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Sample records for aldehyde dehydrogenase activity

  1. Detection of aldehyde dehydrogenase activity in human corneal extracts

    NARCIS (Netherlands)

    Gondhowiardjo, T. D.; van Haeringen, N. J.; Hoekzema, R.; Pels, L.; Kijlstra, A.

    1991-01-01

    The major soluble protein in bovine corneal epithelial extracts is a 54 kD protein (BCP 54) which has recently been identified as the corneal aldehyde dehydrogenase. Although ALDH activity has been reported in human corneal extracts it was not yet clear whether this was identical with the 54 kD

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  4. Correlation of loss of activity of human aldehyde dehydrogenase with reaction of bromoacetophenone with glutamic acid-268 and cysteine-302 residues. Partial-sites reactivity of aldehyde dehydrogenase.

    Science.gov (United States)

    Abriola, D P; MacKerell, A D; Pietruszko, R

    1990-01-01

    Bromoacetophenone (2-bromo-1-phenylethanone) has been characterized as an affinity reagent for human aldehyde dehydrogenase (EC 1.2.1.3) [MacKerell, MacWright & Pietruszko (1986) Biochemistry 25, 5182-5189], and has been shown to react specifically with the Glu-268 residue [Abriola, Fields, Stein, MacKerell & Pietruszko (1987) Biochemistry 26, 5679-5684] with an apparent inactivation stoichiometry of two molecules of bromoacetophenone per molecule of enzyme. The specificity of bromoacetophenone for reaction with Glu-268, however, is not absolute, owing to the extreme reactivity of this reagent. When bromo[14C]acetophenone was used to label the human cytoplasmic E1 isoenzyme radioactively and tryptic fragmentation was carried out, peptides besides that containing Glu-268 were found to have reacted with reagent. These peptides were purified by h.p.l.c. and analysed by sequencing and scintillation counting to quantify radioactive label in the material from each cycle of sequencing. Reaction of bromoacetophenone with the aldehyde dehydrogenase molecule during enzyme activity loss occurs with two residues, Glu-268 and Cys-302. The activity loss, however, appears to be proportional to incorporation of label at Glu-268. The large part of incorporation of label at Cys-302 occurs after the activity loss is essentially complete. With both Glu-268 and Cys-302, however, the incorporation of label stops after one molecule of bromoacetophenone has reacted with each residue. Reaction with other residues continues after activity loss is complete. PMID:1968743

  5. Aldehyde dehydrogenases and cell proliferation.

    Science.gov (United States)

    Muzio, G; Maggiora, M; Paiuzzi, E; Oraldi, M; Canuto, R A

    2012-02-15

    Aldehyde dehydrogenases (ALDHs) oxidize aldehydes to the corresponding carboxylic acids using either NAD or NADP as a coenzyme. Aldehydes are highly reactive aliphatic or aromatic molecules that play an important role in numerous physiological, pathological, and pharmacological processes. ALDHs have been discovered in practically all organisms and there are multiple isoforms, with multiple subcellular localizations. More than 160 ALDH cDNAs or genes have been isolated and sequenced to date from various sources, including bacteria, yeast, fungi, plants, and animals. The eukaryote ALDH genes can be subdivided into several families; the human genome contains 19 known ALDH genes, as well as many pseudogenes. Noteworthy is the fact that elevated activity of various ALDHs, namely ALDH1A2, ALDH1A3, ALDH1A7, ALDH2*2, ALDH3A1, ALDH4A1, ALDH5A1, ALDH6, and ALDH9A1, has been observed in normal and cancer stem cells. Consequently, ALDHs not only may be considered markers of these cells, but also may well play a functional role in terms of self-protection, differentiation, and/or expansion of stem cell populations. The ALDH3 family includes enzymes able to oxidize medium-chain aliphatic and aromatic aldehydes, such as peroxidic and fatty aldehydes. Moreover, these enzymes also have noncatalytic functions, including antioxidant functions and some structural roles. The gene of the cytosolic form, ALDH3A1, is localized on chromosome 17 in human beings and on the 11th and 10th chromosome in the mouse and rat, respectively. ALDH3A1 belongs to the phase II group of drug-metabolizing enzymes and is highly expressed in the stomach, lung, keratinocytes, and cornea, but poorly, if at all, in normal liver. Cytosolic ALDH3 is induced by polycyclic aromatic hydrocarbons or chlorinated compounds, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin, in rat liver cells and increases during carcinogenesis. It has been observed that this increased activity is directly correlated with the degree of

  6. The Diagnostic Significance of Serum Alcohol Dehydrogenase Isoenzymes and Aldehyde Dehydrogenase Activity in Urinary Bladder Cancer Patients.

    Science.gov (United States)

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

    2017-07-01

    The aim of this study was to investigate a potential role of alcohol dehydrogenase and aldehyde dehydrogenase as tumor markers for urinary bladder cancer. Serum samples were obtained from 41 patients with bladder cancer and 52 healthy individuals. Class III and IV of ADH and total ADH activity were measured by the photometric method. For measurement of class I and II ADH and ALDH activity, the fluorometric method was employed. Significantly higher total activity of ADH was found in sera of both, low-grade and high-grade bladder cancer patients. The diagnostic sensitivity for total ADH activity was 81.5%, specificity 98.1%, positive (PPV) and negative (NPV) predictive values were 97.4% and 92.3% respectively. Area under ROC curve for total ADH activity was 0.848. A potential role of total ADH activity as a marker for bladder cancer, is herein proposed. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  7. Corneal aldehyde dehydrogenase and glutathione S-transferase activity after excimer laser keratectomy in guinea pigs.

    Science.gov (United States)

    Bilgihan, K; Bilgihan, A; Hasanreisoğlu, B; Turkozkan, N

    1998-03-01

    The free radical balance of the eye may be changed by excimer laser keratectomy. Previous studies have demonstrated that excimer laser keratectomy increases the corneal temperature, decreases the superoxide dismutase activity of the aqueous, and induces lipid peroxidation in the superficial corneal stroma. Aldehyde dehydrogenase (ALDH) and glutathione S-transferase (GST) are known to play an important role in corneal metabolism, particularly in detoxification of aldehydes, which are generated from free radical reactions. In three groups of guinea pigs mechanical corneal de-epithelialisation was performed in group I, superficial corneal photoablation in group II, and deep corneal photoablation in group III, and the corneal ALDH and GST activities measured after 48 hours. The mean ALDH and GST activities of group I and II showed no differences compared with the controls (p > 0.05). The corneal ALDH activities were found to be significantly decreased (p < 0.05) and GST activities increased (p < 0.05) in group III. These results suggest that excimer laser treatment of high myopia may change the ALDH and GST activities, metabolism, and free radical balance of the cornea.

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

    International Nuclear Information System (INIS)

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

    2011-01-01

    Highlights: ► Isolated ALDH Hi PC3 cells preferentially form primitive holoclone-type colonies. ► Primitive holoclone colonies are predominantly ALDH Lo but contain rare ALDH Hi cells. ► Holoclone-forming cells are not restricted to the ALDH Hi population. ► ALDH phenotypic plasticity occurs in PC3 cells (ALDH Lo to ALDH Hi and vice versa). ► ALDH 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 Lo cells in experimental models. However it is unclear whether all CSC-like cells are within the ALDH Hi population, or whether all ALDH 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 Hi cells enriches for the most primitive holoclone population, however holoclone formation is not restricted to ALDH Hi cells. In addition, we show that ALDH activity undergoes phenotypic plasticity, since the ALDH Lo population can develop ALDH Hi populations comparable to parental cells within 2 weeks in culture. Furthermore, we show that the majority of ALDH Hi cells are found within the least primitive paraclone population, which is circumvented by culturing PC3 cells as spheroids in defined medium favouring stem cell characteristics. Although ALDH Hi status enriches for holoclone formation, this activity may be mediated by a minority of ALDH Hi cells.

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

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    Marc-Estienne Roehrich

    2013-01-01

    Full Text Available 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.

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

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

    2017-06-01

    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) and matrix

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

    Energy Technology Data Exchange (ETDEWEB)

    Doherty, R.E.; Haywood-Small, S.L. [Biomedical Research Centre, Sheffield Hallam University, Sheffield S1 1WB (United Kingdom); Sisley, K. [Department of Oncology, Academic Unit of Ophthalmology and Orthopties, University of Sheffield, Sheffield S10 2RX (United Kingdom); Cross, N.A., E-mail: n.cross@shu.ac.uk [Biomedical Research Centre, Sheffield Hallam University, Sheffield S1 1WB (United Kingdom)

    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

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

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    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.

  13. Aldehyde dehydrogenase protein superfamily in maize.

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    Zhou, Mei-Liang; Zhang, Qian; Zhou, Ming; Qi, Lei-Peng; Yang, Xiong-Bang; Zhang, Kai-Xuan; Pang, Jun-Feng; Zhu, Xue-Mei; Shao, Ji-Rong; Tang, Yi-Xiong; Wu, Yan-Min

    2012-11-01

    Maize (Zea mays ssp. mays L.) is an important model organism for fundamental research in the agro-biotechnology field. Aldehydes were generated in response to a suite of environmental stresses that perturb metabolism including salinity, dehydration, desiccation, and cold and heat shock. Many biologically important aldehydes are metabolized by the superfamily of NAD(P)(+)-dependent aldehyde dehydrogenases. Here, starting from the database of Z. mays, we identified 28 aldehyde dehydrogenase (ALDH) genes and 48 transcripts by the in silico cloning method using the ALDH-conserved domain amino acid sequence of Arabidopsis and rice as a probe. Phylogenetic analysis shows that all 28 members of the ALDH gene families were classified to ten distinct subfamilies. Microarray data and quantitative real-time PCR analysis reveal that ZmALDH9, ZmALDH13, and ZmALDH17 genes involve the function of drought stress, acid tolerance, and pathogens infection. These results suggested that these three ZmALDH genes might be potentially useful in maize genetic improvement.

  14. Prognostic values of aldehyde dehydrogenase 1 isoenzymes in ovarian cancer

    OpenAIRE

    Ma, Yu-mei; Zhao, Shan

    2016-01-01

    Yu-mei Ma,1 Shan Zhao2 1Department of Pathology, 2Department of Cancer Second Division, The Second Hospital of Hebei Medical University, Shijiazhuang City, People’s Republic of China Abstract: Aldehyde dehydrogenase 1 (ALDH1) activity has been used as a functional stem cell marker to isolate cancer stem cells in different cancer types, including ovarian cancer. However, which ALDH1’s isoenzymes are contributing to ALDH1 activity in ovarian cancer remains elusive. In addi...

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

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    Iniesta Pilar

    2011-08-01

    Full Text Available Abstract 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.

  16. Multiple binding of thallium and rubidium to potassium-activated yeast aldehyde dehydrogenase. Influences on tertiary structure, stability and catalytic activity.

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    Bostian, K A; Betts, G F; Man, W K; Hughes, M N

    1982-01-01

    Univalent cation activators of aldehyde dehydrogenase have dual effects, both interpreted as cation-induced or -stabilized conformation changes. These two processes are differentiated by the time scales of their associated changes in activity. Using Tl+ as an activator, under certain conditions, the slower change in activity saturates at a Tl+ concentration which is only 0.1 Ks for the faster change. This, together with evidence for cation-induced rather than cation-stabilized conformation changes, is used to propose separate binding sites for cations responsible for the two activation processes. Equilibrium dialysis indicates 4 binding sites per active site for Rb+ or 6 sites for Tl+. At least one of the additional sites for Tl+ is an inhibitory site which has been differentiated from the activator sites on the basis of steady-state and pre-steady-state kinetic data. PMID:6758767

  17. Differences in the roles of conserved glutamic acid residues in the active site of human class 3 and class 2 aldehyde dehydrogenases.

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    Mann, C J; Weiner, H

    1999-10-01

    Although the three-dimensional structure of the dimeric class 3 rat aldehyde dehydrogenase has recently been published (Liu ZJ et al., 1997, Nature Struct Biol 4:317-326), few mechanistic studies have been conducted on this isoenzyme. We have characterized the enzymatic properties of recombinant class 3 human stomach aldehyde dehydrogenase, which is very similar in amino acid sequence to the class 3 rat aldehyde dehydrogenase. We have determined that the rate-limiting step for the human class 3 isozyme is hydride transfer rather than deacylation as observed for the human liver class 2 mitochondrial enzyme. No enhancement of NADH fluorescence was observed upon binding to the class 3 enzyme, while fluorescence enhancement of NADH has been previously observed upon binding to the class 2 isoenzyme. It was also observed that binding of the NAD cofactor inhibited the esterase activity of the class 3 enzyme while activating the esterase activity of the class 2 enzyme. Site-directed mutagenesis of two conserved glutamic acid residues (209 and 333) to glutamine residues indicated that, unlike in the class 2 enzyme, Glu333 served as the general base in the catalytic reaction and E209Q had only marginal effects on enzyme activity, thus confirming the proposed mechanism (Hempel J et al., 1999, Adv Exp Med Biol 436:53-59). Together, these data suggest that even though the subunit structures and active site residues of the isozymes are similar, the enzymes have very distinct properties besides their oligomeric state (dimer vs. tetramer) and substrate specificity.

  18. aldB, an RpoS-dependent gene in Escherichia coli encoding an aldehyde dehydrogenase that is repressed by Fis and activated by Crp.

    OpenAIRE

    Xu, J; Johnson, R C

    1995-01-01

    Escherichia coli aldB was identified as a gene that is negatively regulated by Fis but positively regulated by RpoS. The complete DNA sequence determined in this study indicates that aldB encodes a 56.3-kDa protein which shares a high degree of homology with an acetaldehyde dehydrogenase encoded by acoD of Alcaligenes eutrophus and an aldehyde dehydrogenase encoded by aldA of Vibrio cholerae and significant homology with a group of other aldehyde dehydrogenases from prokaryotes and eukaryotes...

  19. Evidence that the C-terminal domain of a type B PutA protein contributes to aldehyde dehydrogenase activity and substrate channeling.

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    Luo, Min; Christgen, Shelbi; Sanyal, Nikhilesh; Arentson, Benjamin W; Becker, Donald F; Tanner, John J

    2014-09-09

    Proline utilization A (PutA) is a bifunctional enzyme that catalyzes the oxidation of proline to glutamate. Structures of type A PutAs have revealed the catalytic core consisting of proline dehydrogenase (PRODH) and Δ(1)-pyrroline-5-carboxylate dehydrogenase (P5CDH) modules connected by a substrate-channeling tunnel. Type B PutAs also have a C-terminal domain of unknown function (CTDUF) that is absent in type A PutAs. Small-angle X-ray scattering (SAXS), mutagenesis, and kinetics are used to determine the contributions of this domain to PutA structure and function. The 1127-residue Rhodobacter capsulatus PutA (RcPutA) is used as a representative CTDUF-containing type B PutA. The reaction progress curve for the coupled PRODH-P5CDH activity of RcPutA does not exhibit a time lag, implying a substrate channeling mechanism. RcPutA is monomeric in solution, which is unprecedented for PutAs. SAXS rigid body modeling with target-decoy validation is used to build a model of RcPutA. On the basis of homology to aldehyde dehydrogenases (ALDHs), the CTDUF is predicted to consist of a β-hairpin fused to a noncatalytic Rossmann fold domain. The predicted tertiary structural interactions of the CTDUF resemble the quaternary structural interactions in the type A PutA dimer interface. The model is tested by mutagenesis of the dimerization hairpin of a type A PutA and the CTDUF hairpin of RcPutA. Similar functional phenotypes are observed in the two sets of variants, supporting the hypothesis that the CTDUF mimics the type A PutA dimer interface. These results suggest annotation of the CTDUF as an ALDH superfamily domain that facilitates P5CDH activity and substrate channeling by stabilizing the aldehyde-binding site and sealing the substrate-channeling tunnel from the bulk medium.

  20. aldB, an RpoS-dependent gene in Escherichia coli encoding an aldehyde dehydrogenase that is repressed by Fis and activated by Crp.

    Science.gov (United States)

    Xu, J; Johnson, R C

    1995-06-01

    Escherichia coli aldB was identified as a gene that is negatively regulated by Fis but positively regulated by RpoS. The complete DNA sequence determined in this study indicates that aldB encodes a 56.3-kDa protein which shares a high degree of homology with an acetaldehyde dehydrogenase encoded by acoD of Alcaligenes eutrophus and an aldehyde dehydrogenase encoded by aldA of Vibrio cholerae and significant homology with a group of other aldehyde dehydrogenases from prokaryotes and eukaryotes. Expression of aldB is maximally induced during the transition from exponential phase to stationary phase. Its message levels are elevated three- to fourfold by a fis mutation and abolished by an rpoS mutation. In addition, the expression of an aldB-lacZ fusion was decreased about 20-fold in the absence of crp. DNase I footprinting analysis showed that five Fis binding sites and one Crp binding site are located within the aldB promoter region, suggesting that Fis and Crp are acting directly to control aldB transcription. AldB expression is induced by ethanol, but in contrast to that of most of the RpoS-dependent genes, the expression of aldB is not altered by an increase in medium osmolarity.

  1. Effects of Betaine Aldehyde Dehydrogenase-Transgenic Soybean on Phosphatase Activities and Rhizospheric Bacterial Community of the Saline-Alkali Soil

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    Ying Nie

    2016-01-01

    Full Text Available 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.

  2. [Activity of the octanol dehydrogenase, of the alcool dehydrogenase and aldehyde dehydrogenase on the farnesol metabolism. Photoperiodic and neurhormonale regulation, controlling the metabolism of the juvenile hormone, in Pieris brassicae (author's transl)].

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    L'Hélias, C

    1979-01-01

    The antagonistic photoperiodic behaviour of the farnesol dehydrogenases indicates that the photonic control mechanism of the brain acts on the farnesol derivates. This cerebral control is double. The first system, linked at the allatotrope function is proportionnal at the photoperiod and acts on the octanol dehydrogenase 0,32. The second system controle the deshydrogenases ADH bands 0,50--0,58, is linked at the darkness. It is linked also at the neurocerebral activity then it stops its activity at the 4th day of the 5th stage. This last seems to be the determinating control for the establishment of the diapause since in short photoperiod, when the inhibition by this system ends, the alcool dehydrogenases 0,50-0,58 series is suractivated in rate with the lasting of the scotophase. In darkness, the 1st system functionnes cyclically and has a maximum synchron with the single maximum of the 2nd system. Inversally, in continuous light, the 2nd system is synchronisated with the 1st which has a prolongated action, maybe linked with a prolongated activity of the neurosecretory cells of the pars intercerebralis and corpora allata.

  3. Five Fatty Aldehyde Dehydrogenase Enzymes from Marinobacter and Acinetobacter spp. and Structural Insights into the Aldehyde Binding Pocket

    Energy Technology Data Exchange (ETDEWEB)

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

    2017-04-07

    ABSTRACT

    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 fromMarinobacter aquaeoleiVT8 and an additional enzyme fromAcinetobacter baylyiwere heterologously expressed inEscherichia coliand shown to display FAldDH activity. Five enzymes (Maqu_0438, Maqu_3316, Maqu_3410, Maqu_3572, and the enzyme reported under RefSeq accession no.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 specificities of these enzymes with various aldehydes is presented. Crystallization trials yielded diffraction-quality crystals of one particular FAldDH (Maqu_3316) fromM. aquaeoleiVT8. 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 catalysis by the enzyme is accomplished is also provided.

    IMPORTANCEThis study provides a comparison of multiple enzymes with the ability

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

    Science.gov (United States)

    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

  5. Role and structural characterization of plant aldehyde dehydrogenases from family 2 and family 7

    Czech Academy of Sciences Publication Activity Database

    Končitíková, R.; Vigouroux, A.; Kopečná, M.; Andree, T.; Bartoš, Jan; Šebela, M.; Moréra, S.; Kopečný, D.

    2015-01-01

    Roč. 468, Part: 1 (2015), s. 109-123 ISSN 0264-6021 R&D Projects: GA ČR GA15-22322S; GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : aldehyde dehydrogenase 2 (ALDH2) * aldehyde dehydrogenase 7 (ALDH7) * benzaldehyde Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.562, year: 2015

  6. Targeting aldehyde dehydrogenase: a potential approach for cell labeling

    Energy Technology Data Exchange (ETDEWEB)

    Vaidyanathan, Ganesan [Department of Radiology, Duke University Medical Center, Box 3808, Durham, NC 27710 (United States)], E-mail: ganesan.v@duke.edu; Song, Haijing; Affleck, Donna; McDougald, Darryl L. [Department of Radiology, Duke University Medical Center, Box 3808, Durham, NC 27710 (United States); Storms, Robert W. [Division of Cellular Therapy, Department of Medicine, Duke University Medical Center, Durham, NC 27710 (United States); Zalutsky, Michael R.; Chin, Bennett B. [Department of Radiology, Duke University Medical Center, Box 3808, Durham, NC 27710 (United States)

    2009-11-15

    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 radioiodinated 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 of [{sup 125}I]FMIC and [{sup 125}I]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.

  7. Targeting aldehyde dehydrogenase: a potential approach for cell labeling

    International Nuclear Information System (INIS)

    Vaidyanathan, Ganesan; Song, Haijing; Affleck, Donna; McDougald, Darryl L.; Storms, Robert W.; Zalutsky, 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 radioiodinated 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 of [ 125 I]FMIC and [ 125 I]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.

  8. Nitrite reductase activity of rat and human xanthine oxidase, xanthine dehydrogenase, and aldehyde oxidase: evaluation of their contribution to NO formation in vivo.

    Science.gov (United States)

    Maia, Luisa B; Pereira, Vânia; Mira, Lurdes; Moura, José J G

    2015-01-27

    Nitrite is presently considered a NO "storage form" that can be made available, through its one-electron reduction, to maintain NO formation under hypoxia/anoxia. The molybdoenzymes xanthine oxidase/dehydrogenase (XO/XD) and aldehyde oxidase (AO) are two of the most promising mammalian nitrite reductases, and in this work, we characterized NO formation by rat and human XO/XD and AO. This is the first characterization of human enzymes, and our results support the employment of rat liver enzymes as suitable models of the human counterparts. A comprehensive kinetic characterization of the effect of pH on XO and AO-catalyzed nitrite reduction showed that the enzyme's specificity constant for nitrite increase 8-fold, while the Km(NO2(-)) decrease 6-fold, when the pH decreases from 7.4 to 6.3. These results demonstrate that the ability of XO/AO to trigger NO formation would be greatly enhanced under the acidic conditions characteristic of ischemia. The dioxygen inhibition was quantified, and the Ki(O2) values found (24.3-48.8 μM) suggest that in vivo NO formation would be fine-tuned by dioxygen availability. The potential in vivo relative physiological relevance of XO/XD/AO-dependent pathways of NO formation was evaluated using HepG2 and HMEC cell lines subjected to hypoxia. NO formation by the cells was found to be pH-, nitrite-, and dioxygen-dependent, and the relative contribution of XO/XD plus AO was found to be as high as 50%. Collectively, our results supported the possibility that XO/XD and AO can contribute to NO generation under hypoxia inside a living human cell. Furthermore, the molecular mechanism of XO/AO-catalyzed nitrite reduction was revised.

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

    International Nuclear Information System (INIS)

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

    1990-01-01

    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

  10. Group X Aldehyde Dehydrogenases of Pseudomonas aeruginosa PAO1 Degrade Hydrazones

    OpenAIRE

    Taniyama, Kosuke; Itoh, Hideomi; Takuwa, Atsushi; Sasaki, Yasuyuki; Yajima, Shunsuke; Toyofuku, Masanori; Nomura, Nobuhiko; Takaya, Naoki

    2012-01-01

    Hydrazones are natural and synthetic compounds containing a C=N-N moiety. Here we found that the opportunistic pathogen Pseudomonas aeruginosa PAO1 produced NAD+- or NADP+-dependent hydrazone dehydrogenase (HDH), which converts hydrazones to the corresponding hydrazides and acids rather than to the simple hydrolytic product aldehydes. Gene cloning indicated that the HDH is part of the group X aldehyde dehydrogenase (ALDH) family, which is distributed among bacteria, although the physiological...

  11. Heterologous Expression of Aldehyde Dehydrogenase in Lactococcus lactis for Acetaldehyde Detoxification at Low pH.

    Science.gov (United States)

    Lyu, Yunbin; LaPointe, Gisèle; Zhong, Lei; Lu, Jing; Zhang, Chong; Lu, Zhaoxin

    2018-02-01

    Aldehyde dehydrogenase (E.C. 1.2.1.x) can catalyze detoxification of acetaldehydes. A novel acetaldehyde dehydrogenase (istALDH) from the non-Saccharomyces yeast Issatchenkia terricola strain XJ-2 has been previously characterized. In this work, Lactococcus lactis with the NIsin Controlled Expression (NICE) System was applied to express the aldehyde dehydrogenase gene (istALDH) in order to catalyze oxidation of acetaldehyde at low pH. A recombinant L. lactis NZ3900 was obtained and applied for the detoxification of acetaldehyde as whole-cell biocatalysts. The activity of IstALDH in L. lactis NZ3900 (pNZ8148-istALDH) reached 36.4 U mL -1 when the recombinant cells were induced with 50 ng mL -1 nisin at 20 °C for 2 h. The IstALDH activity of recombinant L. lactis cells showed higher stability at 37 °C and pH 4.0 compared with the crude enzyme. L. lactis NZ3900 (pNZ8148-istALDH) could convert acetaldehyde at pH 2.0 while the crude enzyme could not. Moreover, the resting cells of L. lactis NZ3900 (pNZ8148-istALDH) showed a 2.5-fold higher activity and better stability in catalyzing oxidation of acetaldehyde at pH 2.0 compared with that of Escherichia coli expressing the IstALDH. Taken together, the L. lactis cells expressing recombinant IstALDH are potential whole-cell biocatalysts that can be applied in the detoxification of aldehydes.

  12. Modulation of the reactivity of the essential cysteine residue of betaine aldehyde dehydrogenase from Pseudomonas aeruginosa.

    Science.gov (United States)

    González-Segura, Lilian; Velasco-García, Roberto; Muñoz-Clares, Rosario A

    2002-02-01

    Betaine aldehyde dehydrogenase (BADH) catalyses the irreversible NAD(P)(+)-dependent oxidation of betaine aldehyde to glycine betaine. In the human opportunistic pathogen Pseudomonas aeruginosa this reaction is an obligatory step in the assimilation of carbon and nitrogen when bacteria are growing in choline or choline precursors. As with every aldehyde dehydrogenase studied so far, BADH possesses an essential cysteine residue involved in the formation of the intermediate thiohemiacetal with the aldehyde substrate. We report here that the chemical modification of this residue is conveniently measured by the loss in enzyme activity, which allowed us to explore its reactivity in a pH range around neutrality. The pH dependence of the observed second-order rate constant of BADH inactivation by methyl methanethiosulphonate (MMTS) suggests that at low pH values the essential cysteine residue exists as thiolate by the formation of an ion pair with a positively charged residue. The estimated macroscopic pK values are 8.6 and 4.0 for the free and ion-pair-forming thiolate respectively. The reactivity towards MMTS of both thiolate forms is notably lower than that of model compounds of similar pK, suggesting a considerable steric inhibition by the structure of the protein. Binding of the dinucleotides rapidly induced a significant and transitory increment of thiolate reactivity, followed by a relatively slow change to an almost unreactive form. Thus it seems that to gain protection against oxidation without compromising catalytic efficiency, BADH from P. aeruginosa has evolved a complex and previously undescribed mechanism, involving several conformational rearrangements of the active site, to suit the reactivity of the essential thiol to the availability of coenzyme and substrate.

