity with selective inhibitors has beneficial effects on various conditions, including diabetes, dyslipidemia and obesity, and therefore constitutes a promising strategy to discover novel therapies for metabolic and cardiovascular diseases. A series of novel adamantyl heterocyclic ketones provides potent and selective inhibitors of human 11β-HSD1. Lead compounds display low nanomolar inhibition against human and mouse 11β-HSD1 and are selective with no activity against 11β-HSD2 and 17β-HSD1. Selected potent 11β-HSD1 inhibitors show moderate metabolic stability upon incubation with human liver microsomes and weak inhibition of human CYP450 enzymes.

  17. Human hydroxysteroid dehydrogenases and pre-receptor regulation: Insights into inhibitor design and evaluation

    PubMed Central

    Penning, Trevor M.

    2011-01-01

    Hydroxysteroid dehydrogenases (HSDs) represent a major class of NAD(P)(H) dependent steroid hormone oxidoreductases involved in the pre-receptor regulation of hormone action. This is achieved by HSDs working in pairs so that they can interconvert ketosteroids with hydroxysteroids resulting in a change in ligand potency for nuclear receptors. HSDs belong to two protein superfamilies the aldo-keto reductases and the short-chain dehydrogenase/reductases. In humans, many of the important enzymes have been thoroughly characterized including the elucidation of their three-dimensional structures. Because these enzymes play fundamental roles in steroid hormone action they can be considered to be drug targets for a variety of steroid driven diseases: e.g. metabolic syndrome and obesity, inflammation, and hormone dependent malignancies of the endometrium, prostate and breast. This article will review how fundamental knowledge of these enzymes can be exploited in the development of isoform specific HSD inhibitors from both protein superfamilies. PMID:21272640

  18. The design of novel 17beta-hydroxysteroid dehydrogenase type 3 inhibitors.

    PubMed

    Vicker, Nigel; Sharland, Christopher M; Heaton, Wesley B; Gonzalez, Ana M Ramos; Bailey, Helen V; Smith, Andrew; Springall, Jeremy S; Day, Joanna M; Tutill, Helena J; Reed, Michael J; Purohit, Atul; Potter, Barry V L

    2009-03-25

    17beta-Hydroxysteroid dehydrogenase type 3 (17beta-HSD3) is expressed at high levels in the testes and seminal vesicles but has also been shown to be present in prostate tissue, suggesting its potential involvement in both gonadal and non-gonadal testosterone biosynthesis. The role of 17beta-HSD3 in testosterone biosynthesis makes this enzyme an attractive molecular target for small molecule inhibitors for the treatment of prostate cancer. Here we report the design of selective inhibitors of 17beta-HSD3 as potential anti-cancer agents. Due to 17beta-HSD3 being a membrane-bound protein a crystal structure is not yet available. A homology model of 17beta-HSD3 has been built to aid structure-based drug design. This model has been used with docking studies to identify a series of lead compounds that may give an insight as to how inhibitors interact with the active site. Compound 1 was identified as a potent selective inhibitor of 17beta-HSD3 with an IC(50)=700nM resulting in the discovery of a novel lead series for further optimisation. Using our homology model as a tool for inhibitor design compound 5 was discovered as a novel potent and selective inhibitor of 17beta-HSD3 with an IC(50) approximately 200nM.

  19. Identification of Small-Molecule Inhibitors against Meso-2, 6-Diaminopimelate Dehydrogenase from Porphyromonas gingivalis

    PubMed Central

    Stone, Victoria N.; Parikh, Hardik I.; El-rami, Fadi; Ge, Xiuchun; Chen, Weihau; Zhang, Yan; Kellogg, Glen E.; Xu, Ping