  13. Aldehyde Dehydrogenase 2 Polymorphism Is a Predictor of Smoking Cessation.

    Science.gov (United States)

    Masaoka, Hiroyuki; Gallus, Silvano; Ito, Hidemi; Watanabe, Miki; Yokomizo, Akira; Eto, Masatoshi; Matsuo, Keitaro

    2017-09-01

    Smoking cessation has been known to be associated with drinking behaviors, which are influenced by polymorphisms in genes encoding alcohol metabolizing enzymes. The aim was to evaluate the impact of aldehyde dehydrogenase 2 (ALDH2, rs671) and alcohol dehydrogenase 1B (ADH1B, rs1229984) polymorphisms together with drinking behaviors on smoking cessation. We conducted a cross-sectional study with 1137 former smokers and 1775 current smokers without any cancer at Aichi Cancer Center Hospital between 2001 and 2005. Unconditional logistic regression models were used to estimate odds ratios (OR) and 95% confidence intervals (CI) for successful smoking cessation by comparing former smokers (quitters) with current smokers (non-quitters). Older age, lower amount of cumulative smoking exposure, lower number of cigarettes per day, younger age of smoking initiation, shorter smoking duration, longer time to first cigarette in the morning, and lower amount of drinking among ever drinkers were predictors of smoking cessation. After careful adjustment for age, sex, smoking patterns, and drinking status, the ORs for smoking cessation among subjects with ALDH2 Glu/Lys and Lys/Lys were 1.02 (95% CI 0.84-1.23) and 1.78 (95% CI 1.23-2.58) compared with those with ALDH2 Glu/Glu, respectively Mediation analyses confirmed that the effect of ALDH2 Lys/Lys on smoking cessation was independent by dinking behaviors. No statistically significant association between ADH1B polymorphism and smoking cessation was observed. In our Japanese population, ALDH2 polymorphism predicts smoking cessation, independent by drinking behaviors. Interventions for promoting smoking cessation by ALDH2 polymorphism may be useful in Asian populations. We newly show that subjects with ALDH2 Lys/Lys genotype in a functional polymorphism, rs671, are more likely to quit smoking than those with ALDH2 Glu allele in a Japanese population. Our finding suggests that ALDH2 polymorphism may be useful for promoting smoking

  14. Group X aldehyde dehydrogenases of Pseudomonas aeruginosa PAO1 degrade hydrazones.

    Science.gov (United States)

    Taniyama, Kosuke; Itoh, Hideomi; Takuwa, Atsushi; Sasaki, Yasuyuki; Yajima, Shunsuke; Toyofuku, Masanori; Nomura, Nobuhiko; Takaya, Naoki

    2012-03-01

    Hydrazones are natural and synthetic compounds containing a C=N-N moiety. Here we found that the opportunistic pathogen Pseudomonas aeruginosa PAO1 produced NAD(+)- or NADP(+)-dependent hydrazone dehydrogenase (HDH), which converts hydrazones to the corresponding hydrazides and acids rather than to the simple hydrolytic product aldehydes. Gene cloning indicated that the HDH is part of the group X aldehyde dehydrogenase (ALDH) family, which is distributed among bacteria, although the physiological roles of the ALDH family remain unknown. The PAO1 strain upregulated HDH in the presence of the hydrazone adipic acid bis(ethylidene hydrazide) (AEH). Gene disruption of the HDH-encoding hdhA (PA4022) decreased growth rates in culture medium containing AEH as the sole carbon source, and this effect was more obvious in the double gene disruption of hdhA and its orthologous exaC (PA1984), indicating that these genes are responsible for hydrazone utilization. Recombinant proteins of group X ALDHs from Escherichia coli, Paracoccus denitrificans, and Ochrobactrum anthropi also acted as HDHs in that they produced HDH activity in the cells and degraded hydrazones. These findings indicated the physiological roles of group X ALDHs in bacteria and showed that they comprise a distinct ALDH subfamily.

  15. Blood Leukocyte Counts and Genetic Polymorphisms of Alcohol Dehydrogenase-1B and Aldehyde Dehydrogenase-2 in Japanese Alcoholic Men.

    Science.gov (United States)

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

    2016-03-01

    Roughly 40% of East Asians have inactive aldehyde dehydrogenase-2 (ALDH2) encoded by the ALDH2*2 allele, and 90% have highly active alcohol dehydrogenase-1B (ADH1B) encoded by the ADH1B*2 allele. Macrocytosis and macrocytic anemia in alcoholics have been associated with ADH1B and ALDH2 gene variants which increase acetaldehyde (AcH) levels. We investigated the relationship between ADH1B*2, ALDH2*2, and leukocyte counts of Japanese alcoholic men (N = 1,661). After adjusting for age, drinking habits, smoking habits, body mass index, presence of liver cirrhosis, and serum levels of C-reactive protein, we found that total and differential leukocyte counts were lower in the presence of the ALDH2*1/*2 genotype (vs. ALDH2*1/*1 genotype). ALDH2*2/*2 carriers were not found in our study population. Leukocyte, granulocyte, and monocyte counts were also lower in the presence of ADH1B*2 (vs. ADH1B*1/*1 genotype), but the lymphocyte count was higher. The ALDH2*1/*2 genotype was associated with leukocytopenia (counts. The total and differential blood leukocyte counts of Japanese alcoholics were strongly affected by their ADH1B and ALDH2 gene variants. High AcH exposure levels probably play a critical role in the suppression of blood leukocyte counts in alcoholics. Copyright © 2016 by the Research Society on Alcoholism.

  16. Identification and characterisation of Aedes aegypti aldehyde dehydrogenases involved in pyrethroid metabolism.

    Directory of Open Access Journals (Sweden)

    Nongkran Lumjuan

    Full Text Available Pyrethroid insecticides, especially permethrin and deltamethrin, have been used extensively worldwide for mosquito control. However, insecticide resistance can spread through a population very rapidly under strong selection pressure from insecticide use. The upregulation of aldehyde dehydrogenase (ALDH has been reported upon pyrethroid treatment. In Aedes aegypti, the increase in ALDH activity against the hydrolytic product of pyrethroid has been observed in DDT/permethrin-resistant strains. The objective of this study was to identify the role of individual ALDHs involved in pyrethroid metabolism.Three ALDHs were identified; two of these, ALDH9948 and ALDH14080, were upregulated in terms of both mRNA and protein levels in a DDT/pyrethroid-resistant strain of Ae. aegypti. Recombinant ALDH9948 and ALDH14080 exhibited oxidase activities to catalyse the oxidation of a permethrin intermediate, phenoxybenzyl aldehyde (PBald, to phenoxybenzoic acid (PBacid.ALDHs have been identified in association with permethrin resistance in Ae. aegypti. Characterisation of recombinant ALDHs confirmed the role of this protein in pyrethroid metabolism. Understanding the biochemical and molecular mechanisms of pyrethroid resistance provides information for improving vector control strategies.

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

    Science.gov (United States)

    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

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

    Directory of Open Access Journals (Sweden)

    Tetsuhiko Ikeda

    Full Text Available 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.

  19. Expression of the Vibrio cholerae gene encoding aldehyde dehydrogenase is under control of ToxR, the cholera toxin transcriptional activator.

    OpenAIRE

    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 st...

  20. Spectroscopic characterisation of interaction of ferulic acid with aldehyde dehydrogenase (ALDH).

    Science.gov (United States)

    Kolawole, Ayodele O; Agaba, Ruth J; Oluwole, Matthew O

    2017-05-01

    Interaction of a pharmacological important phenolic, ferulic acid, with Aldehyde dehydrogenase (ALDH) at the simulative pH condition, was studied using spectroscopic approach. Ferulic acid caused a decrease in the fluorescence intensity formed from ALDH-ferulic acid complex resulting in mixed inhibition of ALDH activity (IC 50 =30.65μM). The intrinsic quenching was dynamic and induced altered conformation of ALDH and made the protein less compact but might not unfold it. ALDH has two binding sites for ferulic acid at saturating concentrations having association constant of 1.35×10 3 Lmol -1 and a dissociation constant of 9.7×10 7 Lmol -1 at 25°C indicating ALDH-ferulic acid complex formation is more favourable than its dissociation. The interaction was not spontaneous and endothermic and suggests the involvement of hydrophobic interactions with a FRET binding distance of 4.49nm. Change in pH near and far from isoelectric points of ferulic acid did not affect the bonding interaction. Using trehalose as viscosogen, the result from Stoke-Einstein hypothesis showed that ferulic acid-ALDH binding and dissociation equilibrium was diffusion controlled. These results clearly suggest the unique binding properties and lipophilicity influence of ferulic acid. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    Science.gov (United States)

    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.

  2. Aldehyde Dehydrogenase 1 and Raf Kinase Inhibitor Protein ...

    African Journals Online (AJOL)

    interestingly defines the higher expression in low grade cervical cancer to regulate the tumour, but shows little or no very mild ... Conclusion: ALDH1 and RKIP marker in association correlation with Sox2 aids in defining the proliferative ability of .... endogenous peroxidase activity was blocked by immersing the sections in ...

  3. Characterization of aldehyde dehydrogenase isozymes in ovarian cancer tissues and sphere cultures

    International Nuclear Information System (INIS)

    Saw, Yu-Ting; Thompson, David; Vasiliou, Vasilis; Berkowitz, Ross S; Ng, Shu-Wing; Yang, Junzheng; Ng, Shu-Kay; Liu, Shubai; Singh, Surendra; Singh, Margit; Welch, William R; Tsuda, Hiroshi; Fong, Wing-Ping

    2012-01-01

    Aldehyde dehydrogenases belong to a superfamily of detoxifying enzymes that protect cells from carcinogenic aldehydes. Of the superfamily, ALDH1A1 has gained most attention because current studies have shown that its expression is associated with human cancer stem cells. However, ALDH1A1 is only one of the 19 human ALDH subfamilies currently known. The purpose of the present study was to determine if the expression and activities of other major ALDH isozymes are associated with human ovarian cancer and ovarian cancer sphere cultures. Immunohistochemistry was used to delineate ALDH isozyme localization in clinical ovarian tissues. Western Blot analyses were performed on lysates prepared from cancer cell lines and ovarian cancer spheres to confirm the immunohistochemistry findings. Quantitative reverse transcription-polymerase chain reactions were used to measure the mRNA expression levels. The Aldefluor® assay was used to measure ALDH activity in cancer cells from the four tumor subtypes. Immunohistochemical staining showed significant overexpression of ALDH1A3, ALDH3A2, and ALDH7A1 isozymes in ovarian tumors relative to normal ovarian tissues. The expression and activity of ALDH1A1 is tumor type-dependent, as seen from immunohistochemisty, Western blot analysis, and the Aldefluor® assay. The expression was elevated in the mucinous and endometrioid ovarian epithelial tumors than in serous and clear cell tumors. In some serous and most clear cell tumors, ALDH1A1 expression was found in the stromal fibroblasts. RNA expression of all studied ALDH isozymes also showed higher expression in endometrioid and mucinous tumors than in the serous and clear cell subtypes. The expression of ALDH enzymes showed tumor type-dependent induction in ovarian cancer cells growing as sphere suspensions in serum-free medium. The results of our study indicate that ALDH enzyme expression and activity may be associated with specific cell types in ovarian tumor tissues and vary according to

  4. The dental pulp stem cell niche based on aldehyde dehydrogenase 1 expression

    Science.gov (United States)

    Machado, CV; Passos, ST; Campos, TMC; Bernardi, L; Vilas-Bôas, DS; Nör, JE; Telles, PDS; Nascimento, IL

    2015-01-01

    Aim To detect cells expressing the stem cell marker ALDH1 (aldehyde dehydrogenase1) in the pulp of human permanent teeth and to investigate the expression of ALDH1 in isolated dental pulp cells. Methodology Pulp tissue was collected and processed for immunohistochemistry to detect ALDH1, STRO-1 and CD90 positive cells. In addition, cells were isolated and analyzed by flow cytometry for ALDH1 activity, and for the cell surface markers CD44, CD73, CD90, STRO-1 and CD45. Cells were also examined for multi-differentiation capacity. Within these cells, an ALDH1+ cell subpopulation was selected and evaluated for multi-differentiation capacity. Results The immunohistochemistry analyses showed that ALDH1, CD90 and STRO-1 positive cells were located mainly in the perivascular areas and nerve fibres of dental pulps. Cells on the fifth passage had high expression for CD44, CD73 and CD90, whereas moderate labeling was observed for STRO-1 and ALDH1 in flow cytometry analysis. On the same passages, cells were able to differentiate into osteogenic, adipogenic and chondrogenic lineages. The ALDH1+ cell subpopulation also demonstrated multi-lineage differentiation ability. Conclusions Dental pulp stem cells reside in the vicinity of blood vessels and nerve fibres, indicating the possible existence of more than one stem cell niche in dental pulps. Furthermore, ALDH1 was expressed by isolated dental pulp cells, which had mesenchymal stem cell characteristics. Thus, it can be suggested that ALDH1 may be used as a DPSC marker. PMID:26198909

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

    Energy Technology Data Exchange (ETDEWEB)

    Rogers, G.R.; Markova, N.G.; Compton, J.G. [National Institutes of Health, Bethesda, MD (United States)] [and others

    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.

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

    Directory of Open Access Journals (Sweden)

    Malaspina Patrizia

    2009-01-01

    Full Text Available Abstract 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.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  9. Human aldehyde dehydrogenase genes: alternatively spliced transcriptional variants and their suggested nomenclature.

    Science.gov (United States)

    Black, William J; Stagos, Dimitrios; Marchitti, Satori A; Nebert, Daniel W; Tipton, Keith F; Bairoch, Amos; Vasiliou, Vasilis

    2009-11-01

    The human aldehyde dehydrogenase (ALDH) gene superfamily consists of 19 genes encoding enzymes critical for NAD(P)-dependent oxidation of endogenous and exogenous aldehydes, including drugs and environmental toxicants. Mutations in ALDH genes are the molecular basis of several disease states (e.g. Sjögren-Larsson syndrome, pyridoxine-dependent seizures, and type II hyperprolinemia) and may contribute to the etiology of complex diseases such as cancer and Alzheimer's disease. The aim of this nomenclature update was to identify splice transcriptional variants principally for the human ALDH genes. Data-mining methods were used to retrieve all human ALDH sequences. Alternatively spliced transcriptional variants were determined based on (i) criteria for sequence integrity and genomic alignment; (ii) evidence of multiple independent cDNA sequences corresponding to a variant sequence; and (iii) if available, empirical evidence of variants from the literature. Alternatively spliced transcriptional variants and their encoded proteins exist for most of the human ALDH genes; however, their function and significance remain to be established. When compared with the human genome, rat and mouse include an additional gene, Aldh1a7, in the ALDH1A subfamily. To avoid confusion when identifying splice variants in various genomes, nomenclature guidelines for the naming of such alternative transcriptional variants and proteins are recommended herein. In addition, a web database (www.aldh.org) has been developed to provide up-to-date information and nomenclature guidelines for the ALDH superfamily.

  10. Glu504Lys Single Nucleotide Polymorphism of Aldehyde Dehydrogenase 2 Gene and the Risk of Human Diseases

    Directory of Open Access Journals (Sweden)

    Yan Zhao

    2015-01-01

    Full Text Available Aldehyde dehydrogenase (ALDH 2 is a mitochondrial enzyme that is known for its important role in oxidation and detoxification of ethanol metabolite acetaldehyde. ALDH2 also metabolizes other reactive aldehydes such as 4-hydroxy-2-nonenal and acrolein. The Glu504Lys single nucleotide polymorphism (SNP of ALDH2 gene, which is found in approximately 40% of the East Asian populations, causes defect in the enzyme activity of ALDH2, leading to alterations in acetaldehyde metabolism and alcohol-induced “flushing” syndrome. Evidence suggests that ALDH2 Glu504Lys SNP is a potential candidate genetic risk factor for a variety of chronic diseases such as cardiovascular disease, cancer, and late-onset Alzheimer’s disease. In addition, the association between ALDH2 Glu504Lys SNP and the development of these chronic diseases appears to be affected by the interaction between the SNP and lifestyle factors such as alcohol consumption as well as by the presence of other genetic variations.

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

    Science.gov (United States)

    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

    2017-06-01

    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 approximately 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. Copyright© Ferrata Storti Foundation.

  12. Genetic Polymorphisms of the Mitochondrial Aldehyde Dehydrogenase ALDH2 Gene in a Large Ethnic Hakka Population in Southern China.

    Science.gov (United States)

    Zhong, Zhixiong; Hou, Jingyuan; Li, Bin; Zhang, Qifeng; Li, Cunren; Liu, Zhidong; Yang, Min; Zhong, Wei; Zhao, Pingsen

    2018-04-06

    BACKGROUND Human mitochondrial aldehyde dehydrogenase 2 (ALDH2) plays a critical role in the detoxification of the ethanol metabolite acetaldehyde. The ALDH2*2 (rs671) gene variant is mainly absent among Europeans but is prevalent in populations in East Asia. The aim of this study was to investigate ALDH2*2 mutant alleles and genotype frequencies in the Hakka population of China. MATERIAL AND METHODS Between January 2016 and June 2017, 7,966 unrelated individuals were recruited into the study from the Hakka ethnic population residing in the Meizhou area of Guangdong Province, China, who provided venous blood samples. Genotyping of ALDH2 genotypes were determined using a gene chip platform and confirmed by DNA sequencing. RESULTS In the 7,966 individuals from the Hakka population of China in this study, the frequencies of the ALDH2 genotypes *1/*1, *1/*2 and *2/*2 were 52.03%, 39.67%, and 8.30%, respectively; 47.97% of the individuals were found to carry the ALDH2*2 genotype, which was associated with a deficiency in the aldehyde dehydrogenase (ALDH2) enzyme activity. The frequency of the ALDH2*2 allele was lower than that previously reported in the Japanese population but higher than that reported in other Oriental populations. CONCLUSIONS The findings of this study have provided new information on the ALDH2 gene polymorphisms in the Hakka ethnic population residing in the Meizhou area of Guangdong Province, China, including an understanding of the origin of the atypical ALDH2*2 allele. Also, the study findings may be relevant to the primary care of patients in China.

  13. Aldehyde-induced xanthine oxidase activity in raw milk.

    Science.gov (United States)

    Steffensen, Charlotte L; Andersen, Henrik J; Nielsen, Jacob H

    2002-12-04

    In the present study, the aldehyde-induced pro-oxidative activity of xanthine oxidase was followed in an accelerated raw milk system using spin-trap electron spin resonance (ESR) spectroscopy. The aldehydes acetaldehyde, propanal, hexanal, trans-2-hexenal, trans-2-heptenal, trans-2-nonenal, and 3-methyl-2-butenal were all found to initiate radical reactions when added to milk. Formation of superoxide through aldehyde-induced xanthine oxidase activity is suggested as the initial reaction, as all tested aldehydes were shown to trigger superoxide formation in an ultrahigh temperature (UHT) milk model system with added xanthine oxidase. It was found that addition of aldehydes to milk initially increased the ascorbyl radical concentration with a subsequent decay due to ascorbate depletion, which renders the formation of superoxide in milk with added aldehyde. The present study shows for the first time potential acceleration of oxidative events in milk through aldehyde-induced xanthine oxidase activity.

  14. Aldehyde dehydrogenase 2*2 knock-in mice show increased reactive oxygen species production in response to cisplatin treatment.

    Science.gov (United States)

    Kim, Jeewon; Chen, Che-Hong; Yang, Jieying; Mochly-Rosen, Daria

    2017-05-22

    The aldehyde dehydrogenase (ALDH) enzyme family metabolizes and detoxifies both exogenous and endogenous aldehydes. Since chemotherapeutic agents, such as cisplatin, generate cytotoxic aldehydes and oxidative stress, and chemoresistant cancer cells express high levels of ALDH enzymes, we hypothesized that different ALDH expression within cells may show different chemosensitivity. ALDH2 has the lowest Km for acetaldehyde among ALDH isozymes and detoxifies acetaldehydes in addition to other reactive aldehydes, such as 4-hydroxy-nonenal, malondialdehyde and acrolein produced from lipid peroxidation by reactive oxygen species (ROS). Thus, cells with an ALDH2 variant may sensitize them to these ROS-inducing chemotherapy drugs. Here, we used wild type C57BL/6 mice and ALDH2*2 knock-in mutant mice and compared the basal level of ROS in different tissues. Then, we treated the mice with cisplatin, isolated cells from organs and fractionated them into lysates containing mitochondrial and cytosolic fractions, treated with cisplatin again in vitro, and compared the level of ROS generated. We show that overall ROS production increases with cisplatin treatment in cells with ALDH2 mutation. The treatment of cisplatin in the wild type mice did not change the level of ROS compared to PBS treated controls. In contrast, ALDH2*2 knock-in mutant mice showed a significantly increased level of ROS compared to wild type mice in tongue, lung, kidney and brain tissues without any treatment. ALDH2*2 mutant mice showed 20% of the ALDH2 activity in the kidney compared to wild type mice. Treatment of ALDH2*2 mutant mice with cisplatin showed increased ROS levels in the mitochondrial fraction of kidney. In the cytosolic fraction, treatment of mutant mice with cisplatin increased ROS levels in lung and brain compared to PBS treated controls. Furthermore, ALDH2*2 mutant mice treated with cisplatin showed increased cytotoxicity in the kidney cells compared to PBS treated mutant controls. These data

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

    Directory of Open Access Journals (Sweden)

    Feng-Xia Tian

    Full Text Available 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.

  16. The mutation in the mitochondrial aldehyde dehydrogenase (ALDH2) gene responsible for alcohol-induced flushing increases turnover of the enzyme tetramers in a dominant fashion.

    OpenAIRE

    Xiao, Q; Weiner, H; Crabb, D W

    1996-01-01

    Deficiency in mitochondrial aldehyde dehydrogenase (ALDH2), a tetrameric enzyme, results from inheriting one or two ALDH2*2 alleles. This allele encodes a protein subunit with a lysine for glutamate substitution at position 487 and is dominant over the wild-type allele, ALDH2*1. The ALDH2*2-encoded subunit (ALDH2K) reduces the activity of ALDH2 enzyme in cell lines expressing the wild-type subunit (ALDH2E). In addition to this effect on the enzyme activity, we now report that ALDH2*2 heterozy...

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

    Science.gov (United States)

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

    2014-01-01

    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. PMID:25045085

  18. Inhibitory effect of Nodal on the expression of aldehyde dehydrogenase 1 in endometrioid adenocarcinoma of uterus.

    Science.gov (United States)

    Wang, Yi; Jiang, Yang; Tian, Tian; Hori, Yumiko; Wada, Naoki; Ikeda, Jun-ichiro; Morii, Eiichi

    2013-11-01

    Cancers consist of heterogeneous populations. Recently, it has been demonstrated that cells with tumorigenic potential are limited to a small population, called cancer-initiating cells (CICs). Aldehyde dehydrogenase 1 (ALDH1) is one of the markers of CICs. We previously reported that ALDH1-high cases of uterine endometrioid adenocarcinoma showed poor prognosis, and ALDH1-high population of endometrioid adenocarcinoma cell line was more tumorigenic, resistant to anti-cancer drugs, and invasive than ALDH1-low population. Here, the regulatory signaling for ALDH1 was examined. The inhibition of TGF-β signaling increased ALDH1-high population. Among TGF-β family members, Nodal expression and ALDH1 expression levels were mutually exclusive. Immunohistochemical analysis on clinical samples revealed Nodal-high tumor cells to be ALDH-low and vise versa, suggesting that Nodal may inhibit ALDH1 expression via stimulating TGF-β signaling in uterine endometrioid adenocarcinoma. In fact, the addition of Nodal to endometrioid adenocarcinoma cell line reduced ALDH1-high population. Although ALDH1 mRNA level was not affected, the amount of ALDH1 protein appeared to be reduce by Nodal through ubiquitine-proteasome pathway. The regulation of TGF-β signaling might be a novel therapeutic target of CICs in endometrioid adenocarcinoma. Copyright © 2013. Published by Elsevier Inc.

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

    Science.gov (United States)

    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. © 2014 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  20. Interaction of Aldehyde dehydrogenase with acetaminophen as examined by spectroscopies and molecular docking

    Directory of Open Access Journals (Sweden)

    Ayodele O. Kolawole

    2017-07-01

    Full Text Available The interaction of acetaminophen, a non-substrate anionic ligand, with Aldehyde Dehydrogenase was studied by fluorescence, UV–Vis absorption, and circular dichroism spectroscopies under simulated physiological conditions. The fluorescence spectra and data generated showed that acetaminophen binding to ALDH is purely dynamic quenching mechanism. The acetaminophen-ALDH is kinetically rapid reversible interaction with a binding constant, Ka, of 4.91×103 L mol−1. There was an existence of second binding site of ALDH for acetaminophen at saturating acetaminophen concentration. The binding sites were non-cooperative. The thermodynamic parameters obtained suggest that Van der Waal force and hydrogen bonding played a major role in the binding of acetaminophen to ALDH. The interaction caused perturbation of the ALDH structures with an obvious reduction in the α-helix. The binding distance of 4.43 nm was obtained between Acetaminophen and ALDH. Using Ficoll 400 as macro-viscosogen and glycerol as micro-viscosogen, Stoke-Einstein empirical plot demonstrated that acetaminophen-ALDH binding was diffusion controlled. Molecular docking showed the participation of some amino acids in the complex formation with −5.3 kcal binding energy. With these, ALDH might not an excipient detoxifier of acetaminophen but could be involved in its pegylation/encapsulation.

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

    International Nuclear Information System (INIS)

    Nakamura, Tomofumi; Ichinose, Hirofumi; Wariishi, Hiroyuki

    2010-01-01

    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 + -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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, Tomofumi [Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Fukuoka Institute of Health and Environmental Sciences, 39 Mukaizano, Dazaifu-shi, Fukuoka 818-0135 (Japan); Ichinose, Hirofumi [Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Wariishi, Hiroyuki, E-mail: hirowari@agr.kyushu-u.ac.jp [Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Bio-Architecture Center, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Innovation Center for Medical Redox Navigation, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan)

    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.

  3. Disruption of the coenzyme binding site and dimer interface revealed in the crystal structure of mitochondrial aldehyde dehydrogenase "Asian" variant.

    Science.gov (United States)

    Larson, Heather N; Weiner, Henry; Hurley, Thomas D

    2005-08-26

    Mitochondrial aldehyde dehydrogenase (ALDH2) is the major enzyme that oxidizes ethanol-derived acetaldehyde. A nearly inactive form of the enzyme, ALDH2*2, is found in about 40% of the East Asian population. This variant enzyme is defined by a glutamate to lysine substitution at residue 487 located within the oligomerization domain. ALDH2*2 has an increased Km for its coenzyme, NAD+, and a decreased kcat, which lead to low activity in vivo. Here we report the 2.1 A crystal structure of ALDH2*2. The structure shows a large disordered region located at the dimer interface that includes much of the coenzyme binding cleft and a loop of residues that form the base of the active site. As a consequence of these structural changes, the variant enzyme exhibits rigid body rotations of its catalytic and coenzyme-binding domains relative to the oligomerization domain. These structural perturbations are the direct result of the inability of lysine 487 to form important stabilizing hydrogen bonds with arginines 264 and 475. Thus, the elevated Km for coenzyme exhibited by this variant probably reflects the energetic penalty for reestablishing this site for productive coenzyme binding, whereas the structural alterations near the active site are consistent with the lowered Vmax.

  4. Alcohol and aldehyde dehydrogenase polymorphisms and risk for suicide: a preliminary observation in the Japanese male population.

    Science.gov (United States)

    Hishimoto, A; Fukutake, M; Mouri, K; Nagasaki, Y; Asano, M; Ueno, Y; Nishiguchi, N; Shirakawa, O

    2010-07-01

    Epidemiological studies have shown that excessive alcohol consumption is a potent risk factor to develop suicidal behavior. Genetic factors for suicidal behavior have been observed in family, twin, and adoption studies. Because alcohol dehydrogenase (ADH1B) His47Arg and mitochondrial aldehyde dehydrogenase (ALDH2) Glu487Lys single nucleotide polymorphisms (SNPs), which affect alcohol metabolism, have been reported to exert significant impacts on alcohol consumption and on the risk for alcoholism in East Asia populations, we explored associations of the two functional SNPs with suicide using a case-control study of 283 completed suicides and 319 control subjects in the Japanese population. We found that the inactive ALDH2 allele (487Lys) was significantly less frequent in the completed suicides (19.3%) than in the controls (29.3%), especially in males, whereas this was not the case in females. The males bearing alcoholism-susceptible homozygotes at both loci (inactive ADH1B Arg/Arg and active ALDH2 Glu/Glu genotypes) have a 10 times greater risk for suicide compared with the males bearing alcoholism-protective homozygotes at both loci. Our data show the genetic impact of the two polymorphisms on suicidal behavior in the Japanese population, especially in males. Because we did not verify the daily alcohol consumption, the association of these SNPs with suicide might be due to alcoholism itself. Further studies using case-control subjects, which verifies the details of current and past alcohol consumption and diagnosis for alcoholism, are required to confirm these findings.