    2015-01-01

    Species-specific antimicrobial therapy has the potential to combat the increasing threat of antibiotic resistance and alteration of the human microbiome. We therefore set out to demonstrate the beginning of a pathogen-selective drug discovery method using the periodontal pathogen Porphyromonas gingivalis as a model. Through our knowledge of metabolic networks and essential genes we identified a “druggable” essential target, meso-diaminopimelate dehydrogenase, which is found in a limited number of species. We adopted a high-throughput virtual screen method on the ZINC chemical library to select a group of potential small-molecule inhibitors. Meso-diaminopimelate dehydrogenase from P. gingivalis was first expressed and purified in Escherichia coli then characterized for enzymatic inhibitor screening studies. Several inhibitors with similar structural scaffolds containing a sulfonamide core and aromatic substituents showed dose-dependent inhibition. These compounds were further assayed showing reasonable whole-cell activity and the inhibition mechanism was determined. We conclude that the establishment of this target and screening strategy provides a model for the future development of new antimicrobials. PMID:26544875

  20. Nicotinoprotein (NADH-containing) alcohol dehydrogenase from Rhodococcus erythropolis DSM 1069: an efficient catalyst for coenzyme-independent oxidation of a broad spectrum of alcohols and the interconversion of alcohols and aldehydes.

    PubMed

    Schenkels, P; Duine, J A

    2000-04-01

    Extracts from benzyl-alcohol-grown Rhodococcus erythropolis DSM 1069 showed NAD(P)-independent, N,N-dimethyl-4-nitrosoaniline (NDMA)-dependent alcohol dehydrogenase activity. The enzyme exhibiting this activity was purified to homogeneity and characterized. It appears to be a typical nicotinoprotein as it contains tightly bound NADH acting as cofactor instead of coenzyme. Other characteristics indicate that it is highly similar to the known nicotinoprotein alcohol dehydrogenase (np-ADH) from Amycolatopsis methanolica: it is a homotetramer of 150 kDa; N-terminal amino acid sequencing (22 residues) showed that 77% of these amino acids are identical in the two enzymes; it has optimal activity at pH 7.0; it lacks NAD(P)H-dependent aldehyde reductase activity; it catalyses the oxidation of a broad range of (preferably) primary and secondary alcohols, either aliphatic or aromatic, and formaldehyde, with the concomitant reduction of the artificial electron acceptor NDMA. NDMA could be replaced by an aldehyde, but not formaldehyde, the substrate specificity of the enzyme for the aldehydes reflecting that for the corresponding alcohols. The latter also applied to the low aldehyde dismutase activity displayed by the enzyme. From this, together with the results of the induction studies, it is concluded that np-ADH functions as the main alcohol-oxidizing enzyme in the dissimilation of many, but not all, alcohols by R. erythropolis and may also catalyse coenzyme-independent interconversion of alcohols and aldehydes under certain circumstances. It is anticipated that the enzyme may be of even wider significance since structural data indicate that np-ADH is also present in other (nocardioform) actinomycetes.

  1. Aldehyde dehydrogenase 1A1 up-regulates stem cell markers in benzo[a]pyrene-induced malignant transformation of BEAS-2B cells.

    PubMed

    Liu, Yonghong; Lu, Ruitao; Gu, Junlian; Chen, Yanxuan; Zhang, Xueyan; Zhang, Lan; Wu, Hao; Hua, Wenfeng; Zeng, Jun

    2016-07-01

    Recently, Aldehyde dehydrogenase 1A1 (ALDH1A1) has been proposed to be a common marker of cancer stem cells and can be induced by benzo[a]pyrene (B[a]P) exposure. However, the underlying mechanism of how ALDH1A1 contributes to B[a]P-induced carcinogenesis in human bronchial epithelial cells remains unclear. Here, we found that B[a]P up-regulated expression levels of stem cell markers (ABCG2, SOX2, c-Myc and Klf4), epithelial-mesenchymal transition (EMT) associated genes (SNAIL1, ZEB1, TWIST and β-CATENIN) and cancer-related long non-coding RNAs (lncRNAs; HOTAIR and MALAT-1) in malignant B[a]P-transformed human bronchial epithelial cells (BEAS-2B-T cells), and these up-regulations were dependent on increased expression of ALDH1A1. The inhibition of endogenous ALDH1A1 expression down-regulated expression levels of stem cell markers and reversed the malignant phenotype as well as reduced the chemoresistance of BEAS-2B-T cells. In contrast, the overexpression of ALDH1A1 in BEAS-2B cells increased the expression of stem cell markers, facilitated cell transformation, promoted migratory ability and enhanced the drug resistance of BEAS-2B cells. Overall, our data indicates that ALDH1A1 promotes a stemness phenotype and plays a critical role in the BEAS-2B cell malignant transformation induced by B[a]P.