  5. Aldehyde dehydrogenase 2 polymorphism for development to hepatocellular carcinoma in East Asian alcoholic liver cirrhosis.

    Science.gov (United States)

    Abe, Hiroshi; Aida, Yuta; Seki, Nobuyoshi; Sugita, Tomonori; Tomita, Yoichi; Nagano, Tomohisa; Itagaki, Munenori; Sutoh, Satoshi; Nagatsuma, Keisuke; Itoh, Kyoko; Matsuura, Tomokazu; Aizawa, Yoshio

    2015-09-01

    We aimed to clarify the influences of aldehyde dehydrogenase 2 (ALDH2), alcohol dehydrogenase 1B (ADH1B) polymorphisms, and ethanol consumption profile to hepatocellular carcinoma (HCC) development in alcoholic liver cirrhosis without chronic hepatitis B and C virus infection (non-B non-C). Of 236 freshly diagnosed non-B non-C alcoholic liver cirrhosis patients, 67 were diagnosed as HCC and the remaining 169 as not having HCC. The relationship between the genetic polymorphisms and development to HCC were evaluated in well-matched patients with HCC (HCC group, n = 67) and without HCC (non-HCC group, n = 67) using propensity scores in age, sex, and prevalence of diabetes mellitus. Daily amount of ethanol consumption was significantly lower (P = 0.005), and consumptive period was significantly longer (P = 0.003) in HCC group than non-HCC group. Of 134 well-matched patients, 113 (84.3%) had ALDH2*1/*1 genotype and 21 (15.7%) had ALDH2*1/*2 genotype. In HCC development, consumptive long period (P = 0.007) and carrying ALDH2*1/*2 genotype (P = 0.026) were identified as significant factors independently participated, while there was no relation to ADH1B polymorphism. In addition, consumptive period was significantly longer in HCC group than non-HCC group in ALDH2*1/*1 genotype patients (P = 0.0005), while there was no difference in profile of ethanol consumption in ALDH2*1/*2 genotype patients. Among HCC group, daily (P = 3.78 × 10(-6) ) and cumulative amount (P = 4.89 × 10(-6) ) of ethanol consumption were significantly higher in ALDH2*1/*1 genotype patients than ALDH2*1/*2 genotype patients. In alcoholic liver cirrhosis, investigations of ALDH2 polymorphism and ethanol consumption profile are useful for prediction of HCC development. © 2015 Journal of Gastroenterology and Hepatology Foundation and Wiley Publishing Asia Pty Ltd.

  6. Alcohol and aldehyde dehydrogenase gene polymorphisms and oropharyngolaryngeal, esophageal and stomach cancers in Japanese alcoholics.

    Science.gov (United States)

    Yokoyama, A; Muramatsu, T; Omori, T; Yokoyama, T; Matsushita, S; Higuchi, S; Maruyama, K; Ishii, H

    2001-03-01

    Alcohol dehydrogenase-2 (ADH2) and aldehyde dehydrogenase-2 (ALDH2) gene polymorphisms play roles in ethanol metabolism, drinking behavior and esophageal carcinogenesis in Japanese; however, the combined influence of ADH2 and ALDH2 genotypes on other aerodigestive tract cancers have not been investigated. ADH2/ALDH2 genotyping was performed on lymphocyte DNA samples from Japanese alcoholic men (526 cancer-free; 159 with solitary or multiple aerodigestive tract cancers, including 33 oropharyngolaryngeal, 112 esophageal, 38 stomach and 22 multiple primary cancers in two or three organs). After adjustment for age, drinking and smoking habits, and ADH2/ALDH2 genotypes, the presence of either ADH2*1/2*1 or ALDH2*1/2*2 significantly increased the risk for oropharyngolaryngeal cancer [odds ratios (ORs), 6.68 with ADH2*1/2*1 and 18.52 with ALDH2*1/2*2] and esophageal cancer (ORs, 2.64 and 13.50, respectively). For patients with both ADH2*1/2*1 and ALDH2*1/2*2, the risks for oropharyngolaryngeal and esophageal cancers were enhanced in a multiplicative fashion (OR = 121.77 and 40.40, respectively). A positive association with ALDH2*1/2*2 alone was observed for stomach cancer patients who also had oropharyngolaryngeal and/or esophageal cancer (OR = 110.58), but it was not observed for those with stomach cancer alone. Furthermore, in the presence of ALDH2*1/2*2, the risks for multiple intra-esophageal cancers (OR = 3.43) and for esophageal cancer with oropharyngolaryngeal and/or stomach cancer (OR = 3.95) were higher than the risks for solitary intra-esophageal cancer and for esophageal cancer alone, but these tendencies were not observed for ADH2*1/2*1 genotype. Alcoholics' population attributable risks due to ADH2/ALDH2 polymorphisms were estimated to be 82.0% for oropharyngolaryngeal cancer and 63.9% for esophageal cancer.

  7. Alcohol and aldehyde dehydrogenase gene polymorphisms influence susceptibility to esophageal cancer in Japanese alcoholics.

    Science.gov (United States)

    Yokoyama, A; Muramatsu, T; Omori, T; Matsushita, S; Yoshimizu, H; Higuchi, S; Yokoyama, T; Maruyama, K; Ishii, H

    1999-11-01

    Studies have consistently demonstrated that inactive aldehyde dehydrogenase-2 (ALDH2), encoded by ALDH2*1/2*2, is closely associated with alcohol-related carcinogenesis. Recently, the contributions of alcohol dehydrogenase-2 (ADH2) polymorphism to alcoholism, esophageal cancer, and the flushing response have also been described. To determine the effects of ALDH2 and ADH2 genotypes in genetically based cancer susceptibility, lymphocyte DNA samples from 668 Japanese alcoholic men more than 40 years of age (91 with and 577 without esophageal cancer) were genotyped and the results were expressed as odds ratios (ORs). This study also tested 82 of the alcoholics with esophageal cancer to determine whether cancer susceptibility is associated with patients' responses to simple questions about current or former flushing after drinking a glass of beer. The frequencies of ADH2*1/2*1 and ALDH2*1/2*2 were significantly higher in alcoholics with, than in those without, esophageal cancer (0.473 vs. 0.289 and 0.560 vs. 0.099, respectively). After adjustment for drinking and smoking, the analysis showed significantly increased cancer risk for alcoholics with either ADH2*1/2*I (OR = 2.03) or ALDH2*1/2*2 (OR = 12.76). For those having ADH2*1/2*1 combined with ALDH2*1/2*2, the esophageal cancer risk was enhanced in a multiplicative fashion (OR = 27.66). Responses to flushing questions showed that only 47.8% of the ALDH2*1/2*2 heterozygotes with ADH2*1/ 2*1, compared with 92.3% of those with ALDH2*1/2*2 and the ADH2*2 allele, reported current or former flushing. Genotyping showed that for alcoholics who reported ever flushing, the questionnaire was 71.4% correct in identifying ALDH2*1/2*2 and 87.9% correct in identifying ALDH2*1/2*1. Japanese alcoholics can be divided into cancer susceptibility groups on the basis of their combined ADH2 and ALDH2 genotypes. The flushing questionnaire can predict high risk ALDH2*1/2*2 fairly accurately in persons with ADH2*2 allele, but a reliable

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

    Directory of Open Access Journals (Sweden)

    Jennifer M. Petrosino

    2014-03-01

    Full Text Available 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.

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

    Science.gov (United States)

    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.

  10. Aldehyde dehydrogenase 1A1 circumscribes high invasive glioma cells and predicts poor prognosis

    Science.gov (United States)

    Xu, Sen-Lin; Liu, Sha; Cui, Wei; Shi, Yu; Liu, Qin; Duan, Jiang-Jie; Yu, Shi-Cang; Zhang, Xia; Cui, You-Hong; Kung, Hsiang-Fu; Bian, Xiu-Wu

    2015-01-01

    Glioma is the most aggressive brain tumor with high invasiveness and poor prognosis. More reliable, sensitive and practical biomarkers to reveal glioma high invasiveness remain to be explored for the guidance of therapy. We herein evaluated the diagnostic and prognostic value of aldehyde dehydrogenase 1A1 (ALDH1A1) in the glioma specimens from 237 patients, and found that ADLH1A1 was frequently overexpressed in the high-grade glioma (WHO grade III-IV) as compared to the low-grade glioma (WHO grade I-II) patients. The tumor cells with ALDH1A1 expression were more abundant in the region between tumor and the borderline of adjacent tissue as compared to the central part of the tumor. ALDH1A1 overexpression was associated with poor differentiation and dismal prognosis. Notably, the overall and disease-free survivals of the patients who had ALDH1A1+ tumor cells sparsely located in the adjacent tissue were much worse. Furthermore, ALDH1A1 expression was correlated with the “classical-like” (CL) subtype as we examined GBM specimens from 72 patients. Multivariate Cox regression analysis revealed that ALDH1A1 was an independent marker for glioma patients’ outcome. Mechanistically, both in vitro and in vivo studies revealed that ALDH1A1+ cells isolated from either a glioblastoma cell line U251 or primary glioblastoma cells displayed significant invasiveness, clonogenicity, and proliferation as compared to ALDH1A1- cells, due to increased levels of mRNA and protein for matrix metalloproteinase 2, 7 and 9 (MMP2, MMP7 and MMP9). These results indicate that ALDH1A1+ cells contribute to the progression of glioma including invasion, proliferation and poor prognosis, and suggest that targeting ALDH1A1 may have important implications for the treatment of highly invasive glioma. PMID:26101711

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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

  13. Aldehyde oxidase activity in fresh human skin.

    Science.gov (United States)

    Manevski, Nenad; Balavenkatraman, Kamal Kumar; Bertschi, Barbara; Swart, Piet; Walles, Markus; Camenisch, Gian; Schiller, Hilmar; Kretz, Olivier; Ling, Barbara; Wettstein, Reto; Schaefer, Dirk J; Pognan, Francois; Wolf, Armin; Litherland, Karine

    2014-12-01

    Human aldehyde oxidase (AO) is a molybdoflavoenzyme that commonly oxidizes azaheterocycles in therapeutic drugs. Although high metabolic clearance by AO resulted in several drug failures, existing in vitro-in vivo correlations are often poor and the extrahepatic role of AO practically unknown. This study investigated enzymatic activity of AO in fresh human skin, the largest organ of the body, frequently exposed to therapeutic drugs and xenobiotics. Fresh, full-thickness human skin was obtained from 13 individual donors and assayed with two specific AO substrates: carbazeran and zoniporide. Human skin explants from all donors metabolized carbazeran to 4-hydroxycarbazeran and zoniporide to 2-oxo-zoniporide. Average rates of carbazeran and zoniporide hydroxylations were 1.301 and 0.164 pmol⋅mg skin(-1)⋅h(-1), resulting in 13 and 2% substrate turnover, respectively, after 24 hours of incubation with 10 μM substrate. Hydroxylation activities for the two substrates were significantly correlated (r(2) = 0.769), with interindividual variability ranging from 3-fold (zoniporide) to 6-fold (carbazeran). Inclusion of hydralazine, an irreversible inhibitor of AO, resulted in concentration-dependent decrease of hydroxylation activities, exceeding 90% inhibition of carbazeran 4-hydroxylation at 100 μM inhibitor. Reaction rates were linear up to 4 hours and well described by Michaelis-Menten enzyme kinetics. Comparison of carbazeran and zoniporide hydroxylation with rates of triclosan glucuronidation and sulfation and p-toluidine N-acetylation showed that cutaneous AO activity is comparable to tested phase II metabolic reactions, indicating a significant role of AO in cutaneous drug metabolism. To our best knowledge, this is the first report of AO enzymatic activity in human skin. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

  14. Gastric cytoprotective activity of ilicic aldehyde: structure-activity relationships.

    Science.gov (United States)

    Donadel, Osvaldo J; Guerreiro, Eduardo; María, Alejandra O; Wendel, Graciela; Enriz, Ricardo D; Giordano, Oscar S; Tonn, Carlos E

    2005-08-01

    A series of sesquiterpene compounds possessing both eudesmane and eremophilane skeletons were tested as gastric cytoprotective agents on male Wistar rats. The presence of an alpha,beta-unsaturated aldehyde on the C-7 side chain together with a hydroxyl group at C-4 is the requirement for the observed antiulcerogenic activity. In an attempt to establish new molecular structural requirements for this gastric cytoprotective activity, a structure-activity study was performed.

  15. Alcohol Dehydrogenase-1B (rs1229984 and Aldehyde Dehydrogenase-2 (rs671 Genotypes Are Strong Determinants of the Serum Triglyceride and Cholesterol Levels of Japanese Alcoholic Men.

    Directory of Open Access Journals (Sweden)

    Akira Yokoyama

    Full Text Available 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.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.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.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-metabolizing ADH1B was associated with lower serum LDL

  16. Improved tolerance to various abiotic stresses in transgenic sweet potato (Ipomoea batatas) expressing spinach betaine aldehyde dehydrogenase.

    Science.gov (United States)

    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

  17. Improved Tolerance to Various Abiotic Stresses in Transgenic Sweet Potato (Ipomoea batatas) Expressing Spinach Betaine Aldehyde Dehydrogenase

    Science.gov (United States)

    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

  18. Improved tolerance to various abiotic stresses in transgenic sweet potato (Ipomoea batatas expressing spinach betaine aldehyde dehydrogenase.

    Directory of Open Access Journals (Sweden)

    Weijuan Fan

    Full Text Available 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

  19. Genome-wide characterization and expression analysis of the aldehyde dehydrogenase (ALDH) gene superfamily under abiotic stresses in cotton.

    Science.gov (United States)

    Guo, Xinlei; Wang, Yuanyuan; Lu, Hejun; Cai, Xiaoyan; Wang, Xingxing; Zhou, Zhongli; Wang, Chunying; Wang, Yuhong; Zhang, Zhenmei; Wang, Kunbo; Liu, Fang

    2017-09-10

    In plants, aldehyde dehydrogenases (ALDHs) function as 'aldehyde scavengers' by removing reactive aldehydes and thus play important roles in stress responses. To date, 30 ALDHs have been identified in Gossypium raimondii, whereas ALDHs have not been studied in Gossypium arboreum or in tetraploid cotton. In this study, we identified 30, 59 and 59 aldehyde dehydrogenase (ALDH) genes from G. arboreum, G. hirsutum and G. barbadense, respectively. Gene structure analysis revealed that members of the same family exhibit similar exon-intron structures and structural domains, and all members of the ALDH18 family possess a distinct AA-kinase domain. Synteny analysis showed that segmental and tandem duplications have played an important role in the expansion and evolution of ALDHs in cotton. Phylogenetic and synteny analysis between G. arboreum and G. raimondii demonstrated that all GaALDHs and GrALDHs are orthologous and that most GaALDHs are located in syntenic blocks corresponding to those of G. raimondii, implying that these genes appeared before the divergence of G. arboreum and G. raimondii and that no expansion of the ALDH superfamily has occurred in these two cotton species. Quantitative real-time PCR analysis revealed that the majority of GaALDHs and GhALDHs are up-regulated under conditions of high salinity and drought, indicating that these genes may be stress responsive. The findings of this study, based on genome-wide identification of ALDHs in Gossypium and analysis of their evolution and expression, provide a foundation for further analysis of ALDHs and suggest potential target genes for improving stress resistance in cotton. Copyright © 2017. Published by Elsevier B.V.

  20. YLL056C from Saccharomyces cerevisiae encodes a novel protein with aldehyde reductase activity.

    Science.gov (United States)

    Wang, Han-Yu; Xiao, Di-Fan; Zhou, Chang; Wang, Lin-Lu; Wu, Lan; Lu, Ya-Ting; Xiang, Quan-Ju; Zhao, Ke; Li, Xi; Ma, Meng -Gen

    2017-06-01

    The short-chain dehydrogenase/reductase (SDR) family, the largest family in dehydrogenase/reductase superfamily, is divided into "classical," "extended," "intermediate," "divergent," "complex," and "atypical" groups. Recently, several open reading frames (ORFs) were characterized as intermediate SDR aldehyde reductase genes in Saccharomyces cerevisiae. However, no functional protein in the atypical group has been characterized in S. cerevisiae till now. Herein, we report that an uncharacterized ORF YLL056C from S. cerevisiae was significantly upregulated under high furfural (2-furaldehyde) or 5-(hydroxymethyl)-2-furaldehyde concentrations, and transcription factors Yap1p, Hsf1p, Pdr1/3p, Yrr1p, and Stb5p likely controlled its upregulated transcription. This ORF indeed encoded a protein (Yll056cp), which was grouped into the atypical subgroup 7 in the SDR family and localized to the cytoplasm. Enzyme activity assays showed that Yll056cp is not a quinone or ketone reductase but an NADH-dependent aldehyde reductase, which can reduce at least seven aldehyde compounds. This enzyme showed the best Vmax, Kcat, and Kcat/Km to glycolaldehyde, but the highest affinity (Km) to formaldehyde. The optimum pH and temperature of this enzyme was pH 6.5 for reduction of glycolaldehyde, furfural, formaldehyde, butyraldehyde, and propylaldehyde, and 30 °C for reduction of formaldehyde or 35 °C for reduction of glycolaldehyde, furfural, butyraldehyde, and propylaldehyde. Temperature and pH affected stability of this enzyme and this influence varied with aldehyde substrate. Metal ions, salts, and chemical protective additives, especially at high concentrations, had different influence on enzyme activities for reduction of different aldehydes. This research provided guidelines for study of more uncharacterized atypical SDR enzymes from S. cerevisiae and other organisms.

  1. Immunohistochemical analysis of aldehyde dehydrogenase isoforms and their association with estrogen-receptor status and disease progression in breast cancer

    Directory of Open Access Journals (Sweden)

    Opdenaker LM

    2014-12-01

    Full Text Available Lynn M Opdenaker,1,2 Kimberly M Arnold,1,3 Ryan T Pohlig,3,4 Jayasree S Padmanabhan,1 Daniel C Flynn,1,3 Jennifer Sims-Mourtada1–3 1Center for Translational Cancer Research, Helen F Graham Cancer Center, Christiana Care Health Services, Inc., Newark, Delaware, USA; 2Department of Biological Sciences, 3Department of Medical Laboratory Sciences, 4Biostatistics Core Facility, University of Delaware, Newark, Delaware, USA Abstract: In many types of tumors, especially breast tumors, aldehyde dehydrogenase (ALDH activity has been used to identify cancer stem-like cells within the tumor. The presence and quantity of these cells are believed to predict the response of tumors to chemotherapy. Therefore, identification and eradication of these cells would be necessary to cure the patient. However, there are 19 different ALDH isoforms that could contribute to the enzyme activity. ALDH1A1 and ALDH1A3 are among the isoforms mostly responsible for the increased ALDH activity observed in these stem-like cells, although the main isoforms vary in different tissues and tumor types. In the study reported here, we attempted to determine if ALDH1A1 or ALDH1A3, specifically, correlate with tumor stage, grade, and hormone-receptor status in breast-cancer patients. While there was no significant correlation between ALDH1A1 and any of the parameters tested, we were able to identify a positive correlation between ALDH1A3 and tumor stage in triple-negative cancers. In addition, ALDH1A3 was negatively correlated with estrogen-receptor status. Our data suggest that ALDH1A3 could be utilized as a marker to identify stem-like cells within triple-negative tumors. Keywords: breast tumor, ALDH, ALDH1A1, ALDH1A3, stem-like cells, triple-negative cancer

  2. Plastid-expressed betaine aldehyde dehydrogenase gene in carrot cultured cells, roots, and leaves confers enhanced salt tolerance.

    Science.gov (United States)

    Kumar, Shashi; Dhingra, Amit; Daniell, Henry

    2004-09-01

    Salinity is one of the major factors that limits geographical distribution of plants and adversely affects crop productivity and quality. We report here high-level expression of betaine aldehyde dehydrogenase (BADH) in cultured cells, roots, and leaves of carrot (Daucus carota) via plastid genetic engineering. Homoplasmic transgenic plants exhibiting high levels of salt tolerance were regenerated from bombarded cell cultures via somatic embryogenesis. Transformation efficiency of carrot somatic embryos was very high, with one transgenic event per approximately seven bombarded plates under optimal conditions. In vitro transgenic carrot cells transformed with the badh transgene were visually green in color when compared to untransformed carrot cells, and this offered a visual selection for transgenic lines. BADH enzyme activity was enhanced 8-fold in transgenic carrot cell cultures, grew 7-fold more, and accumulated 50- to 54-fold more betaine (93-101 micromol g(-1) dry weight of beta-Ala betaine and Gly betaine) than untransformed cells grown in liquid medium containing 100 mm NaCl. Transgenic carrot plants expressing BADH grew in the presence of high concentrations of NaCl (up to 400 mm), the highest level of salt tolerance reported so far among genetically modified crop plants. BADH expression was 74.8% in non-green edible parts (carrots) containing chromoplasts, and 53% in proplastids of cultured cells when compared to chloroplasts (100%) in leaves. Demonstration of plastid transformation via somatic embryogenesis utilizing non-green tissues as recipients of foreign DNA for the first time overcomes two of the major obstacles in extending this technology to important crop plants.

  3. Is aldehyde dehydrogenase 2 a credible genetic instrument for alcohol use in Mendelian randomization analysis in Southern Chinese men?

    Science.gov (United States)

    Au Yeung, Shiu Lun; Jiang, ChaoQiang; Cheng, Kar Keung; Liu, Bin; Zhang, WeiSen; Lam, Tai Hing; Leung, Gabriel M; Schooling, C Mary

    2013-02-01

    Mendelian randomization studies provide a means of assessing causal relations without interventions, but require valid genetic instruments. We assessed the credibility of aldehyde dehydrogenase 2 (ALDH2) as a genetic instrument for alcohol use in Southern Chinese men. We genotyped the single nucleotide polymorphism rs671 of ALDH2 in 4867 men from the Guangzhou Biobank Cohort Study. We used linear regression to assess the strength of the association of ALDH2 variants with alcohol use, whether ALDH2 variants were independently associated with socio-economic position or other potential confounders and whether associations of ALDH2 variants with cardiovascular risk factors (systolic and diastolic blood pressure, HDL- and LDL-cholesterol, fasting glucose), triglycerides, body mass index, self reported cardiovascular disease, self-reported ischaemic heart disease, cognitive function (delayed 10-word recall and Mini Mental State Examination score) and liver function (alanine transaminase and aspartate transaminase) were fully mediated by alcohol use. The minor allele frequency (A) of ALDH2 was 0.29. The F statistic for ALDH2 variants was 75.0, suggesting that substantial weak instrument bias is unlikely. ALDH2 variants were not associated with socio-economic position, smoking or physical activity. ALDH2 variants were only associated with diastolic blood pressure and HDL-cholesterol, but these genetic associations with blood pressure and HDL-cholesterol were attenuated after adjusting for alcohol use, suggesting the apparent genetic associations were possibly mediated by alcohol use. ALDH2 variants are a credible genetic instrument for Mendelian randomization studies of alcohol use and many attributes of health in Southern Chinese men.

  4. [Effects of panthenol and carnitine on aldehyde metabolic enzymes in rats with tetrachloromethane-induced liver injury].

    Science.gov (United States)

    Satanovskaia, V I; Pron'ko, P S; Gaĭshmanova, A V; Miskevich, D A

    2009-01-01

    Tetrachloromethane (2 g/kg, intragastric) produced a decrease in the activity of NAD- and NADH- dependent aldehyde dehydrogenases with high Km for aldehydes in rat liver. Panthenol and L-carnitine administered separately normalized the activity of aldehyde dehydrogenases, while a combination of the drugs did not produce any significant effect.

  5. Kinetic and biophysical investigation of the inhibitory effect of caffeine on human salivary aldehyde dehydrogenase: Implications in oral health and chemotherapy

    Science.gov (United States)

    Laskar, Amaj Ahmed; Alam, Md Fazle; Ahmad, Mohammad; Younus, Hina

    2018-04-01

    Human salivary aldehyde dehydrogenase (hsALDH) is primarily a class 3 ALDH (ALDH3A1), and is an important antioxidant enzyme present in the saliva which maintains healthy oral cavity. It detoxifies toxic aldehydes into non-toxic carboxylic acids in the oral cavity. Reduced level of hsALDH activity is a risk factor for oral cancer development. It is involved in the resistance of certain chemotherapeutic drugs. Coffee has been reported to affect the activity of salivary ALDH. In this study, the effect of caffeine on the activity (dehydrogenase and esterase) of hsALDH was investigated. The binding of caffeine to hsALDH was studied using different biophysical methods and molecular docking analysis. Caffeine was found to inhibit both crude and purified hsALDH. The Km increased and the Vmax decreased showing a mixed type of inhibition. Caffeine decreased the nucleophilicity of the catalytic cysteine residue. It binds to the active site of ALDH3A1 by forming a complex through non-covalent interactions with some highly conserved amino acid residues. It partially alters the secondary structure of the enzyme. Therefore, it is very likely that caffeine binds and inhibits the activity of hsALDH by decreasing substrate binding affinity and the catalytic efficiency of the enzyme. The study indicates that oral intake of caffeine may have a harmful effect on the oral health and may increase the risk of carcinogenesis through the inhibition of this important enzyme. Further, the inactivation of oxazaphosphorine based chemotherapeutic drugs by ALDH3A1 may be prevented by using caffeine as an adjuvant during medication which is expected to increase the sensitivity of these drugs through its inhibitory effect on the enzyme.

  6. 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)

    Science.gov (United States)

    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...

  7. The aldehyde dehydrogenase, AldA, is essential for L-1,2-propanediol utilization in laboratory-evolved Escherichia coli

    DEFF Research Database (Denmark)

    Aziz, Ramy K.; Monk, Jonathan M.; Andrews, Kathleen A.

    2017-01-01

    Most Escherichia coli strains are naturally unable to grow on 1,2-propanediol (PDO) as a sole carbon source. Recently, however, a K-12 descendent E. coli strain was evolved to grow on 1,2-PDO, and it was hypothesized that this evolved ability was dependent on the aldehyde dehydrogenase, AldA, whi...

  8. Human class I alcohol dehydrogenases catalyze the interconversion of alcohols and aldehydes in the metabolism of dopamine.

    Science.gov (United States)

    Mårdh, G; Vallee, B L

    1986-11-18

    The class I human liver alcohol dehydrogenases (ADHs) catalyze the interconversion of the intermediary alcohols and aldehydes of dopamine metabolism in vitro, whereas those of the class II and class III do not. The individual, homogeneous class I isozymes oxidize (3,4-dihydroxyphenyl)ethanol and (4-hydroxy-3-methoxyphenyl)ethanol (HMPE) and ethanol with kcat/Km values in the range from 16 to 240 mM-1 min-1 and from 16 to 66 mM-1 min-1, respectively. They reduce the corresponding dopamine aldehydes (3,4-dihydroxyphenyl)acetaldehyde and (4-hydroxy-3-methoxyphenyl)acetaldehyde (HMPAL) with kcat/Km values varying from 7800 to 190,000 mM-1 min-1, considerably more efficient than the reduction of acetaldehyde with kcat/Km values from 780 to 4900 mM-1 min-1. For beta 1 gamma 2 ADH, ethanol competes with HMPE oxidation with a Ki of 23 microM. In addition, 1,10-phenanthroline inhibits HMPE oxidation and HMPAL reduction with Ki values of 20 microM and 12 microM, respectively, both quite similar to that for ethanol, Ki = 22 microM. Thus, both ethanol/acetaldehyde and the dopamine intermediates compete for the same site of ADH, a basis for the ethanol-induced in vivo alterations of dopamine metabolism.

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

    Science.gov (United States)

    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. The ORF slr0091 of Synechocystis sp. PCC6803 encodes a high-light induced aldehyde dehydrogenase converting apocarotenals and alkanals

    KAUST Repository

    Trautmann, Danika

    2013-07-05

    Oxidative cleavage of carotenoids and peroxidation of lipids lead to apocarotenals and aliphatic aldehydes called alkanals, which react with vitally important compounds, promoting cytotoxicity. Although many enzymes have been reported to deactivate alkanals by converting them into fatty acids, little is known about the mechanisms used to detoxify apocarotenals or the enzymes acting on them. Cyanobacteria and other photosynthetic organisms must cope with both classes of aldehydes. Here we report that the Synechocystis enzyme SynAlh1, encoded by the ORF slr0091, is an aldehyde dehydrogenase that mediates oxidation of both apocarotenals and alkanals into the corresponding acids. Using a crude lysate of SynAlh1-expressing Escherichia coli cells, we show that SynAlh1 converts a wide range of apocarotenals and alkanals, with a preference for apocarotenals with defined chain lengths. As suggested by in vitro incubations and using engineered retinal-forming E. coli cells, we found that retinal is not a substrate for SynAlh1, making involvement in Synechocystis retinoid metabolism unlikely. The transcript level of SynAlh1 is induced by high light and cold treatment, indicating a role in the stress response, and the corresponding gene is a constituent of a stress-related operon. The assumptions regarding the function of SynAlh are further supported by the surprisingly high homology to human and plant aldehyde dehydrogenase that have been assigned to aldehyde detoxification. SynAlh1 is the first aldehyde dehydrogenase that has been shown to form both apocarotenoic and fatty acids. This dual function suggests that its eukaryotic homologs may also be involved in apocarotenal metabolism, a function that has not been considered so far. Aldehyde dehydrogenases play an important role in detoxification of reactive aldehydes. Here, we report on a cyanbacterial enzyme capable in converting two classes of lipid-derived aldehydes, apocaotenals and alkanals. The corresponding gene is a

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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

  13. Direct electron transfer-based bioanodes for ethanol biofuel cells using PQQ-dependent alcohol and aldehyde dehydrogenases

    International Nuclear Information System (INIS)

    Aquino Neto, Sidney; Suda, Emily L.; Xu, Shuai; Meredith, Matthew T.; De Andrade, Adalgisa R.; Minteer, Shelley D.

    2013-01-01

    This paper compares the performance of a DET (direct electron transfer) bioanode containing both PQQ-ADH (pyrroloquinoline quinone-dependent alcohol dehydrogenase) and PQQ-AldDH (PQQ-dependent aldehyde dehydrogenase) immobilized onto different modified electrode surfaces employing either a tetrabutylammonium (TBAB)-modified Nafion ® membrane polymer or polyamidoamine (PAMAM) dendrimers for the enzyme immobilization. The electrochemical characterization showed that the prepared bioelectrodes were able to undergo DET onto glassy carbon surface in the presence as well as the absence of multi-walled carbon nanotubes (MWCNTs); also, in the latter case a relevant shift in the oxidation peak of about 180 mV vs. saturated calomel electrode (SCE) was observed. A very similar redox potential was achieved with the self-assembled bioelectrode prepared onto modified-gold surfaces with dendrimers, indicating that both methodologies provide an environment that enables the PQQ-enzymes to undergo DET. The biofuel cell tests confirmed the ease of the DET process and the enhanced performance in the presence of the carbon nanotubes. Considering the bioanodes prepared with PAMAM dendrimers, the power density values vary from 19.4 μW cm −2 without MWCNTs to 25.7 μW cm −2 in the presence of MWCNTs. Similarly, with the bioanodes prepared with the TBAB-modified-Nafion ® polymer, the results indicate power densities of 27.9 and 38.4 μW cm −2 respectively. These electrode modifications represent effective methods for immobilization and direct electrical connection of quinohemoproteins to electrode surfaces.

  14. The Short-Chain Alcohol Dehydrogenase ABA2 Catalyzes the Conversion of Xanthoxin to Abscisic AldehydeW⃞

    Science.gov (United States)

    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

  15. The correlation between aldehyde dehydrogenase-1A1 level and tumor shrinkage after preoperative chemoradiation in locally advanced rectal cancer

    Directory of Open Access Journals (Sweden)

    Rhandyka Rafli

    2015-12-01

    Full Text Available This study was performed to determine the correlation between aldehyde dehydrogenase-1A1 (ALDH1A1 level and tumor shrinkage after chemoradiation in locally advanced rectal cancer. This is a retrospective study of 14 locally advanced rectal cancer patients with long course neoadjuvant chemoradiation. ALDH1A1 level was measured using ELISA from paraffin embedded tissue. Tumor shrinkage was measured from computed tomography (CT scan or magnetic resonance imaging (MRI based on Response Evaluation Criteria in Solid Tumor v1.1 (RECIST v1.1. The mean of ALDH1A1 level was 9.014 ± 3.3 pg/mL and the mean of tumor shrinkage was 7.89 ± 35.7%. Partial response proportion was 28.6%, stable disease proportion was 50% and progressive disease proportion was 21.4%. There was a significant strong negative correlation (r = –0.890, plt; 0.001 between ALDH1A1 and tumor shrinkage. In conclusion, tumor shrinkage in locally advanced rectal cancer after preoperative chemoradiation was influenced by ALDH1A1 level. Higher level of ALDH1A1 suggests decreased tumor shrinkage after preoperative chemoradiation.

  16. Interaction of the SPG21 protein ACP33/maspardin with the aldehyde dehydrogenase ALDH16A1

    Science.gov (United States)

    2017-01-01

    Mast syndrome (SPG21) is an autosomal-recessive complicated form of hereditary spastic paraplegia characterized by dementia, thin corpus callosum, white matter abnormalities, and cerebellar and extrapyramidal signs in addition to spastic paraparesis. A nucleotide insertion resulting in premature truncation of the SPG21 gene product acidic cluster protein 33 (ACP33)/maspardin underlies this disorder, likely causing loss of protein function. However, little is known about the function of maspardin. Here, we report that maspardin localizes prominently to cytoplasm as well as to membranes, possibly at trans-Golgi network/late endosomal compartments. Immunoprecipitation of maspardin with identification of coprecipitating proteins by mass spectrometry revealed the aldehyde dehydrogenase ALDH16A1 as an interacting protein. This interaction was confirmed using overexpressed proteins as well as by fusion protein pull down experiments, and these proteins colocalized in cells. Further studies of the function of ALDH16A1 and the role of the maspardin–ALDH16A1 interaction in neuronal cells may clarify the cellular pathogenesis of Mast syndrome. PMID:19184135

  17. The aldehyde dehydrogenase 2 polymorphisms on neuropsychological performance in bipolar II disorder with or without comorbid anxiety disorder.

    Science.gov (United States)

    Lu, Ru-Band; Chang, Yun-Hsuan; Wang, Tzu-Yun; Lee, Sheng-Yu; Chen, Po See; Yang, Yen Kuang

    2018-01-01

    Anxiety disorders (ADs), the most common comorbid illnesses with bipolar disorder (BP) has been reported to associate with dopamine system. Dopamine, metabolized to 3,4-dihydroxyphenylacetic acid (DOPAC) by aldehyde dehydrogenase 2 (ALDH2), and the distribution of the ALDH2*1/*1, and ALDH2*1/*2+ALDH*2/*2 alleles in the Han Chinese general population is relatively equal. The association between dopamine metabolic enzymes and cognitive performance in patients with bipolar II disorder (BP-II) comorbid with AD is unclear. This study proposed to explore the role of ALDH2 polymorphisms on neuropsychological performance between BP-II comorbid with or without AD. One hundred ninety-seven BP-II patients with and without a comorbid AD were recruited and compared with 130 healthy controls (HCs). A polymerase chain reaction and a restriction fragment length polymorphism analysis were used to determine genotypes for ALDH2, and study participants underwent neuropsychological tests. An interaction between AD comorbidity and the ALDH2 polymorphisms was found in different domain of cognitive dysfunction in the BP-II patients. The ALDH2 polymorphisms might have different effects on the neuropsychological performance of BP-II patients with and without comorbid AD.

  18. The expression of aldehyde dehydrogenase 1 (ALDH1) in ovarian carcinomas and its clinicopathological associations: a retrospective study

    International Nuclear Information System (INIS)

    Huang, Ruixia; Li, Xiaoran; Holm, Ruth; Trope, Claes G.; Nesland, Jahn M.; Suo, Zhenhe

    2015-01-01

    Aldehyde dehydrogenase 1 (ALDH1) is widely used as a specific cancer stem cell marker in a variety of cancers, and may become a promising target for cancer therapy. However, the role of its expression in tumor cells and the microenvironment in different cancers is still controversial. To clarify the clinicopathological effect of ALDH1 expression in ovarian carcinoma, a series of 248 cases of paraffin-embedded formalin fixed ovarian carcinoma tissues with long term follow-up information were studied by immunohistochemistry. The immunostaining of ALDH1was variably detected in both tumor cells and the stromal cells, although the staining in tumor cells was not as strong as that in stromal cells. Statistical analyses showed that high ALDH1 expression in tumor cells was significantly associated with histological subtypes, early FIGO stage, well differentiation grade and better survival probability (p < 0.05). The expression of ALDH1 in the stromal cells had no clinicopathological associations in the present study (p > 0.05). High expression of cancer stem cell marker ALDH1 in ovarian carcinoma cells may thus portend a favorable prognosis, but its expression in tumor microenvironment may have no role in tumor behavior of ovarian carcinomas. More studies are warranted to find out the mechanisms for this

  19. Ovarian cancer stem cells are enriched in side population and aldehyde dehydrogenase bright overlapping population.

    Directory of Open Access Journals (Sweden)

    Kazuyo Yasuda

    Full Text Available Cancer stem-like cells (CSCs/cancer-initiaiting cells (CICs are defined as a small population of cancer cells that have self-renewal capacity, differentiation potential and high tumor-initiating ability. CSCs/CICs of ovarian cancer have been isolated by side population (SP analysis, ALDEFLUOR assay and using cell surface markers. However, these approaches are not definitive markers for CSCs/CICs, and it is necessary to refine recent methods for identifying more highly purified CSCs/CICs. In this study, we analyzed SP cells and aldehyde dehydrogenese bright (ALDH(Br cells from ovarian cancer cells. Both SP cells and ALDH(Br cells exhibited higher tumor-initiating ability and higher expression level of a stem cell marker, sex determining region Y-box 2 (SOX2, than those of main population (MP cells and ALDH(Low cells, respectively. We analyzed an SP and ALDH(Br overlapping population (SP/ALDH(Br, and the SP/ALDH(Br population exhibited higher tumor-initiating ability than that of SP cells or ALDH(Br cells, enabling initiation of tumor with as few as 10(2 cells. Furthermore, SP/ADLH(Br population showed higher sphere-forming ability, cisplatin resistance, adipocyte differentiation ability and expression of SOX2 than those of SP/ALDH(Low, MP/ALDH(Br and MP/ALDH(Low cells. Gene knockdown of SOX2 suppressed the tumor-initiation of ovarian cancer cells. An SP/ALDH(Br population was detected in several gynecological cancer cells with ratios of 0.1% for HEC-1 endometrioid adenocarcinoma cells to 1% for MCAS ovary mucinous adenocarcinoma cells. Taken together, use of the SP and ALDH(Br overlapping population is a promising approach to isolate highly purified CSCs/CICs and SOX2 might be a novel functional marker for ovarian CSCs/CICs.

  20. Inhibition of human alcohol and aldehyde dehydrogenases by aspirin and salicylate: assessment of the effects on first-pass metabolism of ethanol.

    Science.gov (United States)

    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. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. 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

    DEFF Research Database (Denmark)

    Holm, Niels Bjerre; Noble, Carolina; Linnet, Kristian

    2016-01-01

    +)-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...... 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...... 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...

  2. INFLUENCE OF SELECTED PHARMACEUTICALS ON ACTIVATED SLUDGE DEHYDROGENASE ACTIVITY

    Directory of Open Access Journals (Sweden)

    Agnieszka Tomska

    2016-06-01

    The aim of this work was to evaluate the effect of selected antibiotics - sulfanilamide and erythromycin on activated sludge dehydrogenase activity with use of trifenyltetrazolinum chloride (TTC test. Dehydrogenases activity is an indicator of biochemical activity of microorganisms present in activated sludge or the ability to degrade organic compounds in waste water. TTC test is particularly useful for the regularity of the course of treatment, in which the presence of inhibitors of biochemical reactions and toxic compounds are present. It was observed that the dehydrogenase activity decreases with the increase of a antibiotics concentration. The lowest value of the dehydrogenase activity equal to 32.4 μmol TF / gMLSS obtained at sulfanilamide concentration 150mg / l. For this sample, an inhibition of dehydrogenase activity was 31%.

  3. Genome-wide characterization of the aldehyde dehydrogenase gene superfamily in soybean and its potential role in drought stress response.

    Science.gov (United States)

    Wang, Wei; Jiang, Wei; Liu, Juge; Li, Yang; Gai, Junyi; Li, Yan

    2017-07-07

    Aldehyde dehydrogenases (ALDHs) represent a group of enzymes that detoxify aldehydes by facilitating their oxidation to carboxylic acids, and have been shown to play roles in plant response to abiotic stresses. However, the comprehensive analysis of ALDH superfamily in soybean (Glycine max) has been limited. In present study, a total of 53 GmALDHs were identified in soybean, and grouped into 10 ALDH families according to the ALDH Gene Nomenclature Committee and phylogenetic analysis. These groupings were supported by their gene structures and conserved motifs. Soybean ALDH superfamily expanded mainly by whole genome duplication/segmental duplications. Gene network analysis identified 1146 putative co-functional genes of 51 GmALDHs. Gene Ontology (GO) enrichment analysis suggested the co-functional genes of these 51 GmALDHs were enriched (FDR soybean tissues. The expression levels of 13 GmALDHs were significantly up-regulated and 14 down-regulated in response to water deficit. The occurrence frequencies of three drought-responsive cis-elements (ABRE, CRT/DRE, and GTGCnTGC/G) were compared in GmALDH genes that were up-, down-, or non-regulated by water deficit. Higher frequency of these three cis-elements was observed for the group of up-regulated GmALDH genes as compared to the group of down- or non- regulated GmALDHs by drought stress, implying their potential roles in the regulation of soybean response to drought stress. A total of 53 ALDH genes were identified in soybean genome and their phylogenetic relationships and duplication patterns were analyzed. The potential functions of GmALDHs were predicted by analyses of their co-functional gene networks, gene expression profiles, and cis-regulatory elements. Three GmALDH genes, including GmALDH3H2, GmALDH12A2 and GmALDH18B3, were highly induced by drought stress in soybean leaves. Our study provides a foundation for future investigations of GmALDH gene function in soybean.

  4. Metabolism of trans, trans-muconaldehyde, a cytotoxic metabolite of benzene, in mouse liver by alcohol dehydrogenase Adh1 and aldehyde reductase AKR1A4

    International Nuclear Information System (INIS)

    Short, Duncan M.; Lyon, Robert; Watson, David G.; Barski, Oleg A.; McGarvie, Gail; Ellis, Elizabeth M.

    2006-01-01

    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 max of 2141 ± 500 nmol/min/mg and a K m of 11 ± 4 μM. This enzyme was inhibited by pyrazole with a K I of 3.1 ± 0.57 μM. 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 max of 115 nmol/min/mg and a K m of 15 ± 2 μM and was not inhibited by pyrazole

  5. Role of a membrane-bound aldehyde dehydrogenase complex AldFGH in acetic acid fermentation with Acetobacter pasteurianus SKU1108.

    Science.gov (United States)

    Yakushi, Toshiharu; Fukunari, Seiya; Kodama, Tomohiro; Matsutani, Minenosuke; Nina, Shun; Kataoka, Naoya; Theeragool, Gunjana; Matsushita, Kazunobu

    2018-04-03

    Acetic acid fermentation is widely considered a consequence of ethanol oxidation by two membrane-bound enzymes-alcohol dehydrogenase and aldehyde dehydrogenase (ALDH)-of acetic acid bacteria. Here, we used a markerless gene disruption method to construct a mutant of the Acetobacter pasteurianus strain SKU1108 with a deletion in the aldH gene, which encodes the large catalytic subunit of a heterotrimeric ALDH complex (AldFGH), to examine the role of AldFGH in acetic acid fermentation. The ΔaldH strain grew less on ethanol-containing medium, i.e., acetic acid fermentation conditions, than the wild-type strain and significantly accumulated acetaldehyde in the culture medium. Unexpectedly, acetaldehyde oxidase activity levels of the intact ΔaldH cells and the ΔaldH cell membranes were similar to those of the wild-type strain, which might be attributed to an additional ALDH isozyme (AldSLC). The apparent K M values of the wild-type and ΔaldH membranes for acetaldehyde were similar to each other, when the cells were cultured in nonfermentation conditions, where ΔaldH cells grow as well as the wild-type cells. However, the membranes of the wild-type cells grown under fermentation conditions showed a 10-fold lower apparent K M value than those of the cells grown under nonfermentation conditions. Under fermentation conditions, transcriptional levels of a gene for AldSLC were 10-fold lower than those under nonfermentation conditions, whereas aldH transcript levels were not dramatically changed under the two conditions. We suggest that A. pasteurianus SKU1108 has two ALDHs, and the AldFGH complex is indispensable for acetic acid fermentation and is the major enzyme under fermentation conditions.

  6. Structures of Proline Utilization A (PutA) Reveal the Fold and Functions of the Aldehyde Dehydrogenase Superfamily Domain of Unknown Function*

    Science.gov (United States)

    Luo, Min; Gamage, Thameesha T.; Arentson, Benjamin W.; Schlasner, Katherine N.; Becker, Donald F.; Tanner, John J.

    2016-01-01

    Aldehyde dehydrogenases (ALDHs) catalyze the NAD(P)+-dependent oxidation of aldehydes to carboxylic acids and are important for metabolism and detoxification. Although the ALDH superfamily fold is well established, some ALDHs contain an uncharacterized domain of unknown function (DUF) near the C terminus of the polypeptide chain. Herein, we report the first structure of a protein containing the ALDH superfamily DUF. Proline utilization A from Sinorhizobium meliloti (SmPutA) is a 1233-residue bifunctional enzyme that contains the DUF in addition to proline dehydrogenase and l-glutamate-γ-semialdehyde dehydrogenase catalytic modules. Structures of SmPutA with a proline analog bound to the proline dehydrogenase site and NAD+ bound to the ALDH site were determined in two space groups at 1.7–1.9 Å resolution. The DUF consists of a Rossmann dinucleotide-binding fold fused to a three-stranded β-flap. The Rossmann domain resembles the classic ALDH superfamily NAD+-binding domain, whereas the flap is strikingly similar to the ALDH superfamily dimerization domain. Paradoxically, neither structural element performs its implied function. Electron density maps show that NAD+ does not bind to the DUF Rossmann fold, and small-angle X-ray scattering reveals a novel dimer that has never been seen in the ALDH superfamily. The structure suggests that the DUF is an adapter domain that stabilizes the aldehyde substrate binding loop and seals the substrate-channeling tunnel via tertiary structural interactions that mimic the quaternary structural interactions found in non-DUF PutAs. Kinetic data for SmPutA indicate a substrate-channeling mechanism, in agreement with previous studies of other PutAs. PMID:27679491

  7. Structures of Proline Utilization A (PutA) Reveal the Fold and Functions of the Aldehyde Dehydrogenase Superfamily Domain of Unknown Function.

    Science.gov (United States)

    Luo, Min; Gamage, Thameesha T; Arentson, Benjamin W; Schlasner, Katherine N; Becker, Donald F; Tanner, John J

    2016-11-11

    Aldehyde dehydrogenases (ALDHs) catalyze the NAD(P) + -dependent oxidation of aldehydes to carboxylic acids and are important for metabolism and detoxification. Although the ALDH superfamily fold is well established, some ALDHs contain an uncharacterized domain of unknown function (DUF) near the C terminus of the polypeptide chain. Herein, we report the first structure of a protein containing the ALDH superfamily DUF. Proline utilization A from Sinorhizobium meliloti (SmPutA) is a 1233-residue bifunctional enzyme that contains the DUF in addition to proline dehydrogenase and l-glutamate-γ-semialdehyde dehydrogenase catalytic modules. Structures of SmPutA with a proline analog bound to the proline dehydrogenase site and NAD + bound to the ALDH site were determined in two space groups at 1.7-1.9 Å resolution. The DUF consists of a Rossmann dinucleotide-binding fold fused to a three-stranded β-flap. The Rossmann domain resembles the classic ALDH superfamily NAD + -binding domain, whereas the flap is strikingly similar to the ALDH superfamily dimerization domain. Paradoxically, neither structural element performs its implied function. Electron density maps show that NAD + does not bind to the DUF Rossmann fold, and small-angle X-ray scattering reveals a novel dimer that has never been seen in the ALDH superfamily. The structure suggests that the DUF is an adapter domain that stabilizes the aldehyde substrate binding loop and seals the substrate-channeling tunnel via tertiary structural interactions that mimic the quaternary structural interactions found in non-DUF PutAs. Kinetic data for SmPutA indicate a substrate-channeling mechanism, in agreement with previous studies of other PutAs. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. Characterization and Targeting of the Aldehyde Dehydrogenase Subpopulation in Ovarian Cancer

    Science.gov (United States)

    2012-07-01

    but increased expression specific to tumor cells in our laser-microdis- sected tissues suggest that it may play a role in tumor cell chemoresistance...Interests Family activities Wife Donna; Kids Sydney, Nicholson, and Jackson Sports / Crosstraining / Triathlon Hiking / Camping Religion / Philosophy / History

  9. Characterization and Targeting of the Aldehyde Dehydrogenase Subpopulation in Ovarian Cancer

    Science.gov (United States)

    2016-10-01

    Hanks’ balanced salt solution (HBSS; Gibco) and injected intraperitoneally into NOD- SCIDmice in limiting dilutions . Mice were followed for 1 year or...malignant human colonic stem cells (SC) and tracks SC overpopulation during colon tumorigenesis. Cancer Res 2009;69:3382–9. 16. Carpentino JE, HynesMJ...peroxidase activity was quenched with 3% hydrogen peroxide solution in methanol for 15 minutes. Sections were blocked with CytoQ immune diluent and

  10. The oxidative fermentation of ethanol in Gluconacetobacter diazotrophicus is a two-step pathway catalyzed by a single enzyme: alcohol-aldehyde Dehydrogenase (ADHa).

    Science.gov (United States)

    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.

  11. The Oxidative Fermentation of Ethanol in Gluconacetobacter diazotrophicus Is a Two-Step Pathway Catalyzed by a Single Enzyme: Alcohol-Aldehyde Dehydrogenase (ADHa

    Directory of Open Access Journals (Sweden)

    Saúl Gómez-Manzo

    2015-01-01

    Full Text Available 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.

  12. The Oxidative Fermentation of Ethanol in Gluconacetobacter diazotrophicus Is a Two-Step Pathway Catalyzed by a Single Enzyme: Alcohol-Aldehyde Dehydrogenase (ADHa)

    Science.gov (United States)

    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

  13. Identification and Overexpression of a Bifunctional Aldehyde/Alcohol Dehydrogenase Responsible for Ethanol Production in Thermoanaerobacter mathranii

    DEFF Research Database (Denmark)

    Yao, Shuo; Just Mikkelsen, Marie

    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 (Adh....... 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...

  14. Malignant phyllodes tumors display mesenchymal stem cell features and aldehyde dehydrogenase/disialoganglioside identify their tumor stem cells.

    Science.gov (United States)

    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

  15. Preferential antitumor effect of the Src inhibitor dasatinib associated with a decreased proportion of aldehyde dehydrogenase 1-positive cells in breast cancer cells of the basal B subtype

    Directory of Open Access Journals (Sweden)

    Watanabe Mika

    2010-10-01

    Full Text Available Abstract Background Recent studies have suggested that the Src inhibitor dasatinib preferentially inhibits the growth of breast cancer cells of the basal-like subtype. To clarify this finding and further investigate combined antitumor effects of dasatinib with cytotoxic agents, a panel of breast cancer cell lines of various subtypes was treated with dasatinib and/or chemotherapeutic agents. Methods Seven human breast cancer cell lines were treated with dasatinib and/or seven chemotherapeutic agents. Effects of the treatments on c-Src activation, cell growth, cell cycle, apoptosis and the proportion of aldehyde dehydrogenase (ALDH 1-positive cells were examined. Results The 50%-growth inhibitory concentrations (IC50s of dasatinib were much lower in two basal B cell lines than those in the other cell lines. The IC50s of chemotherapeutic agents were not substantially different among the cell lines. Dasatinib enhanced antitumor activity of etoposide in the basal B cell lines. Dasatinib induced a G1-S blockade with a slight apoptosis, and a combined treatment of dasatinib with etoposide also induced a G1-S blockade in the basal B cell lines. Dasatinib decreased the expression levels of phosphorylated Src in all cell lines. Interestingly, dasatinib significantly decreased the proportion of ALDH1-positive cells in the basal B cell lines but not in the other cell lines. Conclusions The present study indicates that dasatinib preferentially inhibits the growth of breast cancer cells of the basal B subtype associated with a significant loss of putative cancer stem cell population. A combined use of dasatinib with etoposide additively inhibits their growth. Further studies targeting breast cancers of the basal B subtype using dasatinib with cytotoxic agents are warranted.

  16. Pharmacological activities of cilantro's aliphatic aldehydes against Leishmania donovani.

    Science.gov (United States)

    Donega, Mateus A; Mello, Simone C; Moraes, Rita M; Jain, Surendra K; Tekwani, Babu L; Cantrell, Charles L

    2014-12-01

    Leishmaniasis is a chronic infectious disease caused by different Leishmania species. Global occurrences of this disease are primarily limited to tropical and subtropical regions. Treatments are available; however, patients complain of side effects. Different species of plants have been screened as a potential source of new drugs against leishmaniasis. In this study, we investigated the antileishmanial activity of cilantro (Coriandrum sativum) essential oil and its main components: (E)-2-undecenal, (E)-2-decenal, (E)-2-dodecenal, decanal, dodecanal, and tetradecanal. The essential oil of C. sativum leaves inhibits growth of Leishmani donovani promastigotes in culture with an IC50 of 26.58 ± 6.11 µg/mL. The aliphatic aldehydes (E)-2-decenal (7.85 ± 0.28 µg/mL), (E)-2-undecenal (2.81 ± 0.21 µg/mL), and (E)-2-dodecenal (4.35 ± 0.15 µg/mL), all isolated from C. sativum essential oil, are effective inhibitors of in vitro cultures of L. donovani promastigotes. Aldehydes (E)-2-decenal, (E)-2-undecenal, and (E)-2-dodecenal were also evaluated against axenic amastigotes and IC50 values were determined to be 2.47 ± 0.25 µg/mL, 1.25 ± 0.11 µg/mL, and 4.78 ± 1.12 µg/mL, respectively. (E)-2-Undecenal and (E)-2-dodecenal demonstrated IC50 values of 5.65 ± 0.19 µg/mL and 9.60 ± 0.89 µg/mL, respectively, against macrophage amastigotes. These cilantro compounds showed no cytotoxicity against THP-1 macrophages. Georg Thieme Verlag KG Stuttgart · New York.

  17. Genetic polymorphisms of alcohol and aldehyde dehydrogenases and glutathione S-transferase M1 and drinking, smoking, and diet in Japanese men with esophageal squamous cell carcinoma.

    Science.gov (United States)

    Yokoyama, Akira; Kato, Hoichi; Yokoyama, Tetsuji; Tsujinaka, Toshimasa; Muto, Manabu; Omori, Tai; Haneda, Tatsumasa; Kumagai, Yoshiya; Igaki, Hiroyasu; Yokoyama, Masako; Watanabe, Hiroshi; Fukuda, Haruhiko; Yoshimizu, Haruko

    2002-11-01

    The genetic polymorphisms of aldehyde dehydrogenase-2 (ALDH2), alcohol dehydrogenase-2 (ADH2), ADH3, and glutathione S-transferase M1 (GSTM1) influence the metabolism of alcohol and other carcinogens. The ALDH2*1/2*2 genotype, which encodes inactive ALDH2, and ADH2*1/2*1, which encodes the low-activity form of ADH2, enhance the risk for esophageal cancer in East Asian alcoholics. This case-control study of whether the enzyme-related vulnerability for esophageal cancer can be extended to a general population involved 234 Japanese men with esophageal squamous cell carcinoma and 634 cancer-free Japanese men who received annual health checkups. The GSTM1 genotype was not associated with the risk for this cancer. Light drinkers (1-8.9 units/week) with ALDH2*1/2*2 had an esophageal cancer risk 5.82 times that of light drinkers with ALDH2*1/2*1 (reference category), and their risk was similar to that of moderate drinkers (9-17.9 units/week) with ALDH2*1/2*1 (odds ratio = 5.58). The risk for moderate drinkers with ALDH2*1/2*2 (OR = 55.84) exceeded that for heavy drinkers (18+ units/week) with ALDH2*1/2*1 (OR = 10.38). Similar increased risks were observed for those with ADH2*1/2*1. A multiple logistic model including ALDH2, ADH2, and ADH3 genotypes showed that the ADH3 genotype does not significantly affect the risk for esophageal cancer. For individuals with both ALDH2*1/2*2 and ADH2*1/2*1, the risk of esophageal cancer was enhanced in a multiplicative fashion (OR = 30.12), whereas for those with either ALDH2*1/2*2 or ADH2*1/2*1 alone the ORs were 7.36 and 4.11. In comparison with the estimated population-attributable risks for preference for strong alcoholic beverages (30.7%), smoking (53.6%) and for lower intake of green and yellow vegetables (25.7%) and fruit (37.6%), an extraordinarily high proportion of the excessive risk for esophageal cancer in the Japanese males can be attributed to drinking (90.9%), particularly drinking by persons with inactive heterozygous ALDH

  18. Aldehyde dehydrogenase 1 is a marker for normal and malignant human colonic stem cells (SC) and tracks SC overpopulation during colon tumorigenesis.