  2. Crosstalk-eliminated quantitative determination of aflatoxin B1-induced hepatocellular cancer stem cells based on concurrent monitoring of CD133, CD44, and aldehyde dehydrogenase1.

    PubMed

    Ju, Hee; Shim, Yumi; Arumugam, Parthasarathy; Song, Joon Myong

    2016-01-22

    Cancer stem cells (CSCs), known as tumor initiating cells, have become a critically important issue for cancer therapy. Although much research has demonstrated the induction of hepato cellular carcinoma by aflatoxin B1, the formation of hepatocellular CSCs and their quantitative determination is hardly reported. In this work, it was found that hepatocellular CSCs were produced from HepG2 cells by aflatoxin B1-induced mutation, and their amount was quantitatively determined using crosstalk-eliminated multicolor cellular imaging based on quantum dot (Qdot) nanoprobes and an acousto-optical tunable filter (AOTF). Hepatocellular CSCs were acquired via magnetic bead-based sorting and observed using concurrent detection of three different markers: CD133, CD44, and aldehyde dehydrogenase1 (ALDH1). The DNA mutation of HepG2 cells caused by aflatoxin B1 was quantitatively observed via absorbance spectra of aflatoxin B1-8, 9-epoxide-DNA adducts. The percentages of hepatocellular CSCs formed in the entire HepG2 cells were determined to be 9.77±0.65%, 10.9±1.39%, 11.4±1.32%, and 12.8±0.7%, respectively, at 0 μM, 5 μM, 10 μM, and 20 μM of aflatoxin B1. The results matched well with those obtained utilizing flow cytometry. This study demonstrates that aflatoxin mediated mutation induced the conversion of hepatic cancer cell to hepatic CSCs by using a Qdot based constructed multicolor cellular imaging system.

  3. Cancer screening of upper aerodigestive tract in Japanese alcoholics with reference to drinking and smoking habits and aldehyde dehydrogenase-2 genotype.

    PubMed

    Yokoyama, A; Ohmori, T; Muramatsu, T; Higuchi, S; Yokoyama, T; Matsushita, S; Matsumoto, M; Maruyama, K; Hayashida, M; Ishii, H

    1996-11-04

    In this study, 1,000 Japanese male alcoholics were consecutively screened by upper gastrointestinal endoscopy with esophageal iodine staining. Associations among cancer-detection rates, drinking and smoking habits, and aldehyde dehydrogenase-2 (ALDH2) genotypes were evaluated. A total of 53 patients (5.3%) had histologically confirmed cancer. Esophageal cancer was diagnosed in 36, gastric cancer in 17, and oropharyngolaryngeal cancer in 9 patients: 8 of the esophageal-cancer patients were multiple-cancer patients, with additional cancer(s) in the stomach and/or oropharyngolaryngeal region. Multiple logistic regression revealed that use of stronger alcoholic beverages (whisky or shochu) in contrast with lighter beverages (sake or beer) and smoking of 50 pack-years or more increased the risks for esophageal (odds ratio 3.2 and 2.8 respectively), oropharyngolaryngeal (4.8 and 5.1 respectively) and multiple cancer (10.5 and 11.8 respectively). The inactive form of ALDH2, encoded by the gene ALDH2*1/2*2 prevalent in Orientals, exposes them to higher blood levels of acetaldehyde, a recognized animal carcinogen, after drinking. This inactive ALDH2 was detected in 19/36 (52.8%) patients with esophageal cancer, in 5/9 (55.6%) patients with oropharyngolaryngeal cancer, and in 7/8 (87.5%) patients with multiple cancer. All of these gene frequencies far exceeded that in a large alcoholic cohort (80/655, 12.2%). The triple combination of the risk factors of the inactive ALDH2, stronger alcoholic beverages and heavy smoking was more commonly associated with multiple-cancer patients than with patients with esophageal cancer alone (62.5% vs. 7.1%). These results show that the 3 risk factors are important for the development of upper-aerodigestive-tract cancer in Japanese alcoholics. For these high-risk drinkers, regimented screening appears to be indicated.