    Science.gov (United States)

    Huang, Emina H; Hynes, Mark J; Zhang, Tao; Ginestier, Christophe; Dontu, Gabriela; Appelman, Henry; Fields, Jeremy Z; Wicha, Max S; Boman, Bruce M

    2009-04-15

    Although the concept that cancers originate from stem cells (SC) is becoming scientifically accepted, mechanisms by which SC contribute to tumor initiation and progression are largely unknown. For colorectal cancer (CRC), investigation of this problem has been hindered by a paucity of specific markers for identification and isolation of SC from normal and malignant colon. Accordingly, aldehyde dehydrogenase 1 (ALDH1) was investigated as a possible marker for identifying colonic SC and for tracking them during cancer progression. Immunostaining showed that ALDH1(+) cells are sparse and limited to the normal crypt bottom, where SCs reside. During progression from normal epithelium to mutant (APC) epithelium to adenoma, ALDH1(+) cells increased in number and became distributed farther up the crypt. CD133(+) and CD44(+) cells, which are more numerous and broadly distributed in normal crypts, showed similar changes during tumorigenesis. Flow cytometric isolation of cancer cells based on enzymatic activity of ALDH (Aldefluor assay) and implantation of these cells in nonobese diabetic-severe combined immunodeficient mice (a) generated xenograft tumors (Aldefluor(-) cells did not), (b) generated them after implanting as few as 25 cells, and (c) generated them dose dependently. Further isolation of cancer cells using a second marker (CD44(+) or CD133(+) serially) only modestly increased enrichment based on tumor-initiating ability. Thus, ALDH1 seems to be a specific marker for identifying, isolating, and tracking human colonic SC during CRC development. These findings also support our original hypothesis, derived previously from mathematical modeling of crypt dynamics, that progressive colonic SC overpopulation occurs during colon tumorigenesis and drives CRC development.

  19. Aldehyde Dehydrogenase 1 Is a Marker for Normal and Malignant Human Colonic Stem Cells (SC) and Tracks SC Overpopulation during Colon Tumorigenesis

    Science.gov (United States)

    Huang, Emina H.; Hynes, Mark J.; Zhang, Tao; Ginestier, Christophe; Dontu, Gabriela; Appelman, Henry; Fields, Jeremy Z.; Wicha, Max S.; Boman, Bruce M.

    2009-01-01

    Although the concept that cancers originate from stem cells (SC) is becoming scientifically accepted, mechanisms by which SC contribute to tumor initiation and progression are largely unknown. For colorectal cancer (CRC), investigation of this problem has been hindered by a paucity of specific markers for identification and isolation of SC from normal and malignant colon. Accordingly, aldehyde dehydrogenase 1 (ALDH1) was investigated as a possible marker for identifying colonic SC and for tracking them during cancer progression. Immunostaining showed that ALDH1+ cells are sparse and limited to the normal crypt bottom, where SCs reside. During progression from normal epithelium to mutant (APC) epithelium to adenoma, ALDH1+ cells increased in number and became distributed farther up the crypt. CD133+ and CD44+ cells, which are more numerous and broadly distributed in normal crypts, showed similar changes during tumorigenesis. Flow cytometric isolation of cancer cells based on enzymatic activity of ALDH (Aldefluor assay) and implantation of these cells in nonobese diabetic–severe combined immunodeficient mice (a) generated xenograft tumors (Aldefluor− cells did not), (b) generated them after implanting as few as 25 cells, and (c) generated them dose dependently. Further isolation of cancer cells using a second marker (CD44+ or CD133+ serially) only modestly increased enrichment based on tumor-initiating ability. Thus, ALDH1 seems to be a specific marker for identifying, isolating, and tracking human colonic SC during CRC development. These findings also support our original hypothesis, derived previously from mathematical modeling of crypt dynamics, that progressive colonic SC overpopulation occurs during colon tumorigenesis and drives CRC development. PMID:19336570

  20. Study on the triphenyl tetrazolium chloride– dehydrogenase activity ...

    African Journals Online (AJOL)

    A quick analysis of the sludge activity method based on triphenyltetrazolium chloride-dehydrogenase activity (TTC-DHA) was developed to change the rule and status of the biological activity of the activated sludge in tomato paste wastewater treatment. The results indicate that dehydrogenase activity (DHA) can effectively ...

  1. The activity of the artemisinic aldehyde Δ11(13) reductase promoter is important for artemisinin yield in different chemotypes of Artemisia annua L.

    Science.gov (United States)

    Yang, Ke; Monfared, Sajad Rashidi; Monafared, Rashidi Sajad; Wang, Hongzhen; Lundgren, Anneli; Brodelius, Peter E

    2015-07-01

    The artemisinic aldehyde double bond reductase (DBR2) plays an important role in the biosynthesis of the antimalarial artemisinin in Artemisia annua. Artemisinic aldehyde is reduced into dihydroartemisinic aldehyde by DBR2. Artemisinic aldehyde can also be oxidized by amorpha-4,11-diene 12-hydroxylase and/or aldehyde dehydrogenase 1 to artemisinic acid, a precursor of arteannuin B. In order to better understand the effects of DBR2 expression on the flow of artemisinic aldehyde into either artemisinin or arteannuin B, we determined the content of dihydroartemisinic aldehyde, artemisinin, artemisinic acid and arteannuin B content of A. annua varieties sorted into two chemotypes. The high artemisinin producers (HAPs), which includes the '2/39', 'Chongqing' and 'Anamed' varieties, produce more artemisinin than arteannuin B; the low artemisinin producers (LAPs), which include the 'Meise', 'Iran#8', 'Iran#14', 'Iran#24' and 'Iran#47' varieties, produce more arteannuin B than artemisinin. Quantitative PCR showed that the relative expression of DBR2 was significantly higher in the HAP varieties. We cloned and sequenced the promoter of the DBR2 gene from varieties of both the LAP and the HAP groups. There were deletions/insertions in the region just upstream of the ATG start codon in the LAP varities, which might be the reason for the different promoter activities of the HAP and LAP varieties. The relevance of promoter variation, DBR2 expression levels and artemisinin biosynthesis capabilities are discussed and a selection method for HAP varieties with a DNA marker is suggested. Furthermore, putative cis-acting regulatory elements differ between the HAP and LAP varieties.

  2. Purification, crystallization and preliminary X-ray analysis of recombinant betaine aldehyde dehydrogenase 2 (OsBADH2), a protein involved in jasmine aroma, from Thai fragrant rice (Oryza sativa L.)

    International Nuclear Information System (INIS)

    Kuaprasert, Buabarn; Silprasit, Kun; Horata, Natharinee; Khunrae, Pongsak; Wongpanya, Ratree; Boonyalai, Nonlawat; Vanavichit, Apichart; Choowongkomon, Kiattawee

    2011-01-01

    Crystals of betaine aldehyde dehydrogenase 2 from rice (O. sativa L.) belonged to a C-centred orthorhombic space group and diffraceted X-rays to 2.6 Å resolution. Fragrant rice (Oryza sativa L.) betaine aldehyde dehydrogenase 2 (OsBADH2) is a key enzyme in the synthesis of fragrance aroma compounds. The extremely low activity of OsBADH2 in catalyzing the oxidation of acetaldehyde is believed to be crucial for the accumulation of the volatile compound 2-acetyl-1-pyrroline (2AP) in many scented plants, including fragrant rice. Recombinant fragrant rice OsBADH2 was expressed in Escherichia coli as an N-terminal hexahistidine fusion protein, purified using Ni Sepharose affinity chromatography and crystallized using the microbatch method. Initial crystals were obtained within 24 h using 0.1 M Tris pH 8.5 with 30%(w/v) PEG 4000 and 0.2 M magnesium chloride as the precipitating agent at 291 K. Crystal quality was improved when the enzyme was cocrystallized with NAD + . Improved crystals were grown in 0.1 M HEPES pH 7.4, 24%(w/v) PEG 4000 and 0.2 M ammonium chloride and diffracted to beyond 2.95 Å resolution after being cooled in a stream of N 2 immediately prior to X-ray diffraction experiments. The crystals belonged to space group C222 1 , with unit-cell parameters a = 66.03, b = 183.94, c = 172.28 Å. An initial molecular-replacement solution has been obtained and refinement is in progress

  3. Aldehyde Dehydrogenase 7A1 (ALDH7A1) Is a Novel Enzyme Involved in Cellular Defense against Hyperosmotic Stress*

    OpenAIRE

    Brocker, Chad; Lassen, Natalie; Estey, Tia; Pappa, Aglaia; Cantore, Miriam; Orlova, Valeria V.; Chavakis, Triantafyllos; Kavanagh, Kathryn L.; Oppermann, Udo; Vasiliou, Vasilis

    2010-01-01

    Mammalian ALDH7A1 is homologous to plant ALDH7B1, an enzyme that protects against various forms of stress, such as salinity, dehydration, and osmotic stress. It is known that mutations in the human ALDH7A1 gene cause pyridoxine-dependent and folic acid-responsive seizures. Herein, we show for the first time that human ALDH7A1 protects against hyperosmotic stress by generating osmolytes and metabolizing toxic aldehydes. Human ALDH7A1 expression in Chinese hamster ovary cells attenuated osmotic...

  4. Aromatic aldehydes at the active site of aldehyde oxidoreductase from Desulfovibrio gigas: reactivity and molecular details of the enzyme-substrate and enzyme-product interaction.

    Science.gov (United States)

    Correia, Hugo D; Marangon, Jacopo; Brondino, Carlos D; Moura, Jose J G; Romão, Maria J; González, Pablo J; Santos-Silva, Teresa

    2015-03-01

    Desulfovibrio gigas aldehyde oxidoreductase (DgAOR) is a mononuclear molybdenum-containing enzyme from the xanthine oxidase (XO) family, a group of enzymes capable of catalyzing the oxidative hydroxylation of aldehydes and heterocyclic compounds. The kinetic studies reported in this work showed that DgAOR catalyzes the oxidative hydroxylation of aromatic aldehydes, but not heterocyclic compounds. NMR spectroscopy studies using (13)C-labeled benzaldehyde confirmed that DgAOR catalyzes the conversion of aldehydes to the respective carboxylic acids. Steady-state kinetics in solution showed that high concentrations of the aromatic aldehydes produce substrate inhibition and in the case of 3-phenyl propionaldehyde a suicide substrate behavior. Hydroxyl-substituted aromatic aldehydes present none of these behaviors but the kinetic parameters are largely affected by the position of the OH group. High-resolution crystallographic structures obtained from single crystals of active-DgAOR soaked with benzaldehyde showed that the side chains of Phe425 and Tyr535 are important for the stabilization of the substrate in the active site. On the other hand, the X-ray data of DgAOR soaked with trans-cinnamaldehyde showed a cinnamic acid molecule in the substrate channel. The X-ray data of DgAOR soaked with 3-phenyl propionaldehyde showed clearly how high substrate concentrations inactivate the enzyme by binding covalently at the surface of the enzyme and blocking the substrate channel. The different reactivity of DgAOR versus aldehyde oxidase and XO towards aromatic aldehydes and N-heterocyclic compounds is explained on the basis of the present kinetic and structural data.

  5. Genetic polymorphisms of alcohol dehydrogense-1B and aldehyde dehydrogenase-2, alcohol flushing, mean corpuscular volume, and aerodigestive tract neoplasia in Japanese drinkers.

    Science.gov (United States)

    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.

  6. Mitochondria-targeted ubiquinone (MitoQ enhances acetaldehyde clearance by reversing alcohol-induced posttranslational modification of aldehyde dehydrogenase 2: A molecular mechanism of protection against alcoholic liver disease

    Directory of Open Access Journals (Sweden)

    Liuyi Hao

    2018-04-01

    Full Text Available Alcohol metabolism in the liver generates highly toxic acetaldehyde. Breakdown of acetaldehyde by aldehyde dehydrogenase 2 (ALDH2 in the mitochondria consumes NAD+ and generates reactive oxygen/nitrogen species, which represents a fundamental mechanism in the pathogenesis of alcoholic liver disease (ALD. A mitochondria-targeted lipophilic ubiquinone (MitoQ has been shown to confer greater protection against oxidative damage in the mitochondria compared to untargeted antioxidants. The present study aimed to investigate if MitoQ could preserve mitochondrial ALDH2 activity and speed up acetaldehyde clearance, thereby protects against ALD. Male C57BL/6 J mice were exposed to alcohol for 8 weeks with MitoQ supplementation (5 mg/kg/d for the last 4 weeks. MitoQ ameliorated alcohol-induced oxidative/nitrosative stress and glutathione deficiency. It also reversed alcohol-reduced hepatic ALDH activity and accelerated acetaldehyde clearance through modulating ALDH2 cysteine S-nitrosylation, tyrosine nitration and 4-hydroxynonenol adducts formation. MitoQ ameliorated nitric oxide (NO donor-mediated ADLH2 S-nitrosylation and nitration in Hepa-1c1c7 cells under glutathion depletion condition. In addition, alcohol-increased circulating acetaldehyde levels were accompanied by reduced intestinal ALDH activity and impaired intestinal barrier. In accordance, MitoQ reversed alcohol-increased plasma endotoxin levels and hepatic toll-like receptor 4 (TLR4-NF-κB signaling along with subsequent inhibition of inflammatory cell infiltration. MitoQ also reversed alcohol-induced hepatic lipid accumulation through enhancing fatty acid β-oxidation. Alcohol-induced ER stress and apoptotic cell death signaling were reversed by MitoQ. This study demonstrated that speeding up acetaldehyde clearance by preserving ALDH2 activity critically mediates the beneficial effect of MitoQ on alcohol-induced pathogenesis at the gut-liver axis. Keywords: Aldehyde dehydrogenase 2

  7. The aldehyde dehydrogenase, AldA, is essential for L-1,2-propanediol utilization in laboratory-evolved Escherichia coli.

    Science.gov (United States)

    Aziz, Ramy K; Monk, Jonathan M; Andrews, Kathleen A; Nhan, Jenny; Khaw, Valerie L; Wong, Hesper; Palsson, Bernhard O; Charusanti, Pep

    2017-01-01

    Most Escherichia coli strains are naturally unable to grow on 1,2-propanediol (PDO) as a sole carbon source. Recently, however, a K-12 descendent E. coli strain was evolved to grow on 1,2-PDO, and it was hypothesized that this evolved ability was dependent on the aldehyde dehydrogenase, AldA, which is highly conserved among members of the family Enterobacteriacea. To test this hypothesis, we first performed computational model simulation, which confirmed the essentiality of the aldA gene for 1,2-PDO utilization by the evolved PDO-degrading E. coli. Next, we deleted the aldA gene from the evolved strain, and this deletion was sufficient to abolish the evolved phenotype. On re-introducing the gene on a plasmid, the evolved phenotype was restored. These findings provide experimental evidence for the computationally predicted role of AldA in 1,2-PDO utilization, and represent a good example of E. coli robustness, demonstrated by the bacterial deployment of a generalist enzyme (here AldA) in multiple pathways to survive carbon starvation and to grow on a non-native substrate when no native carbon source is available. Copyright © 2016 Elsevier GmbH. All rights reserved.

  8. A polymorphism of the aldehyde dehydrogenase 2 gene is a risk factor for multiple lacunar infarcts in Japanese men: the Takahata Study.

    Science.gov (United States)

    Nagasawa, H; Wada, M; Arawaka, S; Kawanami, T; Kurita, K; Daimon, M; Adachi, M; Hosoya, T; Emi, M; Muramatsu, M; Kato, T

    2007-04-01

    The objective of the present study was to examine the association between a polymorphism of the aldehyde dehydrogenase 2 (ALDH2) gene and lacunar infarcts of the brain. We conducted a population-based, cross-sectional study on residents from two age groups (61- and 72-year olds). A total of 376 subjects participated in the study, which included brain magnetic resonance image and genetic analysis of the ALDH2 gene. Of the 61- and 72-year-old subjects, 46.4% and 64.3%, respectively, had one or more lacunar infarcts. The average number of infarcts also increased from 2.0 to 2.8 in men and from 2.3 to 3.5 in women. No significant association between the ALDH2 genotype and the presence of lacunar infarction (> or =1) was found. However, in subjects with lacunar infarction, the genotype of ALDH2 *1/*1 was associated with a larger number of the lesion ['single' versus 'multiple' odds ratio (OR) 3.73, 95%CI: 1.43-9.74] in men. The OR was comparable even after adjusting for alcohol consumption, tobacco habits, age, hypertension, hypercholesterolemia, and diabetes mellitus (DM) (OR 3.88; 95% CI: 1.10-13.66). In women, there was no significant association between the ALDH2 genotypes and lacunar infarcts. The present study revealed that the ALDH2 *1/*1 genotype was significantly associated with the prevalence of multiple lacunar infarcts in Japanese men.

  9. Catalytic activity of bovine glutamate dehydrogenase requires a hexamer structure.

    OpenAIRE

    Bell, E T; Bell, J E

    1984-01-01

    Previous workers have shown that the hexamers of glutamate dehydrogenase are dissociated first into trimers and subsequently into monomers by increasing guanidinium chloride concentrations. In renaturation experiments it is shown that trimers of glutamate dehydrogenase can be reassociated to give the hexamer form of the enzyme, with full regain of activity. Monomeric subunits produced at high guanidinium chloride concentrations cannot be renatured. The trimer form of the enzyme is shown to ha...

  10. Lactate dehydrogenase activity drives hair follicle stem cell activation

    Science.gov (United States)

    Aimee, Flores; John, Schell; Abby, Krall; David, Jelinek; Matilde, Miranda; Melina, Grigorian; Daniel, Braas; White Andrew, C; Jessica, Zhou; Nick, Graham; Thomas, Graeber; Pankaj, Seth; Denis, Evseenko; Hilary, Coller; Jared, Rutter; Heather, Christofk; Lowry William, E

    2017-01-01

    Summary While normally dormant, Hair Follicle Stem Cells (HFSCs) quickly become activated to divide during a new hair cycle. The quiescence of HFSCs is known to be regulated by a number of intrinsic and extrinsic mechanisms. Here we provide several lines of evidence to demonstrate that HFSCs utilize glycolytic metabolism and produce significantly more lactate than other cells in the epidermis. Furthermore, lactate generation appears to be critical for the activation of HFSCs as deletion of lactate dehydrogenase (Ldha) prevented their activation. Conversely, genetically promoting lactate production in HFSCs through mitochondrial pyruvate carrier (Mpc1) deletion accelerated their activation and the hair cycle. Finally, we identify small molecules that increase lactate production by stimulating Myc levels or inhibiting Mpc1 carrier activity and can topically induce the hair cycle. These data suggest that HFSCs maintain a metabolic state that allow them to remain dormant and yet quickly respond to appropriate proliferative stimuli. PMID:28812580

  11. Dehydrogenase activity of forest soils depends on the assay used

    Science.gov (United States)

    Januszek, Kazimierz; Długa, Joanna; Socha, Jarosław

    2015-01-01

    Dehydrogenases are exclusively intracellular enzymes, which play an important role in the initial stages of oxidation of soil organic matter. One of the most frequently used methods to estimate dehydrogenase activity in soil is based on the use of triphenyltetrazolium chloride as an artificial electron acceptor. The purpose of this study was to compare the activity of dehydrogenases of forest soils with varied physicochemical properties using different triphenyltetrazolium chloride assays. The determination was carried out using the original procedure by Casida et al., a modification of the procedure which involves the use of Ca(OH)2 instead of CaCO3, the Thalmann method, and the assay by Casida et al. without addition of buffer or any salt. Soil dehydrogenase activity depended on the assay used. Dehydrogenase determined by the Casida et al. method without addition of buffer or any salt correlated with the pH values of soils. The autoclaved strongly acidic samples of control soils showed high concentrations of triphenylformazan, probably due to chemical reduction of triphenyltetrazolium chloride. There is, therefore, a need for a sterilization method other than autoclaving, ie a process that results in significant changes in soil properties, thus helping to increase the chemical reduction of triphenyltetrazolium chloride.

  12. Association of an aldehyde dehydrogenase 2 (ALDH2) gene polymorphism with hyper-low-density lipoprotein cholesterolemia in a Japanese population.

    Science.gov (United States)

    Kotani, Kazuhiko; Sakane, Naoki; Yamada, Toshiyuki

    2012-01-01

    The relationship among alcohol metabolism, lipid profile and cardiovascular disease has been a matter of concern, and aldehyde dehydrogenase-2 (ALDH2) is one of the key enzymes involved in alcohol metabolism. The frequency of ALDH2 gene G/A polymorphism (with the substitution of glutamic acid to lysine) varies widely among ethnic groups; the polymorphism is prevalent among Asian people but rare in other ethnic groups. The objective of our study was to investigate the association between the ALDH2 gene G/A polymorphism and lipid profile, including the low-density lipoprotein cholesterol (LDL-C) status, in a general Japanese population with no or light-to-moderate alcohol drinking habits. Anthropometric and biochemical variables including lipid- and glucose-related factors were measured in a total of 383 Japanese participants (170 males and 213 females; mean age, 45 +/- 8.6 years), free of cardiovascular disease. All participants were genotyped by an allele-specific DNA assay. The numbers of participants with the G/ G, G/A and A/A genotypes were 213, 139 and 31, respectively. The percentages of hyper-LDL-cholesterolemia (identified by LDL-C > or = 3.63 mmol/L) were 31.9%, 45.3% and 29.0% in participants with the G/G, G/A and A/A genotypes, respectively. Carrying the G/A + AA genotype was a significant and positive factor related to hyper-LDL-cholesterolemia with an odds ratio of 1.62 (95% CI: 1.04-2.52) after adjusting for the other variables including drinking status. Our findings suggest that the ALDH2 gene G/A polymorphism can affect the lipid profile such as LDL-C status in this population. The association between the polymorphism and LDL-C status warrants further investigation.

  13. In vitro oxidative metabolism of 6-mercaptopurine in human liver: insights into the role of the molybdoflavoenzymes aldehyde oxidase, xanthine oxidase, and xanthine dehydrogenase.

    Science.gov (United States)

    Choughule, Kanika V; Barnaba, Carlo; Joswig-Jones, Carolyn A; Jones, Jeffrey P

    2014-08-01

    Anticancer agent 6-mercaptopurine (6MP) has been in use since 1953 for the treatment of childhood acute lymphoblastic leukemia (ALL) and inflammatory bowel disease. Despite being available for 60 years, several aspects of 6MP drug metabolism and pharmacokinetics in humans are unknown. Molybdoflavoenzymes such as aldehyde oxidase (AO) and xanthine oxidase (XO) have previously been implicated in the metabolism of this drug. In this study, we investigated the in vitro metabolism of 6MP to 6-thiouric acid (6TUA) in pooled human liver cytosol. We discovered that 6MP is metabolized to 6TUA through sequential metabolism via the 6-thioxanthine (6TX) intermediate. The role of human AO and XO in the metabolism of 6MP was established using the specific inhibitors raloxifene and febuxostat. Both AO and XO were involved in the metabolism of the 6TX intermediate, whereas only XO was responsible for the conversion of 6TX to 6TUA. These findings were further confirmed using purified human AO and Escherichia coli lysate containing expressed recombinant human XO. Xanthine dehydrogenase (XDH), which belongs to the family of xanthine oxidoreductases and preferentially reduces nicotinamide adenine dinucleotide (NAD(+)), was shown to contribute to the overall production of the 6TX intermediate as well as the final product 6TUA in the presence of NAD(+) in human liver cytosol. In conclusion, we present evidence that three enzymes, AO, XO, and XDH, contribute to the production of 6TX intermediate, whereas only XO and XDH are involved in the conversion of 6TX to 6TUA in pooled HLC. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

  14. A Novel Approach for Overcoming Drug Resistance in Breast Cancer Chemotherapy by Targeting new Synthetic Curcumin Analogues Against Aldehyde Dehydrogenase 1 (ALDH1A1) and Glycogen Synthase Kinase-3 β (GSK-3β).

    Science.gov (United States)

    Kesharwani, Rajesh Kumar; Srivastava, Vandana; Singh, Prabhakar; Rizvi, Syed Ibrahim; Adeppa, Kuruba; Misra, Krishna

    2015-08-01

    Breast cancer stem cells are well known to resist the traditional methods like chemo and radio therapy. Aldehyde dehydrogenase 1 (ALDHIA1) and glycogen synthase kinase-3 β (GSK-3β) are the two selected proteins for study, due to their overexpression and upregulation in breast cancer cells. Curcumin, the yellow pigment of the spice turmeric, is widely reported as an antioxidant and acts as a synergist along with traditional drugs. Under hypoxic conditions, it gets converted to free radical which causes apoptosis. Three naturally occurring curcuminoids, i.e. curcumin, demethoxycurcumin, and bisdemethoxycurcumin along with five derivatives/analogues of curcumin, viz. 4,4'-di-O-(carboxy-methyl)-curcumin, 4-O-(2-hydroxyethyl)curcumin, 4,4'-di-O-allyl-curcumin, 4,4'-di-O-(acetyl)-curcumin, and 3,3'-bisdemethylcurcumin were synthesized and evaluated for their anti-breast cancer potential by docking simulation and assessment of their antioxidant character, studied via 2, 2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS(·+)) radical cation scavenging assay, 2,2-diphenyl-1-picrylhydrazyl (DPPH·) radical, and ferric reducing ability potential (FRAP) assay. A co-relation between structure and activity of curcuminoids/its analogues and derivatives has been worked out.

  15. Aminotransferase and glutamate dehydrogenase activities in lactobacilli and streptococci

    Directory of Open Access Journals (Sweden)

    Guillermo Hugo Peralta

    Full Text Available ABSTRACT Aminotransferases and glutamate dehydrogenase are two main types of enzymes involved in the initial steps of amino acid catabolism, which plays a key role in the cheese flavor development. In the present work, glutamate dehydrogenase and aminotransferase activities were screened in twenty one strains of lactic acid bacteria of dairy interest, either cheese-isolated or commercial starters, including fifteen mesophilic lactobacilli, four thermophilic lactobacilli, and two streptococci. The strains of Streptococcus thermophilus showed the highest glutamate dehydrogenase activity, which was significantly elevated compared with the lactobacilli. Aspartate aminotransferase prevailed in most strains tested, while the levels and specificity of other aminotransferases were highly strain- and species-dependent. The knowledge of enzymatic profiles of these starter and cheese-isolated cultures is helpful in proposing appropriate combinations of strains for improved or increased cheese flavor.

  16. Serum creatine kinase and lactate dehydrogenase activities in ...

    African Journals Online (AJOL)

    P = 0.002). Conclusion: The significant elevation in serum CK and LDH activities indicates that these can be used as parameters for screening hypothyroid patients but not hyperthyroid patients. Key words: Hyperthyroidism, hypothyroidism, lactate dehydrogenase, serum creatine kinase. Date of Acceptance: 28-Aug-2011.

  17. A new cotton SDR family gene encodes a polypeptide possessing aldehyde reductase and 3-ketoacyl-CoA reductase activities.

    Science.gov (United States)

    Pang, Yu; Song, Wen-Qiang; Chen, Fang-Yuan; Qin, Yong-Mei

    2010-03-01

    To understand regulatory mechanisms of cotton fiber development, microarray analysis has been performed for upland cotton (Gossypium hirsutum). Based on this, a cDNA (GhKCR3) encoding a polypeptide belonging to short-chain alcohol dehydrogenase/reductase family was isolated and cloned. It contains an open reading frame of 987 bp encoding a polypeptide of 328 amino acid residues. Following its overexpression in bacterial cells, the purified recombinant protein specifically uses NADPH to reduce a variety of short-chain aldehydes. A fragment between Gly180 and Gly191 was found to be essential for its catalytic activity. Though the GhKCR3 gene shares low sequence similarities to the ortholog of Saccharomyces cerevisiae YBR159w that encodes 3-ketoacyl-CoA reductase (KCR) catalyzing the second step of fatty acid elongation, it was surprisingly able to complement the yeast ybr159wDelta mutant. Gas chromatography-mass spectrometry analysis showed that very long-chain fatty acids, especially C26:0, were produced in the ybr159wDelta mutant cells expressing GhKCR3. Applying palmitoyl-CoA and malonyl-CoA as substrates, GhKCR3 showed KCR activity in vitro. Quantitative real time-PCR analysis indicated GhKCR3 transcripts accumulated in rapidly elongating fibers, roots, and stems. Our results suggest that GhKCR3 is probably a novel KCR contributing to very long-chain fatty acid biosynthesis in plants.

  18. Expression of aldehyde dehydrogenase family 1, member A3 in glycogen trophoblast cells of the murine placenta.

    Science.gov (United States)

    Outhwaite, J E; Natale, B V; Natale, D R C; Simmons, D G

    2015-03-01

    Retinoic acid (RA) signaling is a well known regulator of trophoblast differentiation and placental development, and maternal decidual cells are recognized as the source of much of this RA. We explored possible trophoblast-derived sources of RA by examining the expression of RA synthesis enzymes in the developing mouse placenta, as well as addressed potential sites of RA action by examining the ontogeny of gene expression for other RA metabolizing and receptor genes. Furthermore, we investigated the effects of endogenous RA production on trophoblast differentiation. Placental tissues were examined by in situ hybridization and assayed for RARE-LacZ transgene activity to locate sites of RAR signaling. Trophoblast stem cell cultures were differentiated in the presence of ALDH1 inhibitors (DEAB and citral), and expression of labyrinth (Syna, Ctsq) and junctional zone (Tpbpa, Prl7b1, Prl7a2) marker genes were analyzed by qRT-PCR. We show Aldh1a3 is strongly expressed in a subset of ectoplacental cone cells and in glycogen trophoblast cells of the definitive murine placenta. Most trophoblast subtypes of the placenta express RA receptor combinations that would enable them to respond to RA signaling. Furthermore, expression of junctional zone markers decrease in differentiating trophoblast cultures when endogenous ALDH1 enzymes are inhibited. Aldh1a3 is a novel marker for glycogen trophoblast cells and their precursors and may play a role in the differentiation of junctional zone cell types via production of a local source of RA. Copyright © 2014 Elsevier Ltd. All rights reserved.

  19. Antibacterial activity of Mannich bases derived from 2-naphthols, aromatic aldehydes and secondary aliphatic amines.

    Science.gov (United States)

    Roman, Gheorghe; Năstasă, Valentin; Bostănaru, Andra-Cristina; Mareş, Mihai

    2016-05-15

    A small library of 1-aminoalkyl 2-naphthols has been synthesized through the direct Mannich reaction of 2-naphthols with (hetero)aromatic aldehydes and secondary amines. All of the Mannich bases having a thiophen-2-yl ring in their structure had good activity against Gram-positive bacteria, irrespective of the nature of the amino moiety. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. [Studies On Lactic Dehydrogenase Activity In Parasitic Helminths

    Science.gov (United States)

    Lee, Soon Hyung

    1967-06-01

    A series of experiments was performed to determine the lactic dehydrogenase activity of various parasitic helminths. The enzyme activity was determined by the modified method of Wroblewshi and LaDue (1955) using tissue homogenate of 16 kinds of worm parasites. The worms were mostly collected alive from local abattoir and removed from the organ or tissues of the naturally infected animal host and some materials were also obtained from the human host. They were thoroughly washed and homogenized in chilled glass tissue grinder, and then centrifuged. The supernatants were designated as enzyme preparations, and their enzyme activity was measured by spectrophotometry at the wave length of 340 millimicron. In order to know the effects of temperature and substrate concentration on the enzyme activity, the extinction of reduced Coenzyme I(NADH) was measured at the various conditions of incubation temperature and substrate concentration. The results of this experiments were as follows: 1. The lactic dehydrogenase activity occurred over all kinds of parasites used in this study. 2. Most worms of nematodes and trematodes displayed their maximum activity in the range of pH 2.7~3.5, and cestodes revealed their maximum activity in the ranges of both pH 2.7~3.5 and pH 7.4. 3. In nematodes and trematodes, the lactic dehydrogenase activity increased slowly as incubation temperature increases except in the case of Eurytrema pancreaticum, while the activity in cestodes decreased inversely. 4. The lactic dehydrogenase activity increased in proportion to the increase of substrate concentration in most of worm parasites.

  1. High ethanol and acetaldehyde impair spatial memory in mouse models: opposite effects of aldehyde dehydrogenase 2 and apolipoprotein E on memory.

    Science.gov (United States)

    Jamal, Mostofa; Ameno, Kiyoshi; Miki, Takanori; Tanaka, Naoko; Ono, Junichiro; Shirakami, Gotaro; Sultana, Ruby; Yu, Nakamura; Kinoshita, Hiroshi

    2012-05-01

    Aldehyde dehydrogenase 2 deficiency may directly contribute to excess acetaldehyde (AcH) accumulation after ethanol (EtOH) drinking and AcH mediates some of the behavioral effects of EtOH. Apolipoprotein E has been suggested to be involved in the alteration of attention and memory. We have chosen Aldh2-knockout (Aldh2-KO), ApoE-KO, and their wild-type (WT) control mice to examine the effects of EtOH and AcH on spatial memory and to compare the possible relationship between genetic deficiency and memory using two behavioral assessments. Mice were trained for 4 days, with EtOH (0.5, 1.0, 2.0 g/kg) being given intraperitoneally on day 4. A probe trial was given on day 5 in the non-EtOH state in the Morris water maze (MWM). The results showed that 2.0 g/kg EtOH increased errors, indicating memory impairment on the eight-arm radial maze (RAM) for all the mice studied. One gram per kilogram EtOH impaired the performance of Aldh2-KO and ApoE-KO mice, but not WT mice. We found similar effects of EtOH on the MWM performance, with 2.0 g/kg EtOH increasing the latencies. One gram per kilogram EtOH increased the latencies of Aldh2-KO and WT mice, but not ApoE-KO mice. The 2.0 g/kg EtOH-induced memory impairment in Aldh2-KO mice was greater, suggesting an AcH effect. Furthermore, time spent on the probe trial was shorter in mice that had previously received 2.0 g/kg EtOH. ApoE-KO mice learned more slowly, while Aldh2-KO mice learned more quickly. Both the RAM and MWM results suggest that high EtOH and AcH impair spatial memory in mice, while lower doses do not have consistent memory effects. In addition, we conclude that genetic differences might underlie some of EtOH's effects on memory. Copyright © 2012 Elsevier Inc. All rights reserved.

  2. Anisaldehyde production and aryl-alcohol oxidase and dehydrogenase activities in ligninolytic fungi of the genus Pleurotus.

    Science.gov (United States)

    Gutiérrez, A; Caramelo, L; Prieto, A; Martínez, M J; Martínez, A T

    1994-01-01

    A variety of simple aromatic compounds were identified in liquid cultures of the basidiomycetes Pleurotus cornucopiae, P. eryngii, P. floridanus, P. pulmonarius, P. ostreatus, and P. sajor-caju by using gas chromatography-mass spectrometry. Such compounds were detected in fungal cultures on lignin- and straw-containing media, but it was found that they were also produced in the absence of aromatic precursors. Anisylic and hydroxybenzylic compounds (such as alcohols, aldehydes, and acids) were identified, p-anisaldehyde being the most characteristic extracellular metabolite synthesized by these ligninolytic fungi. Small amounts of 3-chloro-p-anisaldehyde were also detected in several species. It is postulated that the balance between the more-or-less-oxidized aromatic compounds can be explained in terms of the activity of fungal enzymes, including aryl-alcohol oxidase and dehydrogenase. The former enzyme shows high affinity for p-anisyl alcohol, which is oxidized to p-anisaldehyde with production of H2O2. The aryl-alcohol dehydrogenase was detected only in the mycelium, where it reduces aromatic aldehydes in the presence of NADPH. Both enzymes could be involved in the redox cycling of these aromatic compounds, providing H2O2 to ligninolytic peroxidases. PMID:8031078

  3. Deficiency in the amino aldehyde dehydrogenase encoded by GmAMADH2, the homologue of rice Os2AP, enhances 2-acetyl-1-pyrroline biosynthesis in soybeans (Glycine max L.).

    Science.gov (United States)

    Arikit, Siwaret; Yoshihashi, Tadashi; Wanchana, Samart; Uyen, Tran T; Huong, Nguyen T T; Wongpornchai, Sugunya; Vanavichit, Apichart

    2011-01-01

    2-Acetyl-1-pyrroline (2AP), the volatile compound that provides the 'popcorn-like' aroma in a large variety of cereal and food products, is widely found in nature. Deficiency in amino aldehyde dehydrogenase (AMADH) was previously shown to be the likely cause of 2AP biosynthesis in rice (Oryza sativa L.). In this study, the validity of this mechanism was investigated in soybeans (Glycine max L.). An assay of AMADH activity in soybeans revealed that the aromatic soybean, which contains 2AP, also lacked AMADH enzyme activity. Two genes, GmAMADH1 and GmAMADH2, which are homologous to the rice Os2AP gene that encodes AMADH, were characterized. The transcription level of GmAMADH2 was lower in aromatic varieties than in nonaromatic varieties, whereas the expression of GmAMADH1 did not differ. A double nucleotide (TT) deletion was found in exon 10 of GmAMADH2 in all aromatic varieties. This variation caused a frame-shift mutation and a premature stop codon. Suppression of GmAMADH2 by introduction of a GmAMADH2-RNAi construct into the calli of the two nonaromatic wild-type varieties inhibited the synthesis of AMADH and induced the biosynthesis of 2AP. These results suggest that deficiency in the GmAMADH2 product, AMADH, plays a similar role in soybean as in rice, which is to promote 2AP biosynthesis. This phenomenon might be a conserved mechanism among plant species. © 2010 The Authors. Plant Biotechnology Journal © 2010 Society for Experimental Biology and Blackwell Publishing Ltd.

  4. Influence of thorax irradiation on lactic dehydrogenase isoenzyme activity

    International Nuclear Information System (INIS)

    Valle, C.; Munnich, A.; Pasquier, C.

    The right hemi-thorax of rats was irradiated with 1200 and 3000 rads ( 60 Co) and blood samples were taken sequentially. The five lactic dehydrogenase (LDH) isoenzymes which have proved to be useful as biochemical indicators of acute pulmonary injury in other experimental animals (dogs), were assayed, after irradiation, as a function of time and as a functon of dose. There was no significant change in LDH isoenzyme activities after lung irradiation in rats [fr

  5. Investigation of antimicrobial activities of indole-3-aldehyde hydrazide/hydrazone derivatives.

    Science.gov (United States)

    Gurkok, Gokce; Altanlar, Nurten; Suzen, Sibel

    2009-01-01

    Indoles and hydrazone-type compounds constitute an important class of compounds for new drug development in order to discover an effective compound against multi-drug-resistant microbial infections. A series of indole-3-aldehyde and 5-bromoindole-3-aldehyde hydrazide and hydrazones was evaluated for their in vitro antimicrobial activities using the 2-fold serial dilution technique against Staphylococcus aureus, methicillin-resistant S. aureus (MRSA), Escherichia coli, Bacillus subtilis and Candida albicans. The minimum inhibitory concentration (MIC) was determined for test compounds and for the reference standards sultamicillin, ampicillin, fluconazole and ciprofloxacin. Compounds possessed a broad spectrum of activity having MIC values of 6.25-100 mg/ml against the tested microorganisms. Compounds 1a-1j, in particular, displayed better activity against MSRA and significant activity against S. aureus relative to ampicillin. Unexpectedly, indole nicotinic acid hydrazides showed no significant activity while indole anisic acid hydrazides displayed better activity. The results may be instructive to researchers attempting to gain more understanding of the antimicrobial activity of indole hydrazide/hydrazone-type compounds. (c) 2008 S. Karger AG, Basel.

  6. Antisickling activity evaluation of 4 aromatic aldehydes using proton magnetic relaxation

    International Nuclear Information System (INIS)

    Falcon Dieguez, J.E.; Grisel del Toro Garcia; Yamirka Alonso Geli; Lores Guevara, M.A.

    2006-12-01

    The formation of a Shiff base aduct hemoglobin-aromatic aldehyde, has been reported as inhibitor of the hemoglobin S polymerization. Using the Proton Magnetic Resonance methodology, the polymerization kinetics can be studied and the delay time can be determined. Our studies in vitro show the inhibitor effect of the isovanillin, o-vanillin, m-hydroxybenzaldehyde and the p-hydroxybenzaldehyde, using molar ratio (hemoglobin S/compound) 1:1, 1:4 and 1:8. The td increment (expressed in percents) obtained for each one of the molar ratio was the following: isovanillin: 34±6% (1:1), 68±16% (1:4), ovanillin: 26±10% (1:1), 63±20% (1:4), m-hydroxybelzaldehyde: 16±4% (1:1), 44±12% (1:4) and the phydroxybenzaldehyde: 10±3% (1:1), 32±8% (1:4). In the case of 1:8, the characteristic kinetics curve was not obtained. At the used concentrations, hemolytic activity was not found on the red blood cell. These results confirm the antisickling activity of these aromatic aldehydes, for a technique different to that reported in literature that also allows the quantification of concentration effect. The same ones will facilitate the study of the therapeutic usefulness of these compounds in the sickle cell anemia treatment. (author)

  7. Oxydoreductases activation by γ rayonnement : lipoxygenase and alcohol dehydrogenase

    International Nuclear Information System (INIS)

    Mejri, Sonia

    2004-01-01

    Ionising radiations, which have already been used for increasing the conservation length of food products, amelioration physicochemical properties of some products of some products and enhancing the rate of some reactions. In fact, we have selected soybean lipoxygenase and yest alcohol dehydrogenase in order to check the comportment of those oxydoreductases with the 60Co-gamma rays. Results reveal a significant effect on the increase of the enzymes activity, stability and productivity. His effect was so much more important when it is anhydrous enzyme to the aqueous one. Enhanced activity and productivity of the enzymes were not affected by the same dose level of irradiation. The irradiating dose 12 KGy active the soybean lipoxygenase as aqueous solution and 8 KGy active this last one under his form anhydrous. Nevertheless, alcohol deshydrogenase is activated by the doses of the interval 30-50Gy. (author). 99 refs

  8. [Activity of blood serum lactate dehydrogenase in diabetes mellitus].

    Science.gov (United States)

    Vizir, O O

    1977-01-01

    The activity of lactic dehydrogenase of the blood serum was studied under clinical conditions in 120 patients suffering from diabetes mellitus. Electrophoretic separation of plasma enzymes was used for this purpose. The shifts in the LDH activity proved to be characteristic of all the degrees of diabetes severity, and were expressed in a significant elevation of total LDH, LDH4,5 activity and a decrease of LDH1,2 activity in comparison with healthy individuals. No change of LDH3 activity was noted in mild forms of diabetes. But in severe form the isoenzymatic spectrum was mostly changed on account of LDH3 hyperfermentemia. In mild form of diabetes it approached control values.

  9. Orthodontic Force Application in Correlation with Salivary Lactate Dehydrogenase Activity

    Directory of Open Access Journals (Sweden)

    Erik Husin

    2013-07-01

    Full Text Available Orthodontic tooth movement generate mechanical forces to periodontal ligament and alveolar bone. The forces correlate with initial responses of periodontal tissues and involving many metabolic changes. One of the metabolic changes detected in saliva is lactate dehydrogenase (LDH activity. Objectives: To evaluate the correlation between orthodontic interrupted force application, lactate dehydrogenase activity and the distance of tooth movement. Methods: upper premolar, pre-retraction of upper canine and 1, 7, 14, 21 and 28 days post-retraction of upper canine with 100g interrupted orthodontic force. Results: duration of force (F=11.926 p 14 and 28 days post-retraction of canine. The region of retraction correlated with the distance of tooth movement (F=7.377 p=0.007. The duration of force correlated with the distance of tooth movement (F=66.554 p=0.000. retraction of canine. Conclusion: This study concluded that orthodontic interrupted force application on canine could increase the distance of tooth movement and LDH activity in saliva.

  10. Identification of a Novel Activator of Mammalian Glutamate Dehydrogenase.

    Science.gov (United States)

    Smith, Hong Q; Smith, Thomas J

    2016-11-29

    Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of l-glutamate and in animals is highly regulated. GDH in hyperinsulinism/hyperammonemia syndrome patients lacks GTP inhibition, resulting in hypersecretion of insulin upon protein consumption. This suggests insulin secretion could be stimulated with GDH activators. A high-throughput screen yielded one potent activator, N1-[4-(2-aminopyrimidin-4-yl)phenyl]-3-(trifluoromethyl)benzene-1-sulfonamide (75-E10). 75-E10 is ∼1000-fold more efficacious than the synthetic activator, BCH, and is at least as effective as ADP. 75-E10 compound is highly effective at alleviating GTP inhibition and may be binding to the ADP site. Unlike ADP, 75-E10 is activated over a broad range of conditions.

  11. Identification of lactaldehyde dehydrogenase and glycolaldehyde dehydrogenase as functions of the same protein in Escherichia coli.

    Science.gov (United States)

    Caballero, E; Baldomá, L; Ros, J; Boronat, A; Aguilar, J

    1983-06-25

    Lactaldehyde dehydrogenase is an enzyme involved in the aerobic metabolism of fucose in wild type Escherichia coli, and glycolaldehyde dehydrogenase is an enzyme involved in the metabolism of ethylene glycol in mutant cells able to utilize this glycol. Both enzyme sources display oxidative activity on either substrate with a constant ratio between these activities. We have found that both enzymatic activities present the same electrophoretic mobility when crude extracts were electrophoresed in polyacrylamide gels and the gels stained for enzyme activities. Furthermore, both enzymatic activities co-chromatograph in a DEAE-Sephadex column. If lactaldehyde dehydrogenase of wild type cells is purified near homogeneity and the purification procedure is screened for both aldehydes as substrates, only one enzyme is apparent, giving again a constant ratio between lactaldehyde and glycolaldehyde dehydrogenase activities. Genetic evidence of the fact that both activities are functions of the same protein is provided by the observation that mutation to thermosensitivity for the production of lactaldehyde dehydrogenase affected in the same way the production of glycolaldehyde dehydrogenase. Glycolaldehyde dehydrogenase from mutant cells is purified in a procedure coincident with the lactaldehyde dehydrogenase purification, yielding a single enzyme electrophoretically indistinguishable from the purified lactaldehyde dehydrogenase. Peptide mapping of the purified preparation after digestion with chymotrypsin or Staphylococcus aureus protease V8 gives an indistinguishable band pattern between both enzymes.

  12. Circadian rhythm in succinate dehydrogenase activity in Neurospora crassa

    Directory of Open Access Journals (Sweden)

    Claudia Patricia Álvarez Barón

    2004-07-01

    Full Text Available Neurospora crassa is a widely studied model of circadian rhythmicity. In this fungus, metabolism is controlled by multiple factors which include development, medium characteristics and the circadian clock. The study of the circadian control of metabolism in this fungus could be masked by the use of restrictive media that inhibit growth and development. In this report, the presence of a circadian rhythm in the activity of the enzyme Succinate Dehydrogenase in Neurospora crassa is demonstrated. Rhythmic and arrhythmic Neurospora strains were grown in complete medium without conidiation restriction. A circadian change in the enzymatic activity was found with high values in hours corresponding to the night and a low level during the day. This finding highlights the importance of deeper studies in the circadian control of metabolism in this fungus, given the existence of multiple pathways of regulation of metabolic enzymes and a circadian clock control at the transcriptional and post-transcriptional levels.

  13. Lanthanide(III) ion - luminescent and catalytically active center of aniline condensation with butyric aldehyde

    International Nuclear Information System (INIS)

    Bulgakov, R.G.; Kuleshov, S.P.; Makhmutov, A.R.

    2007-01-01

    New type of chemiluminescent-catalytic transformation, where lanthanide(III) ion performs as luminescent and high effective catalytic active center, is observed. The chemiluminescent (CL) is generated in the reaction of aniline condensation with butyric aldehyde in DMFA with the formation of 2-propyl-3-ethyl quinoline that is catalyzed by LnCl 3 ·6H 2 O (Ln=Eu, Tb and Ho). Excited ions Eu* 3+ and Tb* 3+ are served as emitters of CL when using salts EuCl 3 ·6H 2 O and TbCl 3 ·6H 2 O by way of catalysts, and in the case of HoCl 3 ·6H 2 O triplet-excited state of 2-propyl-3-ethyl quinoline ( 3 C 14 H 17 N*) is an emitter of CL [ru

  14. 17 beta-hydroxysteroid dehydrogenase activity in canine pancreas

    International Nuclear Information System (INIS)

    Mendoza-Hernandez, G.; Lopez-Solache, I.; Rendon, J.L.; Diaz-Sanchez, V.; Diaz-Zagoya, J.C.

    1988-01-01

    The mitochondrial fraction of the dog pancreas showed NAD(H)-dependent enzyme activity of 17 beta-hydroxysteroid dehydrogenase. The enzyme catalyzes oxidoreduction between androstenedione and testosterone. The apparent Km value of the enzyme for androstenedione was 9.5 +/- 0.9 microM, the apparent Vmax was determined as 0.4 nmol mg-1 min-1, and the optimal pH was 6.5. In phosphate buffer, pH 7.0, maximal rate of androstenedione reduction was observed at 37 degrees C. The oxidation of testosterone by the enzyme proceeded at the same rate as the reduction of the androstenedione at a pH of 6.8-7.0. The apparent Km value and the optimal pH of the enzyme for testosterone were 3.5 +/- 0.5 microM and 7.5, respectively

  15. 17 beta-hydroxysteroid dehydrogenase activity in canine pancreas

    Energy Technology Data Exchange (ETDEWEB)

    Mendoza-Hernandez, G.; Lopez-Solache, I.; Rendon, J.L.; Diaz-Sanchez, V.; Diaz-Zagoya, J.C.

    1988-04-15

    The mitochondrial fraction of the dog pancreas showed NAD(H)-dependent enzyme activity of 17 beta-hydroxysteroid dehydrogenase. The enzyme catalyzes oxidoreduction between androstenedione and testosterone. The apparent Km value of the enzyme for androstenedione was 9.5 +/- 0.9 microM, the apparent Vmax was determined as 0.4 nmol mg-1 min-1, and the optimal pH was 6.5. In phosphate buffer, pH 7.0, maximal rate of androstenedione reduction was observed at 37 degrees C. The oxidation of testosterone by the enzyme proceeded at the same rate as the reduction of the androstenedione at a pH of 6.8-7.0. The apparent Km value and the optimal pH of the enzyme for testosterone were 3.5 +/- 0.5 microM and 7.5, respectively.

  16. [Effect Of Polyelectrolytes on Catalytic Activity of Alcohol Dehydrogenase].

    Science.gov (United States)

    Dubrovsky, A V; Musina, E V; Kim, A L; Tikhonenko, S A

    2016-01-01

    Fluorescent and optical spectroscopy were used to study the interaction of alcohol dehydrogenase (ADH) with negatively charged polystyrene sulfonate (PSS) and dextran sulfate (DS), as well as positively charged poly(diallyldimethylammonium) (PDADMA). As found, DS and PDADMA did not affect the structural and catalytic enzyme properties. In contrast, PSS slightly decreased the protein self-fluorescence over 1 h of incubation, which is associated with partial destruction of its quaternary (globular) structure. Investigation of the ADH activity with and without PSS showed its dependency on the incubation time and the PSS presence. Sodium chloride (2.0 M and 0.2 M) or ammonium sulfate (0.1 M) added to the reaction mixture did not completely protect the enzyme quaternary structure from the PSS action. However ammonium sulfate or 0.2 M sodium chloride stabilized the enzyme and partially inhibited the negative PSS effect.

  17. A Case of Hyperammonemia Associated with High Dihydropyrimidine Dehydrogenase Activity

    Directory of Open Access Journals (Sweden)

    Keiki Nagaharu

    2016-01-01

    Full Text Available Over the past decades, 5-Fluorouracil (5-FU has been widely used to treat several types of carcinoma, including esophageal squamous cell carcinoma. In addition to its common side effects, including diarrhea, mucositis, neutropenia, and anemia, 5-FU treatment has also been reported to cause hyperammonemia. However, the exact mechanism responsible for 5-FU-induced hyperammonemia remains unknown. We encountered an esophageal carcinoma patient who developed hyperammonemia when receiving 5-FU-containing chemotherapy but did not exhibit any of the other common adverse effects of 5-FU treatment. At the onset of hyperammonemia, laboratory tests revealed high dihydropyrimidine dehydrogenase (DPD activity and rapid 5-FU clearance. Our findings suggested that 5-FU hypermetabolism may be one of the key mechanisms responsible for hyperammonemia during 5-FU treatment.

  18. Evaluation of Serum Lactate Dehydrogenase Activity in a Virtual Environment

    Directory of Open Access Journals (Sweden)

    V.M.T. Trindade

    2013-05-01

    Full Text Available Introduction: Lactate dehydrogenase is a citosolic enzyme involved in reversible transformation of pyruvate to lactate. It participates in anaerobic glycolysis of skeletal muscle and red blood cells, in liver gluconeogenesis and in aerobic metabolism of heart muscle. The determination of its activity helps in the diagnosis of various diseases, because it is increased in serum of patients suffering from myocardial infarction, acute hepatitis, muscular dystrophy and cancer. This paper presents a learning object, mediated by computer, which contains the simulation of the laboratory determination serum lactate dehydrogenase activity measured by the spectrophotometric method, based in the decrease of absorbance at 340 nm. Materials and Methods: Initially, pictures and videos were obtained recording the procedure of the methodology. The most representative images were selected, edited and inserted into an animation developed with the aid of the tool Adobe ® Flash ® CS3. The validation of the object was performed by the students of Biochemistry I (Pharmacy-UFRGS from the second semester of 2009 and both of 2010. Results and Discussion: The analysis of students' answers revealed that 80% attributed the excellence of the navigation program, the display format and to aid in learning. Conclusion: Therefore, this software can be considered an adequate teaching resource as well as an innovative support in the construction of theoretical and practical knowledge of Biochemistry. Available at: http://www6.ufrgs.br/gcoeb/LDH

  19. Brønsted acid-catalyzed Mannich reaction through dual activation of aldehydes and N-Boc-imines.

    Science.gov (United States)

    Kano, Taichi; Aota, Yusuke; Asakawa, Daisuke; Maruoka, Keiji

    2015-11-28

    In the presence of a Brønsted acid catalyst, both aldehydes and N-Boc-aminals were converted to enecarbamates and N-Boc-iminium salts as activated nucleophiles and electrophiles, respectively, giving unprecedented Mannich adducts. The asymmetric variant of the present Mannich reaction has also been demonstrated with a chiral phosphoric acid catalyst.

  20. The D-Lactate Dehydrogenase from Sporolactobacillus inulinus Also Possessing Reversible Deamination Activity

    OpenAIRE

    Zhu, Lingfeng; Xu, Xiaoling; Wang, Limin; Dong, Hui; Yu, Bo

    2015-01-01

    Hydroxyacid dehydrogenases are responsible for the conversion of 2-keto acids to 2-hydroxyacids and have a wide range of biotechnological applications. In this study, a D-lactate dehydrogenase (D-LDH) from a Sporolactobacillus inulinus strain was experimentally verified to have both the D-LDH and glutamate dehydrogenase (GDH) activities (reversible deamination). The catalytic mechanism was demonstrated by identification of key residues from the crystal structure analysis and site-directed mut...

  1. Variants of glycerol dehydrogenase having D-lactate dehydrogenase activity and uses thereof

    Science.gov (United States)

    Wang, Qingzhao; Shanmugam, Keelnatham T.; Ingram, Lonnie O'Neal

    2017-08-29

    The present invention provides methods of designing and generating glycerol dehydrogenase (GlyDH) variants that have altered function as compared to a parent polypeptide. The present invention further provides nucleic acids encoding GlyDH polypeptide variants having altered function as compared to the parent polypeptide. Host cells comprising polynucleotides encoding GlyDH variants and methods of producing lactic acids are also provided in various aspects of the invention.