  4. Long-term inhibition of ethanol intake by the administration of an aldehyde dehydrogenase-2 (ALDH2)-coding lentiviral vector into the ventral tegmental area of rats.

    PubMed

    Karahanian, Eduardo; Rivera-Meza, Mario; Tampier, Lutske; Quintanilla, María Elena; Herrera-Marschitz, Mario; Israel, Yedy

    2015-03-01

    Previous studies suggest that acetaldehyde generated from ethanol in the brain is reinforcing. The present studies tested the feasibility of achieving a long-term reduction of chronic and post-deprivation binge ethanol drinking by a single administration into the brain ventral tegmental area (VTA) of a lentiviral vector that codes for aldehyde dehydrogenase-2 (ALDH2), which degrades acetaldehyde. The ALDH2 gene coding vector or a control lentiviral vector were microinjected into the VTA of rats bred for their alcohol preference. In the chronic alcohol administration model, naïve animals administered the control vector and subsequently offered 10% ethanol and water ingested 8-9 g ethanol/kg body weight/day. The single administration of the ALDH2-coding vector prior to allowing ethanol availability reduced ethanol drinking by 85-90% (P < 0.001) for the 45 days tested. In the post-deprivation binge-drinking model, animals that had previously consumed ethanol chronically for 81 days were administered the lentiviral vector and were thereafter deprived of ethanol for three 7-day periods, each interrupted by a single 60-minute ethanol re-access after the last day of each deprivation period. Upon ethanol re-access, control vector-treated animals consumed intoxicating 'binge' amounts of ethanol, reaching intakes of 2.7 g ethanol/kg body weight in 60 minutes. The administration of the ALDH2-coding vector reduced re-access binge drinking by 75-80% (P < 0.001). This study shows that endowing the ventral tegmental with an increased ability to degrade acetaldehyde greatly reduces chronic alcohol consumption and post-deprivation binge drinking for prolonged periods and supports the hypothesis that brain-generated acetaldehyde promotes alcohol drinking. © 2014 Society for the Study of Addiction.

  5. Essential role of aldehyde dehydrogenase 1A3 for the maintenance of non-small cell lung cancer stem cells is associated with the STAT3 pathway.

    PubMed

    Shao, Chunli; Sullivan, James P; Girard, Luc; Augustyn, Alexander; Yenerall, Paul; Rodriguez-Canales, Jaime; Liu, Hui; Behrens, Carmen; Shay, Jerry W; Wistuba, Ignacio I; Minna, John D

    2014-08-01

    Lung cancer stem cells (CSC) with elevated aldehyde dehydrogenase (ALDH) activity are self-renewing, clonogenic, and tumorigenic. The purpose of our study is to elucidate the mechanisms by which lung CSCs are regulated. A genome-wide gene expression analysis was performed to identify genes differentially expressed in the ALDH(+) versus ALDH -: cells. RT-PCR, Western blot analysis, and Aldefluor assay were used to validate identified genes. To explore the function in CSCs, we manipulated their expression followed by colony and tumor formation assays. We identified a subset of genes that were differentially expressed in common in ALDH(+) cells, among which ALDH1A3 was the most upregulated gene in ALDH(+) versus ALDH -: cells. shRNA-mediated knockdown of ALDH1A3 in non-small cell lung cancer (NSCLC) resulted in a dramatic reduction in ALDH activity, clonogenicity, and tumorigenicity, indicating that ALDH1A3 is required for tumorigenic properties. In contrast, overexpression of ALDH1A3 by itself it was not sufficient to increase tumorigenicity. The ALDH(+) cells also expressed more activated STAT3 than ALDH -: cells. Inhibition of STAT3 or its activator EZH2 genetically or pharmacologically diminished the level of ALDH(+) cells and clonogenicity. Unexpectedly, ALDH1A3 was highly expressed in female, never smokers, well-differentiated tumors, or adenocarcinoma. ALDH1A3 low expression was associated with poor overall survival. Our data show that ALDH1A3 is the predominant ALDH isozyme responsible for ALDH activity and tumorigenicity in most NSCLCs, and that inhibiting either ALDH1A3 or the STAT3 pathway are potential therapeutic strategies to eliminate the ALDH(+) subpopulation in NSCLCs. ©2014 American Association for Cancer Research.