  2. The D-Lactate Dehydrogenase from Sporolactobacillus inulinus Also Possessing Reversible Deamination Activity.

    Directory of Open Access Journals (Sweden)

    Lingfeng Zhu

    Full Text Available Hydroxyacid dehydrogenases are responsible for the conversion of 2-keto acids to 2-hydroxyacids and have a wide range of biotechnological applications. In this study, a D-lactate dehydrogenase (D-LDH from a Sporolactobacillus inulinus strain was experimentally verified to have both the D-LDH and glutamate dehydrogenase (GDH activities (reversible deamination. The catalytic mechanism was demonstrated by identification of key residues from the crystal structure analysis and site-directed mutagenesis. The Arg234 and Gly79 residues of this enzyme play a significant role in both D-LDH and GDH activities. His295 and Phe298 in DLDH744 were identified to be key residues for lactate dehydrogenase (LDH activity only whereas Tyr101 is a unique residue that is critical for GDH activity. Characterization of the biochemical properties contributes to understanding of the catalytic mechanism of this novel D-lactate dehydrogenase enzyme.

  3. Alcohol Dehydrogenase Activities of Wine Yeasts in Relation to Higher Alcohol Formation

    Science.gov (United States)

    Singh, Rajendra; Kunkee, Ralph E.

    1976-01-01

    Alcohol dehydrogenase activities were examined in cell-free extracts of 10 representative wine yeast strains having various productivities of higher alcohols (fusel oil). The amount of fusel alcohols (n-propanol, isobutanol, active pentanol, and isopentanol) produced by the different yeasts and the specific alcohol dehydrogenase activities with the corresponding alcohols as substrates were found to be significantly related. No such relationship was found for ethanol. The amounts of higher alcohols formed during vinification could be predicted from the specific activities of the alcohol dehydrogenases with high accuracy. The results suggest a close relationship between the control of the activities of alcohol dehydrogenase and the formation of fusel oil alcohols. Also, new procedures for the prediction of higher alcohol formation during alcoholic beverage fermentation are suggested. PMID:16345179

  4. Site directed mutagenesis of amino acid residues at the active site of mouse aldehyde oxidase AOX1.

    Directory of Open Access Journals (Sweden)

    Silvia Schumann

    Full Text Available Mouse aldehyde oxidase (mAOX1 forms a homodimer and belongs to the xanthine oxidase family of molybdoenzymes which are characterized by an essential equatorial sulfur ligand coordinated to the molybdenum atom. In general, mammalian AOs are characterized by broad substrate specificity and an yet obscure physiological function. To define the physiological substrates and the enzymatic characteristics of mAOX1, we established a system for the heterologous expression of the enzyme in Escherichia coli. The recombinant protein showed spectral features and a range of substrate specificity similar to the native protein purified from mouse liver. The EPR data of recombinant mAOX1 were similar to those of AO from rabbit liver, but differed from the homologous xanthine oxidoreductase enzymes. Site-directed mutagenesis of amino acids Val806, Met884 and Glu1265 at the active site resulted in a drastic decrease in the oxidation of aldehydes with no increase in the oxidation of purine substrates. The double mutant V806E/M884R and the single mutant E1265Q were catalytically inactive enzymes regardless of the aldehyde or purine substrates tested. Our results show that only Glu1265 is essential for the catalytic activity by initiating the base-catalyzed mechanism of substrate oxidation. In addition, it is concluded that the substrate specificity of molybdo-flavoenzymes is more complex and not only defined by the three characterized amino acids in the active site.

  5. Studies in Wild Oat Seed Dormancy: II. ACTIVITIES OF PENTOSE PHOSPHATE PATHWAY DEHYDROGENASES.

    Science.gov (United States)

    Adkins, S W; Ross, J D

    1981-07-01

    A selected strain of wild oat (Avena fatua L.) seed has been shown to lose dormancy rapidly during moist soil incubation at 25 C, whereas seed kept similarly at 5 C maintained a high level of dormancy.The activities of cytosolic dehydrogenase enzymes of the pentose phosphate pathway were assayed throughout a period of moist soil incubation at these two temperatures. A distinction was made between extractable dehydrogenases from the embryo and the endosperm regions of the caryopsis.Dehydrogenase activities monitored in seeds incubated at 25 C gradually increased over the course of the investigation. The largest increases in activity occurred during incubation at 5 C, the situation in which dormancy is maintained. No obvious connection could be found between dormancy breakage and increased activity of the pentose phosphate pathway dehydrogenases.

  6. In vitro effects of metals and pesticides on dehydrogenase activity in ...

    African Journals Online (AJOL)

    In vitro effects of metals and pesticides on dehydrogenase activity in microbial community of cowpea (Vigna unguiculata) rhizoplane. CO Nweke, C Ntinugwa, IF Obah, SC Ike, GE Eme, EC Opara, JC Okolo, CE Nwanyanwu ...

  7. Assessment of the activity of glucose-6-phosphate dehydrogenase ...

    African Journals Online (AJOL)

    Glucose-6-phosphate dehydrogenase (G-6-PD) is an enzyme in the pentose phosphate pathway (PPP) which reduces NADP to NADPH while oxidizing glucose-6-phosphate. In turn, NADPH then provides reducing equivalents needed for the conversion of oxidized glutathione to reduced glutathione, which protects against ...

  8. Assessment of creatine kinase and lactate dehydrogenase activities ...

    African Journals Online (AJOL)

    Ina bid to investigate the influence of menopausal on coronary heart disease, plasma creatine kinase (CK) and lactate dehydrogenase (LDH) enzymes were analysed on a prospective cohort of 100 women attending Irrua Specialist Teaching Hospital (ISTH), Irrua, Edo state-Nigeria. They were divided into two groups; ...

  9. Inosine monophosphate dehydrogenase messenger RNA expression is correlated to clinical outcomes in mycophenolate mofetil-treated kidney transplant patients, whereas inosine monophosphate dehydrogenase activity is not

    NARCIS (Netherlands)

    Sombogaard, Ferdi; Peeters, Annemiek M. A.; Baan, Carla C.; Mathot, Ron A. A.; Quaedackers, Monique E.; Vulto, Arnold G.; Weimar, Willem; van Gelder, Teun

    2009-01-01

    Measurement of the pharmacodynamic biomarker inosine monophosphate dehydrogenase (IMPDH) activity in renal transplant recipients has been proposed to reflect the biological effect better than using pharmacokinetic parameters to monitor mycophenolate mofetil therapy. The IMPDH assays are however

  10. Construction of mutant glucose oxidases with increased dye-mediated dehydrogenase activity.

    Science.gov (United States)

    Horaguchi, Yohei; Saito, Shoko; Kojima, Katsuhiro; Tsugawa, Wakako; Ferri, Stefano; Sode, Koji

    2012-11-02

    Mutagenesis studies on glucose oxidases (GOxs) were conducted to construct GOxs with reduced oxidase activity and increased dehydrogenase activity. We focused on two representative GOxs, of which crystal structures have already been reported—Penicillium amagasakiense GOx (PDB ID; 1gpe) and Aspergillus niger GOx (PDB ID; 1cf3). We constructed oxygen-interacting structural models for GOxs, and predicted the residues responsible for oxidative half reaction with oxygen on the basis of the crystal structure of cholesterol oxidase as well as on the fact that both enzymes are members of the glucose/methanol/choline (GMC) oxidoreductase family. Rational amino acid substitution resulted in the construction of an engineered GOx with drastically decreased oxidase activity and increased dehydrogenase activity, which was higher than that of the wild-type enzyme. As a result, the dehydrogenase/oxidase ratio of the engineered enzyme was more than 11-fold greater than that of the wild-type enzyme. These results indicate that alteration of the dehydrogenase/oxidase activity ratio of GOxs is possible by introducing a mutation into the putative functional residues responsible for oxidative half reaction with oxygen of these enzymes, resulting in a further increased dehydrogenase activity. This is the first study reporting the alteration of GOx electron acceptor preference from oxygen to an artificial electron acceptor.

  11. Construction of Mutant Glucose Oxidases with Increased Dye-Mediated Dehydrogenase Activity

    Directory of Open Access Journals (Sweden)

    Koji Sode

    2012-11-01

    Full Text Available Mutagenesis studies on glucose oxidases (GOxs were conducted to construct GOxs with reduced oxidase activity and increased dehydrogenase activity. We focused on two representative GOxs, of which crystal structures have already been reported—Penicillium amagasakiense GOx (PDB ID; 1gpe and Aspergillus niger GOx (PDB ID; 1cf3. We constructed oxygen-interacting structural models for GOxs, and predicted the residues responsible for oxidative half reaction with oxygen on the basis of the crystal structure of cholesterol oxidase as well as on the fact that both enzymes are members of the glucose/methanol/choline (GMC oxidoreductase family. Rational amino acid substitution resulted in the construction of an engineered GOx with drastically decreased oxidase activity and increased dehydrogenase activity, which was higher than that of the wild-type enzyme. As a result, the dehydrogenase/oxidase ratio of the engineered enzyme was more than 11-fold greater than that of the wild-type enzyme. These results indicate that alteration of the dehydrogenase/oxidase activity ratio of GOxs is possible by introducing a mutation into the putative functional residues responsible for oxidative half reaction with oxygen of these enzymes, resulting in a further increased dehydrogenase activity. This is the first study reporting the alteration of GOx electron acceptor preference from oxygen to an artificial electron acceptor.

  12. Dimerization and enzymatic activity of fungal 17β-hydroxysteroid dehydrogenase from the short-chain dehydrogenase/reductase superfamily

    Directory of Open Access Journals (Sweden)

    Kristan Katja

    2005-12-01

    Full Text Available Abstract Background 17β-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17β-HSDcl is a member of the short-chain dehydrogenase/reductase (SDR superfamily. SDR proteins usually function as dimers or tetramers and 17β-HSDcl is also a homodimer under native conditions. Results We have investigated here which secondary structure elements are involved in the dimerization of 17β-HSDcl and examined the importance of dimerization for the enzyme activity. Sequence similarity with trihydroxynaphthalene reductase from Magnaporthe grisea indicated that Arg129 and His111 from the αE-helices interact with the Asp121, Glu117 and Asp187 residues from the αE and αF-helices of the neighbouring subunit. The Arg129Asp and His111Leu mutations both rendered 17β-HSDcl monomeric, while the mutant 17β-HSDcl-His111Ala was dimeric. Circular dichroism spectroscopy analysis confirmed the conservation of the secondary structure in both monomers. The three mutant proteins all bound coenzyme, as shown by fluorescence quenching in the presence of NADP+, but both monomers showed no enzymatic activity. Conclusion We have shown by site-directed mutagenesis and structure/function analysis that 17β-HSDcl dimerization involves the αE and αF helices of both subunits. Neighbouring subunits are connected through hydrophobic interactions, H-bonds and salt bridges involving amino acid residues His111 and Arg129. Since the substitutions of these two amino acid residues lead to inactive monomers with conserved secondary structure, we suggest dimerization is a prerequisite for catalysis. A detailed understanding of this dimerization could lead to the development of compounds that will specifically prevent dimerization, thereby serving as a new type of inhibitor.

  13. Soil dehydrogenase activity of natural macro aggregates in a toposequence of forest soil

    Directory of Open Access Journals (Sweden)

    Maira Kussainova

    2013-01-01

    Full Text Available The main objective of this study was to determine changes in soil dehydrogenase activity in natural macro aggregates development along a slope in forest soils. This study was carried out in Kocadag, Samsun, Turkey. Four landscape positions i.e., summit, shoulder backslope and footslope, were selected. For each landseape position, soil macro aggregates were separated into six aggregate size classes using a dry sieving method and then dehydrogenase activity was analyzed. In this research, topography influenced the macroaggregate size and dehydrogenase activity within the aggregates. At all landscape positions, the contents of macro aggregates (especially > 6.3 mm and 2.00–4.75 mm in all soil samples were higher than other macro aggregate contents. In footslope position, the soils had generally the higher dehydrogenase activity than the other positions at all landscape positions. In all positions, except for shoulder, dehydrogenase activity was greater macro aggregates of <1 mm than in the other macro aggregate size.

  14. Cytosolic malate dehydrogenase activity helps support glycolysis in actively proliferating cells and cancer.

    Science.gov (United States)

    Hanse, E A; Ruan, C; Kachman, M; Wang, D; Lowman, X H; Kelekar, A

    2017-07-06

    Increased glucose consumption is a hallmark of cancer cells. The increased consumption and subsequent metabolism of glucose during proliferation creates the need for a constant supply of NAD, a co-factor in glycolysis. Regeneration of the NAD required to support enhanced glycolysis has been attributed to the terminal glycolytic enzyme, lactate dehydrogenase (LDH). However, loss of glucose carbons to biosynthetic pathways early in glycolysis reduces the carbon supply to LDH. Thus, alternative routes for NAD regeneration must exist to support the increased glycolytic rate while allowing for the diversion of glucose to generate biomass and support proliferation. Here we demonstrate, using a variety of cancer cell lines as well as activated primary T cells, that cytosolic malate dehydrogenase 1 (MDH1) is an alternative to LDH as a supplier of NAD. Moreover, our results indicate that MDH1 generates malate with carbons derived from glutamine, thus enabling utilization of glucose carbons for glycolysis and for biomass. Amplification of MDH1 occurs at an impressive frequency in human tumors and correlates with poor prognosis. Together, our findings suggest that proliferating cells rely on both MDH1 and LDH to replenish cytosolic NAD, and that therapies designed at targeting glycolysis must consider both dehydrogenases.

  15. Glucose-6-phosphate dehydrogenase activity decreases during storage of leukoreduced red blood cells

    NARCIS (Netherlands)

    Peters, Anna L.; van Bruggen, Robin; de Korte, Dirk; van Noorden, Cornelis J. F.; Vlaar, Alexander P. J.

    2016-01-01

    During storage, the activity of the red blood cell (RBC) antioxidant system decreases. Glucose-6-phosphate dehydrogenase (G6PD) is essential for protection against oxidative stress by producing NADPH. G6PD function of RBC transfusion products is reported to remain stable during storage, but activity

  16. Magnetic-Resonance Studies of the Geometry of Bound Substrate, Coenzyme and Activator on Bovine-Liver Glutamate Dehydrogenase

    NARCIS (Netherlands)

    Zantema, Alt; de Smet, Marie-José; Robillard, George T.

    ADP and ATP with a spin-label linked to the terminal phosphate are activators of glutamate dehydrogenase and bind to the same site as the activator ADP. There is hardly any interaction with the coenzyme site. Glutamate dehydrogenase can be modified with a ketone spin-label at a site in the active

  17. Aldehyde dehydrogenase 2 deficiency increases resting-state glutamate and expression of the GluN1 subunit of N-methyl-D-aspartate receptor in the frontal cortex of mice.

    Science.gov (United States)

    Jamal, Mostofa; Ono, Junichiro; Ameno, Kiyoshi; Shirakami, Gotaro; Tanaka, Naoko; Takakura, Ayaka; Kinoshita, Hiroshi

    2015-01-15

    Our previous study showed that Aldh2-knockout (Aldh2-KO) mice, an animal model of inactive aldehyde dehydrogenase 2 (ALDH2), have better spatial memory when compared with wild-type (WT) mice. Given that the neurotransmitter glutamate has been associated with learning and memory, the goal of the present study was to investigate whether the strain-dependent difference in spatial memory was associated with changes in glutamate transmitter levels or receptor function in the frontal cortex of Aldh2-KO and WT mice. Thus, we first measured extracellular glutamate levels in free-moving mice using microdialysis. Second, we studied protein expression of the N-methyl-D-aspartate (NMDA) receptor (GluN1) subunit and the α-amino-3-hydroxy-5 methylisoxazole-4-propionic acid (AMPA) receptor (GluA1) subunit in lipid raft fractions using Western blot (WB). The samples were collected for WB, and lipid rafts were prepared from the insoluble fraction of homogenate tissue. Protein concentration was measured in the whole cell lysate (WCL) and in five separate lipid raft fractions. Cholesterol was also measured in all fractions 1-5. The microdialysis study revealed that basal glutamate concentration in the dialysates was approximately three-fold (0.27 ± 0.12 μM) higher in Aldh2-KO mice than in WT (0.10 ± 0.03 μM) mice. We also found an increase in the expression of GluN1 in Aldh2-KO mice compared with WT mice, both in the WCL and fraction 5, but GluA1 levels were unchanged as measured by WB. Our novel findings provide the first evidence for the role of ALDH2 in glutamate release and GluN1 protein expression in the frontal cortex. The observed strain differences in glutamate levels and GluN1 expression may suggest that enhanced glutamatergic function facilitates improved spatial memory in Aldh2-KO mice and such observation deserves further investigation. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Potentiation of insulin release in response to amino acid methyl esters correlates to activation of islet glutamate dehydrogenase activity

    DEFF Research Database (Denmark)

    Kofod, Hans; Lernmark, A; Hedeskov, C J

    1986-01-01

    Column perifusion of mouse pancreatic islets was used to study the ability of amino acids and their methyl esters to influence insulin release and activate islet glutamate dehydrogenase activity. In the absence of L-glutamine, L-serine and the methyl ester of L-phenylalanine, but neither L...... glutamate dehydrogenase activity showed that only the two methyl esters of L-phenylalanine and L-serine activated the enzyme. It is concluded that the mechanism by which methyl esters of amino acids potentiate insulin release is most likely to be mediated by the activation of pancreatic beta-cell glutamate...

  19. Compartmentalized activities of the pyruvate dehydrogenase complex sustain lipogenesis in prostate cancer.

    Science.gov (United States)

    Chen, Jingjing; Guccini, Ilaria; Mitri, Diletta Di; Brina, Daniela; Revandkar, Ajinkya; Sarti, Manuela; Pasquini, Emiliano; Alajati, Abdullah; Pinton, Sandra; Losa, Marco; Civenni, Gianluca; Catapano, Carlo V; Sgrignani, Jacopo; Cavalli, Andrea; D'Antuono, Rocco; Asara, John M; Morandi, Andrea; Chiarugi, Paola; Crotti, Sara; Agostini, Marco; Montopoli, Monica; Masgras, Ionica; Rasola, Andrea; Garcia-Escudero, Ramon; Delaleu, Nicolas; Rinaldi, Andrea; Bertoni, Francesco; Bono, Johann de; Carracedo, Arkaitz; Alimonti, Andrea

    2018-02-01

    The mechanisms by which mitochondrial metabolism supports cancer anabolism remain unclear. Here, we found that genetic and pharmacological inactivation of pyruvate dehydrogenase A1 (PDHA1), a subunit of the pyruvate dehydrogenase complex (PDC), inhibits prostate cancer development in mouse and human xenograft tumor models by affecting lipid biosynthesis. Mechanistically, we show that in prostate cancer, PDC localizes in both the mitochondria and the nucleus. Whereas nuclear PDC controls the expression of sterol regulatory element-binding transcription factor (SREBF)-target genes by mediating histone acetylation, mitochondrial PDC provides cytosolic citrate for lipid synthesis in a coordinated manner, thereby sustaining anabolism. Additionally, we found that PDHA1 and the PDC activator pyruvate dehydrogenase phosphatase 1 (PDP1) are frequently amplified and overexpressed at both the gene and protein levels in prostate tumors. Together, these findings demonstrate that both mitochondrial and nuclear PDC sustain prostate tumorigenesis by controlling lipid biosynthesis, thus suggesting this complex as a potential target for cancer therapy.

  20. Amino ketone formation and aminopropanol-dehydrogenase activity in rat-liver preparations

    Science.gov (United States)

    Turner, J. M.; Willetts, A. J.

    1967-01-01

    1. Rat tissue homogenates convert dl-1-aminopropan-2-ol into aminoacetone. Liver homogenates have relatively high aminopropanol-dehydrogenase activity compared with kidney, heart, spleen and muscle preparations. 2. Maximum activity of liver homogenates is exhibited at pH9·8. The Km for aminopropanol is approx. 15mm, calculated for a single enantiomorph, and the maximum activity is approx. 9mμmoles of aminoacetone formed/mg. wet wt. of liver/hr.at 37°. Aminoacetone is also formed from l-threonine, but less rapidly. An unidentified amino ketone is formed from dl-4-amino-3-hydroxybutyrate, the Km for which is approx. 200mm at pH9·8. 3. Aminopropanol-dehydrogenase activity in homogenates is inhibited non-competitively by dl-3-hydroxybutyrate, the Ki being approx. 200mm. EDTA and other chelating agents are weakly inhibitory, and whereas potassium chloride activates slightly at low concentrations, inhibition occurs at 50–100mm. 4. It is concluded that aminopropanol-dehydrogenase is located in mitochondria, and in contrast with l-threonine dehydrogenase can be readily solubilized from mitochondrial preparations by ultrasonic treatment. 5. Soluble extracts of disintegrated mitochondria exhibit maximum aminopropanol-dehydrogenase activity at pH9·1 At this pH, Km values for the amino alcohol and NAD+ are approx. 200 and 1·3mm respectively. Under optimum conditions the maximum velocity is approx. 70mμmoles of aminoacetone formed/mg. of protein/hr. at 37°. Chelating agents and thiol reagents appear to have little effect on enzyme activity, but potassium chloride inhibits at all concentrations tested up to 80mm. dl-3-Hydroxybutyrate is only slightly inhibitory. 6. Dehydrogenase activities for l-threonine and dl-4-amino-3-hydroxybutyrate appear to be distinct from that for aminopropanol. 7. Intraperitoneal injection of aminopropanol into rats leads to excretion of aminoacetone in the urine. Aminoacetone excretion proportional to the amount of the amino alcohol

  1. Evolution of D-lactate dehydrogenase activity from glycerol dehydrogenase and its utility for D-lactate production from lignocellulose

    Science.gov (United States)

    Wang, Qingzhao; Ingram, Lonnie O.; Shanmugam, K. T.

    2011-01-01

    Lactic acid, an attractive, renewable chemical for production of biobased plastics (polylactic acid, PLA), is currently commercially produced from food-based sources of sugar. Pure optical isomers of lactate needed for PLA are typically produced by microbial fermentation of sugars at temperatures below 40 °C. Bacillus coagulans produces L(+)-lactate as a primary fermentation product and grows optimally at 50 °C and pH 5, conditions that are optimal for activity of commercial fungal cellulases. This strain was engineered to produce D(−)-lactate by deleting the native ldh (L-lactate dehydrogenase) and alsS (acetolactate synthase) genes to impede anaerobic growth, followed by growth-based selection to isolate suppressor mutants that restored growth. One of these, strain QZ19, produced about 90 g L-1 of optically pure D(−)-lactic acid from glucose in coagulans and the QZ19 derivative can be used to produce either L(+) or D(−) optical isomers of lactic acid (respectively) at high titers and yields from nonfood carbohydrates. PMID:22065761

  2. Exercise-induced pyruvate dehydrogenase activation is not affected by 7 days of bed rest

    DEFF Research Database (Denmark)

    Kiilerich, Kristian; Jørgensen, Stine Ringholm; Biensø, Rasmus Sjørup

    2011-01-01

    To test the hypothesis that physical inactivity impairs the exercise-induced modulation of pyruvate dehydrogenase (PDH), 6 healthy normally physically active male subjects completed 7 days of bed rest. Before and immediately after the bed rest, the subjects completed an OGTT and a one-legged knee...

  3. In situ glucose-6-phosphate dehydrogenase activity during development of pre-implantation mouse embryos

    NARCIS (Netherlands)

    de Schepper, G. G.; Vander Perk, C.; Westerveld, A.; Oosting, J.; van Noorden, C. J.

    1993-01-01

    Glucose-6-phosphate dehydrogenase activity was analysed cytophotometrically in oocytes and pre-implantation embryos of mice. A bimodal distribution pattern was not found. Therefore, female and male embryos could not be discriminated on the basis of linkage of the enzyme with the X-chromosome during

  4. Activity and isozyme content of lactate dehydrogenase under long-term oral taurine administration to rats

    Directory of Open Access Journals (Sweden)

    R. D. Ostapiv

    2015-08-01

    Full Text Available The effect of long-term oral taurine administration to rats on activity of lactate dehydrogenase (LDH, its isozyme content and activity in the whole blood, liver, thigh muscle, brain and testes tissues were studied in the present work. For this purpose male Wistar rats with body weight 190–220 g were randomly divided into three groups, they were orally administered drinking water (control group or taurine solution 40 and 100 mg per kg of body weight ( groups I and II, respectively. The total lactate dehydrogenase activity was measured spectrophotometrically, the percentage content of isozymes was determined by electrophoresis in 7.5% poliacrylamide gel with further staining according to J. Garbus. It was found that the total lactate dehydrogenase activity increased in all studied tissues. In testes of animals of both groups and in brain of group I animals, the total percentage contents of isozymes that are responsible for lactate production (LDH4+LDH5 increased. In liver of animals of both groups and in whole blood of group II animals, the total percentage content of isozymes that produce pyruvate (LDH1+LDH2 increased. In thigh muscle of both groups and in brain of group II animals the balance between LDH1+LDH2 and LDH4+LDH5 content did not differ from control values, though total lactate dehydrogenase activity was significantly higher, than that in the control group. Thus, the increase in the lactate dehydrogenase activity under long-term oral taurine administration in different rat tissues was found to be tissue- and dose-dependent and was caused by the increase in the content of different isozymes. Such increase in group I animals might be explained by adaptive mechanisms to hypoxia caused by high doses of taurine. For group II animals high doses of taurine were toxic and directly affected metabolic processes in the animal bodies.

  5. ACTIVITY AND ISOZYME CONTENT OF LACTATE DEHYDROGENASE UNDER LONG-TERM ORAL TAURINE ADMINISTRATION TO RATS.

    Science.gov (United States)

    Ostapiv, R D; Humenyuk, S L; Manko, V V

    2015-01-01

    The effect of long-term oral taurine administration to rats on activity of lactate dehydrogenase (LDH), its isozyme content and activity in the whole blood, liver, thigh muscle, brain and testes tissues were studied in the present work. For this purpose male Wistar rats with body weight 190-220 g were randomly divided into three groups, they were orally administered drinking water (control group) or taurine solution 40 and 100 mg per kg of body weight ( groups I and II, respectively). The total lactate dehydrogenase activity was measured spectrophotometrically, the percentage content of isozymes was determined by electrophoresis in 7.5% poliacrylamide gel withfurther staining according to J. Garbus. It was found that the total lactate dehydrogenase activity increased in all studied tissues. In testes of animals of both groups and in brain of group I animals, the total percentage contents of isozymes that are responsible for lactate production (LDH4+LDH5) increased. In liver of animals of both groups and in whole blood of group II animals, the total percentage content of isozymes that produce pyruvate (LDH1+LDH2) increased. In thigh muscle of both groups and in brain of group II animals the balance between LDH1+LDH2 and LDH4+LDH5 content did not differ from control values, though total lactate dehydrogenase activity was significantly higher, than that in the control group. Thus, the increase in the lactate dehydrogenase activity under long-term oral taurine administration in different rat tissues was found to be tissue- and dose-dependent and was caused by the increase in the content of different isozymes. Such increase in group I animals might be explained by adaptive mechanisms to hypoxia caused by high doses of taurine. For group II animals high doses of taurine were toxic and directly affected metabolic processes in the animal bodies.

  6. Simultaneous immobilization of dehydrogenases on polyvinylidene difluoride resin after separation by non-denaturing two-dimensional electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Shimazaki, Youji [Graduate School of Science and Engineering (Science Section) and Venture Business Laboratory, Ehime University, Bunkyo-cho 2-5, Matsuyama City 790-8577 (Japan)], E-mail: yoji@dpc.ehime-u.ac.jp; Kadota, Mariko [Faculty of Science, Ehime University, Matsuyama (Japan)

    2008-06-16

    We detected mouse liver malate, sorbitol and aldehyde dehydrogenases by negative staining, analysis of malate and sorbitol dehydrogenase activities using each substrate, and electron transfers including nicotinamide adenine dinucleotide (NAD) and nitroblue tetrazolium in non-denaturing two-dimensional electrophoresis (2-DE) gel. Dehydrogenases were also identified by electrospray ionization tandem mass spectrometry (ESI-MS/MS) after 2-DE separation and protein detection by negative staining. Spots of dehydrogenases separated by 2-DE were excised, and simultaneously transferred and immobilized on polyvinylidene difuoride (PVDF) resin by electrophoresis. The dehydrogenase activities remained intact after immobilization. In conclusion, resin-immobilized dehydrogenases can be simultaneously obtained after separation by non-denaturing 2-DE, detection by negative staining and transferring to resins.