  6. Immobilisation and characterisation of biocatalytic co-factor recycling enzymes, glucose dehydrogenase and NADH oxidase, on aldehyde functional ReSyn™ polymer microspheres.

    PubMed

    Twala, Busisiwe V; Sewell, B Trevor; Jordaan, Justin

    2012-05-10

    The use of enzymes in industrial applications is limited by their instability, cost and difficulty in their recovery and re-use. Immobilisation is a technique which has been shown to alleviate these limitations in biocatalysis. Here we describe the immobilisation of two biocatalytically relevant co-factor recycling enzymes, glucose dehydrogenase (GDH) and NADH oxidase (NOD) on aldehyde functional ReSyn™ polymer microspheres with varying functional group densities. The successful immobilisation of the enzymes on this new high capacity microsphere technology resulted in the maintenance of activity of ∼40% for GDH and a maximum of 15.4% for NOD. The microsphere variant with highest functional group density of ∼3500 μmol g⁻¹ displayed the highest specific activity for the immobilisation of both enzymes at 33.22 U mg⁻¹ and 6.75 U mg⁻¹ for GDH and NOD with respective loading capacities of 51% (0.51 mg mg⁻¹) and 129% (1.29 mg mg⁻¹). The immobilised GDH further displayed improved activity in the acidic pH range. Both enzymes displayed improved pH and thermal stability with the most pronounced thermal stability for GDH displayed on ReSyn™ A during temperature incubation at 65 °C with a 13.59 fold increase, and NOD with a 2.25-fold improvement at 45 °C on the same microsphere variant. An important finding is the suitability of the microspheres for stabilisation of the multimeric protein GDH.

  7. Comparison of quantum dot technology with conventional immunohistochemistry in examining aldehyde dehydrogenase 1A1 as a potential biomarker for lymph node metastasis of head and neck cancer.

    PubMed

    Xu, Jing; Müller, Susan; Nannapaneni, Sreenivas; Pan, Lin; Wang, Yuxiang; Peng, Xianghong; Wang, Dongsheng; Tighiouart, Mourad; Chen, Zhengjia; Saba, Nabil F; Beitler, Jonathan J; Shin, Dong M; Chen, Zhuo Georgia

    2012-07-01

    This study explored whether the expression of aldehyde dehydrogenase 1 (ALDH1A1) in the primary tumour correlated with lymph node metastasis (LNM) of squamous cell carcinoma of the head and neck (HNSCC). We used both quantum dot (QD)-based immunohistofluorescence (IHF) and conventional immunohistochemistry (IHC) to quantify ALDH1A1 expression in primary tumour samples taken from 96 HNSCC patients, 50 with disease in the lymph nodes and 46 without. The correlation between the quantified level of ALDH1A1 expression and LNM in HNSCC patients was evaluated with univariate and multivariate analysis. The prognostic value of ALDH1A1 was examined by Kaplan-Meier analysis and Wald test. ALDH1A1 was highly correlated with LNM in HNSCC patients (p<0.0001 by QD-based IHF and 0.039 by IHC). The two methods (QD-based IHF and conventional IHC) for quantification of ALDH1A1 were found to be comparable (R=0.75, p<0.0001), but QD-IHF was more sensitive and objective than IHC. The HNSCC patients with low ALDH1A1 expression had a higher 5-year survival rate than those with high ALDH1A1 level (p=0.025). Our study suggests that ALDH1A1 is a potential biomarker for predicting LNM in HNSCC patients, though it is not an independent prognostic factor for survival of HNSCC patients. Furthermore, QD-IHF has advantages over IHC in quantification of ALDH1A1 expression in HNSCC tissues.

  8. Structural and functional consequences of coenzyme binding to the inactive asian variant of mitochondrial aldehyde dehydrogenase: roles of residues 475 and 487.

    PubMed

    Larson, Heather N; Zhou, Jianzhong; Chen, Zhiqiang; Stamler, Jonathan S; Weiner, Henry; Hurley, Thomas D

    2007-04-27

    The common mitochondrial aldehyde dehydrogenase (ALDH2) ALDH2(*)2 polymorphism is associated with impaired ethanol metabolism and decreased efficacy of nitroglycerin treatment. These physiological effects are due to the substitution of Lys for Glu-487 that reduces the k(cat) for these processes and increases the K(m) for NAD(+), as compared with ALDH2. In this study, we sought to understand the nature of the interactions that give rise to the loss of structural integrity and low activity in ALDH2(*)2 even when complexed with coenzyme. Consequently, we have solved the crystal structure of ALDH2(*)2 complexed with coenzyme to 2.5A(.) We have also solved the structures of a mutated form of ALDH2 where Arg-475 is replaced by Gln (R475Q). The structural and functional properties of the R475Q enzyme are intermediate between those of wild-type and the ALDH2(*)2 enzymes. In both cases, the binding of coenzyme restores most of the structural deficits observed in the apoenzyme structures. The binding of coenzyme to the R475Q enzyme restores its structure and catalytic properties to near wild-type levels. In contrast, the disordered helix within the coenzyme binding pocket of ALDH2(*)2 is reordered, but the active site is only partially reordered. Consistent with the structural data, ALDH2(*)2 showed a concentration-dependent increase in esterase activity and nitroglycerin reductase activity upon addition of coenzyme, but the levels of activity do not approach those of the wild-type enzyme or that of the R475Q enzyme. The data presented shows that Glu-487 maintains a critical function in linking the structure of the coenzyme-binding site to that of the active site through its interactions with Arg-264 and Arg-475, and in doing so, creates the stable structural scaffold conducive to catalysis.

  9. Structural and Functional Consequences of Coenzyme Binding to the Inactive Asian Variant of Mitochondrial Aldehyde Dehydrogenase: Roles of Residues 475 and 487

    SciTech Connect

    Larson,H.; Zhou, J.; Chen, Z.; Stamler, J.; Weiner, H.; Hurley, T.

    2007-01-01

    The common mitochondrial aldehyde dehydrogenase (ALDH2) ALDH2*2 polymorphism is associated with impaired ethanol metabolism and decreased efficacy of nitroglycerin treatment. These physiological effects are due to the substitution of Lys for Glu-487 that reduces the k{sub cat} for these processes and increases the K{sub m} for NAD{sup +}, as compared with ALDH2. In this study, we sought to understand the nature of the interactions that give rise to the loss of structural integrity and low activity in ALDH2*2 even when complexed with coenzyme. Consequently, we have solved the crystal structure of ALDH2*2 complexed with coenzyme to 2.5 {angstrom}. We have also solved the structures of a mutated form of ALDH2 where Arg-475 is replaced by Gln (R475Q). The structural and functional properties of the R475Q enzyme are intermediate between those of wild-type and the ALDH2*2 enzymes. In both cases, the binding of coenzyme restores most of the structural deficits observed in the apoenzyme structures. The binding of coenzyme to the R475Q enzyme restores its structure and catalytic properties to near wild-type levels. In contrast, the disordered helix within the coenzyme binding pocket of ALDH2*2 is reordered, but the active site is only partially reordered. Consistent with the structural data, ALDH2*2 showed a concentration-dependent increase in esterase activity and nitroglycerin reductase activity upon addition of coenzyme, but the levels of activity do not approach those of the wild-type enzyme or that of the R475Q enzyme. The data presented shows that Glu-487 maintains a critical function in linking the structure of the coenzyme binding site to that of the active site through its interactions with Arg-264 and Arg-475, and in doing so, creates the stable structural scaffold conducive to catalysis.

  10. Effects of Betaine Aldehyde Dehydrogenase-Transgenic Soybean on Phosphatase Activities and Rhizospheric Bacterial Community of the Saline-Alkali Soil

    PubMed Central

    Wang, Da-qing; Yu, Song

    2016-01-01

    The development of transgenic soybean has produced numerous economic benefits; however the potential impact of root exudates upon soil ecological systems and rhizospheric soil microbial diversity has also received intensive attention. In the present study, the influence of saline-alkali tolerant transgenic soybean of betaine aldehyde dehydrogenase on bacterial community structure and soil phosphatase during growth stages was investigated. The results showed that, compared with nontransgenic soybean as a control, the rhizospheric soil pH of transgenic soybean significantly decreased at the seedling stage. Compared to HN35, organic P content was 13.5% and 25.4% greater at the pod-filling stage and maturity, respectively. The acid phosphatase activity of SRTS was significantly better than HN35 by 12.74% at seedling, 14.03% at flowering, and 59.29% at podding, while alkaline phosphatase achieved maximum activity in the flowering stage and was markedly lower than HN35 by 13.25% at pod-filling. The 454 pyrosequencing technique was employed to investigate bacterial diversity, with a total of 25,499 operational taxonomic units (OTUs) obtained from the 10 samples. Notably, the effect of SRTS on microbial richness and diversity of rhizospheric soil was marked at the stage of podding and pod-filling. Proteobacteria, Acidobacteria, and Actinobacteria were the dominant phyla among all samples. Compared with HN35, the relative abundance of Proteobacteria was lower by 2.01%, 2.06%, and 5.28% at the stage of seedling, at pod-bearing, and at maturity. In genus level, the relative abundance of Gp6, Sphingomonas sp., and GP4 was significantly inhibited by SRTS at the stage of pod-bearing and pod-filling. PMID:27689079

  11. Association between Carotid Intima-media Thickness and Aldehyde Dehydrogenase 2 Glu504Lys Polymorphism in Chinese Han with Essential Hypertension

    PubMed Central

    Ma, Xiao-Xiang; Zheng, Shu-Zhan; Shu, Yan; Wang, Yong; Chen, Xiao-Ping

    2016-01-01

    Background: Aldehyde dehydrogenase 2 (ALDH2) is involved in the pathophysiological processes of cardiovascular diseases. Recent studies showed that mutant ALDH2 could increase oxidative stress and is a susceptible factor for hypertension. In addition, wild-type ALDH2 could improve the endothelial functions, therefore reducing the risk of developing atherosclerosis. The aim of the present study was to explore the frequency of the Glu504Lys polymorphism of the ALDH2 gene and its relation to carotid intima-media thickness (CIMT) in a group of patients with essential hypertension (EH) and to investigate the association between the Glu504Lys polymorphism and CIMT in Chinese Han patients with EH. Methods: In this study, 410 Chinese Han patients with EH who received physical examinations at the People's Hospital of Sichuan Province (China) were selected. DNA microarray chip was used for the genotyping of the Glu504Lys polymorphism of the ALDH2 gene. The differences in CIMT among patients with different Glu504Lys ALDH2 genotypes were analyzed. Results: The mean CIMT of the patients carrying AA/AG and GG genotypes was 1.02 ± 0.31 mm and 0.78 ± 0.28 mm, respectively. One-way ANOVA showed that the CIMT of the patients carrying the AA/AG genotype was significantly higher than in the ones carrying the GG genotype (P < 0.001). Multivariate logistic regression showed that the Glu504Lys AA/AG genotype of the ALDH2 gene was one of the major factors influencing the CIMT in patients with EH (odds ratio = 3.731, 95% confidence interval = 1.589–8.124, P = 0.001). Conclusions: The Glu504Lys polymorphism of the ALDH2 gene is associated with the CIMT of Chinese Han patients with EH in Sichuan, China. PMID:27270535

  12. Identification of 2-amino-5-aryl-pyrazines as inhibitors of human lactate dehydrogenase.

    PubMed

    Fauber, Benjamin P; Dragovich, Peter S; Chen, Jinhua; Corson, Laura B; Ding, Charles Z; Eigenbrot, Charles; Giannetti, Anthony M; Hunsaker, Thomas; Labadie, Sharada; Liu, Yichin; Liu, Yingchun; Malek, Shiva; Peterson, David; Pitts, Keith; Sideris, Steve; Ultsch, Mark; VanderPorten, Erica; Wang, Jing; Wei, BinQing; Yen, Ivana; Yue, Qin

    2013-10-15

    A 2-amino-5-aryl-pyrazine was identified as an inhibitor of human lactate dehydrogenase A (LDHA) via a biochemical screening campaign. Biochemical and biophysical experiments demonstrated that the compound specifically interacted with human LDHA. Structural variation of the screening hit resulted in improvements in LDHA biochemical inhibition and pharmacokinetic properties. A crystal structure of an improved compound bound to human LDHA was also obtained and it explained many of the observed structure-activity relationships. Copyright © 2013 Elsevier Ltd. All rights reserved.

  13. Endogenous selective inhibitors of 11beta-hydroxysteroid dehydrogenase isoforms 1 and 2 of adrenal origin.

    PubMed

    Latif, Syed A; Pardo, Hector A; Hardy, Matthew P; Morris, David J

    2005-11-24

    In earlier studies [Latif, S.A., Sheff, M.F., Ribeiro, C.E., Morris, D.J., 1997. Selective inhibition of sheep kidney 11beta-hydroxysteroid-dehydrogenase isoform 2 activity by 5alpha-reduced (but not 5beta) derivatives of adrenocorticosteroids. Steroids 62, 230-237], only derivatives of steroid hormones possessing the 5alpha-Ring A-reduced configuration selectively inhibited 11beta-HSD2-dehydrogenase, whereas their 5beta-derivatives were inactive. This present study focuses on an expanded group of endogenous 11-oxygenated, 5alpha and 5beta-Ring A-reduced metabolites of adrenocorticosteroids, and progestogen and androgen steroid hormones. These substances were tested for their inhibitory properties against 11beta-HSD2, 11beta-HSD1-dehydrogenase and 11beta-HSD1 reductase. The present studies showed that the following compounds stand out as potent inhibitors. These are 5alpha-DH-corticosterone, 3alpha,5alpha-TH-corticosterone, 11beta-OH-progesterone, 11beta-OH-allopregnanolone, 11beta-OH-testosterone, and 11beta-OH-androstanediol, inhibitors of 11beta-HSD1-dehydrogenase; 3alpha,5alpha-TH-11-dehydro-corticosterone, 11-keto-progesterone, 11-keto-allopregnanolone, and 11-keto-3beta,5alpha-TH-testosterone, inhibitors of 11beta-HSD1 reductase; 3alpha,5alpha-TH-aldosterone, 5alpha-DH-corticosterone, 3alpha,5alpha-TH-corticosterone,11-dehydro-corticosterone, 3alpha,5alpha-TH-11-dehydro-corticosterone, 11beta-OH-progesterone, 11-keto-progesterone, 11beta-OH-allopregnanolone, 11-keto-allopregnanolone, 11beta-OH-testosterone, and 11-keto-testosterone, inhibitors of 11beta-HSD2. All of these substances have the potential to be derived from adrenally synthesized corticosteroids. Substances with similar structures to those described may help in the design of exogenous agents for the management of a variety of disease states involving 11beta-HSD isoenzymes.

  14. A new potent inhibitor of horse liver alcohol dehydrogenase: p-methylbenzyl hydroperoxide.

    PubMed

    Skurský, L; Khan, A N; Saleem, M N; al-Tamer, Y Y

    1992-04-01

    A product of p-xylene auto-oxidation, p-methylbenzyl hydroperoxide, acts as a very strong reversible inhibitor of the ethanol dehydrogenating activity of horse liver alcohol dehydrogenase. Concentrations of hydroperoxide as low as that of the enzyme active site (about 10(-8) mol.dm-3) in the assay depresses the activity by 50%. Somewhat less potent is benzyl hydroperoxide (derived from toluene) while the (secondary) hydroperoxide derived from ethylbenzene and tert.butyl hydroperoxide and cumyl hydroperoxide do not inhibit HLAD appreciably.

  15. Identification of 3,6-disubstituted dihydropyrones as