  7. Human cord blood progenitors with high aldehyde dehydrogenase activity improve vascular density in a model of acute myocardial infarction

    Directory of Open Access Journals (Sweden)

    Creer Michael H

    2010-03-01

    Full Text Available Abstract Human stem cells from adult sources have been shown to contribute to the regeneration of muscle, liver, heart, and vasculature. The mechanisms by which this is accomplished are, however, still not well understood. We tested the engraftment and regenerative potential of human umbilical cord blood-derived ALDHhiLin-, and ALDHloLin- cells following transplantation to NOD/SCID or NOD/SCID β2m null mice with experimentally induced acute myocardial infarction. We used combined nanoparticle labeling and whole organ fluorescent imaging to detect human cells in multiple organs 48 hours post transplantation. Engraftment and regenerative effects of cell treatment were assessed four weeks post transplantation. We found that ALDHhiLin- stem cells specifically located to the site of injury 48 hours post transplantation and engrafted the infarcted heart at higher frequencies than ALDHloLin- committed progenitor cells four weeks post transplantation. We found no donor derived cardiomyocytes and few endothelial cells of donor origin. Cell treatment was not associated with any detectable functional improvement at the four week endpoint. There was, however, a significant increase in vascular density in the central infarct zone of ALDHhiLin- cell-treated mice, as compared to PBS and ALDHloLin- cell-treated mice. Conclusions Our data indicate that adult human stem cells do not become a significant part of the regenerating tissue, but rapidly home to and persist only temporarily at the site of hypoxic injury to exert trophic effects on tissue repair thereby enhancing vascular recovery.

  8. A novel aldehyde dehydrogenase-3 activator (Alda-89) protects submandibular gland function from irradiation without accelerating tumor growth.

    Science.gov (United States)

    Xiao, Nan; Cao, Hongbin; Chen, Che-Hong; Kong, Christina S; Ali, Rehan; Chan, Cato; Sirjani, Davud; Graves, Edward; Koong, Albert; Giaccia, Amato; Mochly-Rosen, Daria; Le, Quynh-Thu

    2013-08-15

    To determine the effect of Alda-89 (an ALDH3 activitor) on (i) the function of irradiated (radiotherapy) submandibular gland (SMG) in mice, (ii) its toxicity profile, and (iii) its effect on the growth of head and neck cancer (HNC) in vitro and in vivo. Adult mice were infused with Alda-89 or vehicle before, during, and after radiotherapy. Saliva secretion was monitored weekly. Hematology, metabolic profile, and postmortem evaluation for toxicity were examined at the time of sacrifice. Alda-89 or vehicle was applied to HNC cell lines in vitro, and severe combined immunodeficient (SCID) mice transplanted with HNC in vivo with or without radiation; HNC growth was monitored. The ALDH3A1 and ALDH3A2 protein expression was evaluated in 89 patients with HNC and correlated to freedom from relapse (FFR) and overall survival (OS). Alda-89 infusion significantly resulted in more whole saliva production and a higher percentage of preserved acini after radiotherapy compared with vehicle control. There was no difference in the complete blood count, metabolic profile, and major organ morphology between the Alda-89 and vehicle groups. Compared with vehicle control, Alda-89 treatment neither accelerated HNC cell proliferation in vitro, nor did it affect tumor growth in vivo with or without radiotherapy. Higher expression of ALDH3A1 or ALDH3A2 was not significantly associated with worse FFR or OS in either human papillomavirus (HPV)-positive or HPV-negative group. Alda-89 preserves salivary function after radiotherapy without affecting HNC growth or causing measurable toxicity in mice. It is a promising candidate to mitigate radiotherapy-related xerostomia. ©2013 AACR.

  9. Role of cysteine residues in the structure, stability, and alkane producing activity of cyanobacterial aldehyde deformylating oxygenase.

    Directory of Open Access Journals (Sweden)

    Yuuki Hayashi

    Full Text Available Aldehyde deformylating oxygenase (AD is a key enzyme for alkane biosynthesis in cyanobacteria, and it can be used as a catalyst for alkane production in vitro and in vivo. However, three free Cys residues in AD may impair its catalytic activity by undesired disulfide bond formation and oxidation. To develop Cys-deficient mutants of AD, we examined the roles of the Cys residues in the structure, stability, and alkane producing activity of AD from Nostoc punctiforme PCC 73102 by systematic Cys-to-Ala/Ser mutagenesis. The C71A/S mutations reduced the hydrocarbon producing activity of AD and facilitated the formation of a dimer, indicating that the conserved Cys71, which is located in close proximity to the substrate-binding site, plays crucial roles in maintaining the activity, structure, and stability of AD. On the other hand, mutations at Cys107 and Cys117 did not affect the hydrocarbon producing activity of AD. Therefore, we propose that the C107A/C117A double mutant is preferable to wild type AD for alkane production and that the double mutant may be used as a pseudo-wild type protein for further improvement of the alkane producing activity of AD.

  10. Glucose -6- phosphate dehydrogenase (g6pd) activity and ...

    African Journals Online (AJOL)

    The enzyme activity was determined quantitatively by spectrophotometer assay method. The activity of red cell G6PD enzyme was subnormal in 20% of the population studied. This agrees with previous report of the prevalence of G6PD deficiency in Nigerian males from the Western region of the country which is between 20 ...

  11. YNL134C from Saccharomyces cerevisiae encodes a novel protein with aldehyde reductase activity for detoxification of furfural derived from lignocellulosic biomass.

    Science.gov (United States)

    Zhao, Xianxian; Tang, Juan; Wang, Xu; Yang, Ruoheng; Zhang, Xiaoping; Gu, Yunfu; Li, Xi; Ma, Menggen

    2015-05-01

    Furfural and 5-hydroxymethylfurfural (HMF) are the two main aldehyde compounds derived from pentoses and hexoses, respectively, during lignocellulosic biomass pretreatment. These two compounds inhibit microbial growth and interfere with subsequent alcohol fermentation. Saccharomyces cerevisiae has the in situ ability to detoxify furfural and HMF to the less toxic 2-furanmethanol (FM) and furan-2,5-dimethanol (FDM), respectively. Herein, we report that an uncharacterized gene, YNL134C, was highly up-regulated under furfural or HMF stress and Yap1p and Msn2/4p transcription factors likely controlled its up-regulated expression. Enzyme activity assays showed that YNL134C is an NADH-dependent aldehyde reductase, which plays a role in detoxification of furfural to FM. However, no NADH- or NADPH-dependent enzyme activity was observed for detoxification of HMF to FDM. This enzyme did not catalyse the reverse reaction of FM to furfural or FDM to HMF. Further studies showed that YNL134C is a broad-substrate aldehyde reductase, which can reduce multiple aldehydes to their corresponding alcohols. Although YNL134C is grouped into the quinone oxidoreductase family, no quinone reductase activity was observed using 1,2-naphthoquinone or 9,10-phenanthrenequinone as a substrate, and phylogenetic analysis indicates that it is genetically distant to quinone reductases. Proteins similar to YNL134C in sequence from S. cerevisiae and other microorganisms were phylogenetically analysed. Copyright © 2015 John Wiley & Sons, Ltd.

  12. Easy access to aroma active unsaturated γ-lactones by addition of modified titanium homoenolate to aldehydes.

    Science.gov (United States)

    Frerot, Eric; Bagnoud, Alain

    2011-04-27

    The homo-Reformatsky reaction, in which a metal homoenolate of an ester is added to an aldehyde, was adapted to produce γ-lactones from unsaturated, enolizable aldehydes. By use of titanium homoenolate, 11 different γ-lactones were synthesized in one step with moderate to good yields from readily available aldehydes. In particular, this procedure allowed the rapid preparation of a series of C(12) unsaturated γ-lactones differing in the position and configuration of the double bond. These reference compounds will be used to identify previously unknown lactones in butter oil. The chromatographic, spectral, and sensory descriptions of the synthesized lactones are provided.

  13. 15-hydroxyprostaglandin dehydrogenase activity in vitro in lung and kidney of essential fatty acid-deficient rats

    DEFF Research Database (Denmark)

    Hansen, Harald S.; Toft, B.S.

    1978-01-01

    Weanling rats were fed for 6 months on a diet deficient in essential fatty acids: either fat-free, or with 28% (w/w) partially hydrogenated fish oil. Control rats were fed a diet with 28% (w/w) arachis oil for 6 months. 15-Hydroxyprostaglandin dehydrogenase activity was determined as initial rates...... of the two groups on diets deficient in essential fatty acids as compared to the control group. No difference was observed in dehydrogenase activity in the kidneys. The dehydrogenase may be of importance for the regulation of the level of endogenous prostaglandins and, thus, a decrease in activity could...

  14. Serum creatine kinase and lactate dehydrogenase activities in ...

    African Journals Online (AJOL)

    Background and Objectives: There is the recognition of a pattern of elevations of serum enzymes in hyperthyroid and hypothyroid patients. The aims of this study were to determine the activities of serum creatine kinase (CK) and lactate deydrogenase (LDH) in thyroid disorders, and to evaluate the relationship between CK, ...

  15. Determination of dehydrogenase activities involved in D-glucose oxidation in Gluconobacter and Acetobacter strains

    Directory of Open Access Journals (Sweden)

    Florencia Sainz

    2016-08-01

    Full Text Available Acetic acid bacteria (AAB are known for rapid and incomplete oxidation of an extensively variety of alcohols and carbohydrates, resulting in the accumulation of organic acids as the final products. These oxidative fermentations in AAB are catalyzed by PQQ- or FAD- dependent membrane bound dehydrogenases. In the present study, the enzyme activity of the membrane bound dehydrogenases (membrane-bound PQQ-glucose dehydrogenase (mGDH, D-gluconate dehydrogenase (GADH and membrane-bound glycerol dehydrogenase (GLDH involved in the oxidation of D-glucose and D-gluconic acid (GA was determined in six strains of three different species of AAB (three natural and three type strains. Moreover, the effect of these activities on the production of related metabolites (GA, 2-keto-D-gluconic acid (2KGA and 5-keto-D-gluconic acid (5KGA was analyzed. The natural strains belonging to Gluconobacter showed a high mGDH activity and low activity in GADH and GLDH, whereas the A. malorum strain presented low activity in the three enzymes. Nevertheless, no correlation was observed between the activity of these enzymes and the concentration of the corresponding metabolites. In fact, all the tested strains were able to oxidize D-glucose to GA, being maximal at the late exponential phase of the AAB growth (24 h, which coincided with glucose exhaustion and the maximum mGDH activity. Instead, only some of the tested strains were capable of producing 2KGA and/or 5KGA. In the case of G. oxydans strains, no 2KGA production was detected which is related to the absence of GADH activity after 24 h, while in the remaining strains, detection of GADH activity after 24h resulted in a high accumulation of 2KGA. Therefore, it is possible to choose the best strain depending on the desired product composition.Moreover, the sequences of these genes were used to construct phylogenetic trees. According to the sequence of gcd, gene coding for mGDH, Acetobacter and Komagataeibacter were

  16. The effect of phenobarbitone on cytoplasmic NADP-linked dehydrogenase activities in rat liver.

    Science.gov (United States)

    Shas, S; Pearson, D J

    1978-02-13

    Phenobarbitone administered in drinking water (0.5 g/l) or by daily intraperitoneal injection (100 mg/kg body weight) consistently caused an elevation of hepatic NADP-linked malic enzyme in rats maintained on a pellet diet. Three to four days appeared to be required for maximum response. The effect was also observed in animals maintained on a protein rich diet, in which the basic hepatic malic enzyme activity was low, but not in animals maintained on a sucrose rich diet, in which the basic enzyme activity was almost twice normal. Methyl cholanthrene, administered by daily intraperitoneal injection (40 mg/kg body weight), resulted in elevated hepatic levels not only of malic enzyme but also of the pentose phosphate pathway dehydrogenases. The timing of the "starve-refeed" response of the hepatic NADP-linked dehydrogenases in phenobarbitone-treated rats was similar to that in controls, and similar maximum enzyme activities were reached. The role of cytoplasmic NADP-linked dehydrogenases in the provision of reducing equivalent is discussed, particularly in relation to hepatic microsomal drug metabolism.

  17. Influence of adrenaline on the activity of succinate dehydrogenase in peripheral blood lymphocytes of irradiated rats

    International Nuclear Information System (INIS)

    Koroleva, L.V.; Vasin, M.V.

    1988-01-01

    In experiments with albino mongrel female rats, the influence of adrenaline on succinate dehydrogenase (SDG) activity in the peripheral blood lymphocytes of irradiated and intact animals has been investigated. Two minutes after the intraperitoneal administration of adrenaline (1 mg/kg) to intact rats SDG activity sharply rises and 3-4 min it drastically falls. In 6 to 8 min the second peak in the enzyme activity is registered. Twenty minutes after irradiation of rats in the crano-caudal direction with a dose of 75 Gy delivered to head, the reaction to adrenaline, manifested by the rise in SDG activity, is absent

  18. Diatom-derived polyunsaturated aldehydes activate cell death in human cancer cell lines but not normal cells.

    Directory of Open Access Journals (Sweden)

    Clementina Sansone

    Full Text Available Diatoms are an important class of unicellular algae that produce bioactive polyunsaturated aldehydes (PUAs that induce abortions or malformations in the offspring of invertebrates exposed to them during gestation. Here we compare the effects of the PUAs 2-trans,4-trans-decadienal (DD, 2-trans,4-trans-octadienal (OD and 2-trans,4-trans-heptadienal (HD on the adenocarcinoma cell lines lung A549 and colon COLO 205, and the normal lung/brunch epithelial BEAS-2B cell line. Using the viability MTT/Trypan blue assays, we show that PUAs have a toxic effect on both A549 and COLO 205 tumor cells but not BEAS-2B normal cells. DD was the strongest of the three PUAs tested, at all time-intervals considered, but HD was as strong as DD after 48 h. OD was the least active of the three PUAs. The effect of the three PUAs was somewhat stronger for A549 cells. We therefore studied the death signaling pathway activated in A549 showing that cells treated with DD activated Tumor Necrosis Factor Receptor 1 (TNFR1 and Fas Associated Death Domain (FADD leading to necroptosis via caspase-3 without activating the survival pathway Receptor-Interacting Protein (RIP. The TNFR1/FADD/caspase pathway was also observed with OD, but only after 48 h. This was the only PUA that activated RIP, consistent with the finding that OD causes less damage to the cell compared to DD and HD. In contrast, cells treated with HD activated the Fas/FADD/caspase pathway. This is the first report that PUAs activate an extrinsic apoptotic machinery in contrast to other anticancer drugs that promote an intrinsic death pathway, without affecting the viability of normal cells from the same tissue type. These findings have interesting implications also from the ecological viewpoint considering that HD is one of the most common PUAs produced by diatoms.

  19. Cytophotometric analysis of reaction rates of succinate and lactate dehydrogenase activity in rat liver, heart muscle and tracheal epithelium

    NARCIS (Netherlands)

    van Noorden, C. J.; Vogels, I. M.

    1989-01-01

    Reaction rates of succinate and lactate dehydrogenase activity in cryostat sections of rat liver, tracheal epithelium and heart muscle were monitored by continuous measurement of formazan formation by cytophotometry at room temperature. Incubation media contained polyvinyl alcohol as tissue

  20. SDR-type human hydroxysteroid dehydrogenases involved in steroid hormone activation.

    Science.gov (United States)

    Wu, Xiaoqiu; Lukacik, Petra; Kavanagh, Kathryn L; Oppermann, Udo

    2007-02-01

    Hydroxysteroid dehydrogenases catalyze the NAD(P)(H)-dependent oxidoreduction of hydroxyl and oxo-functions at distinct positions of steroid hormones. This reversible reaction constitutes an important pre-receptor control mechanism for nuclear receptor ligands of the androgen, estrogen and glucocorticoid classes, since the conversion "switches" between receptor ligands and their inactive metabolites. The major reversible activities found in mammals acting on steroid hormones comprise 3alpha-, 11beta- and 17beta-hydroxysteroid dehydrogenases, and for each group several distinct isozymes have been described. The enzymes differ in their expression pattern, nucleotide cofactor preference, steroid substrate specificity and subcellular localization, and thus constitute a complex system ensuring cell-specific adaptation and regulation of steroid hormone levels. Several isoforms constitute promising drug targets, of particular importance in cancer, metabolic diseases, neurodegeneration and immunity.

  1. Structural characteristics of active and inactive glutamate dehydrogenases from the hyperthermophile Pyrobaculum islandicum.

    Science.gov (United States)

    Ohshima, Toshihisa

    2012-01-01

    The enzymes from hyperthermophiles are generally extremely thermostable and lose little or no activity during long periods under a variety conditions. This high stability is very attractive, in that it gives the enzymes potential for use in numerous bioprocesses. My research group has investigated this high stability from the viewpoint of the relationship between function and structure. In this review, I describe the molecular mechanism underlying the extreme stability of unboiled NAD-dependent glutamate dehydrogenase from the hyperthermophile Pyrobaculum islandicum. I also describe the activation of the inactive recombinant enzyme produced in mesophilic Escherichia coli from the viewpoint of the relationship between structure and activity.

  2. Potentiation of insulin release in response to amino acid methyl esters correlates to activation of islet glutamate dehydrogenase activity

    DEFF Research Database (Denmark)

    Kofod, Hans; Lernmark, A; Hedeskov, C J

    1986-01-01

    Column perifusion of mouse pancreatic islets was used to study the ability of amino acids and their methyl esters to influence insulin release and activate islet glutamate dehydrogenase activity. In the absence of L-glutamine, L-serine and the methyl ester of L-phenylalanine, but neither L......-phenylalanine nor L-serine methyl ester, stimulate insulin secretion. In the presence of L-glutamine, however, the effect of L-serine was additive, while the methyl esters of L-serine and L-phenylalanine as well as native L-phenylalanine, potentiated the glucose-stimulated release of insulin. Measurements of islet...... glutamate dehydrogenase activity showed that only the two methyl esters of L-phenylalanine and L-serine activated the enzyme. It is concluded that the mechanism by which methyl esters of amino acids potentiate insulin release is most likely to be mediated by the activation of pancreatic beta-cell glutamate...

  3. Synthesis of Aryl-(E)-2-(azidomethyl)alkenoate and Aryl-(Z)-2-(azidomethyl)acrylonitrile from Aryl Aldehydes and Activated Alkenes via One-Pot Way

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Sang-Jin; Yang, Hae-Won; Han, Tae-Hwi; Yoon, Cheol Min [Korea University, Sejong (Korea, Republic of)

    2016-05-15

    Aryl-(E)-2-(azidomethyl)alkenoate and aryl-(Z)-2-(azidomethyl)acrylonitrile from aryl aldehydes and activated alkenes (acrylate and acrylonitrile) were synthesized in one-pot consecutive way with excellent stereoselectivity in good to high yield. The method include a three-step one-pot processes: Baylis-Hillman reaction of aryl aldehydes and activated alkenes using DABCO followed by acetylation using acetic anhydride and a catalytic amount of DMAP and nucleophilic substitution using sodium azide in DMF at room temperature. Our one-pot synthetic protocol is efficient and simple. Organoazides are one of the most important synthetic intermediates for the preparation of nitrogen-containing organic compounds, for example, amines via reduction, imines via rearrangement, and other nitrogen-containing heterocycles via cycloaddition reactions. Because allyl azides among them are versatile building skeletons for the synthesis of biologically active nitrogen-containing heterocycles, the synthesis of allyl azides have become an attractive research area.

  4. Proline dehydrogenase activity of the transcriptional repressor PutA is required for induction of the put operon by proline.

    Science.gov (United States)

    Muro-Pastor, A M; Maloy, S

    1995-04-28

    The proline utilization (put) operon from Salmonella typhimurium consists of the putP gene, encoding a proline transporter, and the putA gene, encoding an enzyme with both proline dehydrogenase and 1-pyrroline-5-carboxylate dehydrogenase activities. In addition to these two enzymatic activities, the PutA protein is a transcriptional repressor that regulates the expression of putP and putA in response to the availability of proline. We report the isolation of super-repressor mutants of PutA that decrease expression from the putA promoter in the presence or absence of proline. None of the mutants exhibited increased affinity for the DNA in the put regulatory region in vitro. Although DNA binding by wild-type PutA was prevented by the addition of proline and an artificial electron acceptor, DNA binding by the two strongest super-repressors was not prevented under identical conditions. The proline dehydrogenase activity of the purified mutant proteins showed altered kinetic properties (increased Km(Pro), reduced Vmax, or a completely null phenotype). The observation that these mutations simultaneously affect induction by proline and proline dehydrogenase activity suggests that a single proline-binding site is involved in both proline dehydrogenase activity and induction of the expression of the put operon. Furthermore, the results indicate that the proline dehydrogenase activity of PutA is essential for induction of the put operon by proline.

  5. Changes of α-glycerophosphate dehydrogenase activity in fatty liver of rats by amino acid imbalance

    International Nuclear Information System (INIS)

    Ogura, Masaji; Katsunuma, Eiichi; Akabane, Tomoko; Ogawa, Seiichi

    1976-01-01

    The previous study on the lipogenesis in the fatty livers of rats, which was induced by feeding the diet with imbalanced amino acid, revealed that the induction of this type of fatty livers was due mainly to the acceleration of triglyceride synthesis by the increase in both synthesis and esterification of fatty acid in the livers. Although many studies have been carried out on the dietary control of α-glycerophosphate dehydrogenase activity in rat livers, the enzyme change in amino acid imbalance has not been reported. In the present study, in order to elucidate the difference in the supply of glycerol moiety of triglyceride due to the imbalance, the change of the α-glycerophosphate dehydrogenase activity in livers was investigated. The experimental diets were 8% casein basal diet and basal + 0.3% DL-methionine imbalanced diet. 5 rats of each group were killed after 0.5 and 10 days on the diet, and the analysis of the lipid content in the livers and the determination of the α-glycerophosphate dehydrogenase activity were carried out. The linear response of the enzyme activity to time and protein concentration was obtained. The development of fatty livers was observed in the imbalanced diet group in the feeding period of 10 days. It was found that the specific activity of the imbalanced diet group increased significantly in 5 and 10 days as compared with that of the basal diet group. The elevation in the enzyme activity may suggest that the supply of α-glycerophosphate for triglyceride synthesis is also increased in this type of fatty livers. (Kako, I.)

  6. Differential effects of acute and chronic fructose administration on pyruvate dehydrogenase activity and lipogenesis

    International Nuclear Information System (INIS)

    Wilson, L.

    1988-01-01

    These studies were undertaken to distinguish between the acute and chronic effects of fructose administration. In vivo, liver lipogenesis, as measured by 3 H 2 O incorporation, was greater in rats fed 60% fructose than in their glucose fed controls. Both fructose feeding, and fructose feeding plus intraperitoneal fructose injection increased the activities of 6-phosphogluconate dehydrogenase and malic enzyme. Liver PDH activity was increased by fructose feeding, and was increased even more by fructose feeding and injection of fructose, but this was not associated with any changes in hepatic ATP concentrations

  7. Differential effects of acute and chronic fructose administration on pyruvate dehydrogenase activity and lipogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Wilson, L.

    1988-01-01

    These studies were undertaken to distinguish between the acute and chronic effects of fructose administration. In vivo, liver lipogenesis, as measured by {sup 3}H{sub 2}O incorporation, was greater in rats fed 60% fructose than in their glucose fed controls. Both fructose feeding, and fructose feeding plus intraperitoneal fructose injection increased the activities of 6-phosphogluconate dehydrogenase and malic enzyme. Liver PDH activity was increased by fructose feeding, and was increased even more by fructose feeding and injection of fructose, but this was not associated with any changes in hepatic ATP concentrations.

  8. Physiological and fermentation properties of Bacillus coagulans and a mutant lacking fermentative lactate dehydrogenase activity.

    Science.gov (United States)

    Su, Yue; Rhee, Mun Su; Ingram, Lonnie O; Shanmugam, K T

    2011-03-01

    Bacillus coagulans, a sporogenic lactic acid bacterium, grows optimally at 50-55 °C and produces lactic acid as the primary fermentation product from both hexoses and pentoses. The amount of fungal cellulases required for simultaneous saccharification and fermentation (SSF) at 55 °C was previously reported to be three to four times lower than for SSF at the optimum growth temperature for Saccharomyces cerevisiae of 35 °C. An ethanologenic B. coagulans is expected to lower the cellulase loading and production cost of cellulosic ethanol due to SSF at 55 °C. As a first step towards developing B. coagulans as an ethanologenic microbial biocatalyst, activity of the primary fermentation enzyme L-lactate dehydrogenase was removed by mutation (strain Suy27). Strain Suy27 produced ethanol as the main fermentation product from glucose during growth at pH 7.0 (0.33 g ethanol per g glucose fermented). Pyruvate dehydrogenase (PDH) and alcohol dehydrogenase (ADH) acting in series contributed to about 55% of the ethanol produced by this mutant while pyruvate formate lyase and ADH were responsible for the remainder. Due to the absence of PDH activity in B. coagulans during fermentative growth at pH 5.0, the l-ldh mutant failed to grow anaerobically at pH 5.0. Strain Suy27-13, a derivative of the l-ldh mutant strain Suy27, that produced PDH activity during anaerobic growth at pH 5.0 grew at this pH and also produced ethanol as the fermentation product (0.39 g per g glucose). These results show that construction of an ethanologenic B. coagulans requires optimal expression of PDH activity in addition to the removal of the LDH activity to support growth and ethanol production.

  9. [Activity of liver mitochondrial NAD+-dependent dehydrogenases of the krebs cycle in rats with acetaminophen-induced hepatitis developed under conditions of alimentary protein deficiency].

    Science.gov (United States)

    Voloshchuk, O N; Kopylchuk, G P

    2016-01-01

    Activity of isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, malate dehydrogenase, and the NAD(+)/NADН ratio were studied in the liver mitochondrial fraction of rats with toxic hepatitis induced by acetaminophen under conditions of alimentary protein deprivation. Acetaminophen-induced hepatitis was characterized by a decrease of isocitrate dehydrogenase, α-ketoglutarate dehydrogenase and malate dehydrogenase activities, while the mitochondrial NAD(+)/NADН ratio remained at the control level. Modeling of acetaminophen-induced hepatitis in rats with alimentary protein caused a more pronounced decrease in the activity of NAD(+)-dependent dehydrogenases studied and a 2.2-fold increase of the mitochondrial NAD(+)/NADН ratio. This suggests that alimentary protein deprivation potentiated drug-induced liver damage.

  10. Effect of x-irradiation on succinate dehydrogenase activity in mouse Mus booduga

    International Nuclear Information System (INIS)

    Rajarami Reddy, G.; Pavan Kumar, T.; Vijayalakshmi, S.; Sasira Babu, K.

    1979-01-01

    Succinate dehydrogenase (SDH) activity in brain and liver of lethally X-irradiated Mus booduga exhibited increase (7 to 11 percent) during early (2 hrs) hours and a gradual fall reaching the normal level by 5th day of post-irradiation period. Enzyme substrate affinity, though it dropped during first three days, was highest by 5th day. But in sartorius muscle, both enzyme level and its affinity to substrate had decreased. It is suggested that brain and liver are more radioresistant than sartorius muscle. (auth.)

  11. Effects of Al(III) and nano-Al13 species on malate dehydrogenase activity.

    Science.gov (United States)

    Yang, Xiaodi; Cai, Ling; Peng, Yu; Li, Huihui; Chen, Rong Fu; Shen, Ren Fang

    2011-01-01

    The effects of different aluminum species on malate dehydrogenase (MDH) activity were investigated by monitoring amperometric i-t curves for the oxidation of NADH at low overpotential using a functionalized multi-wall nanotube (MWNT) modified glass carbon electrode (GCE). The results showed that Al(III) and Al(13) can activate the enzymatic activity of MDH, and the activation reaches maximum levels as the Al(III) and Al(13) concentration increase. Our study also found that the effects of Al(III) and Al(13) on the activity of MDH depended on the pH value and aluminum speciation. Electrochemical and circular dichroism spectra methods were applied to study the effects of nano-sized aluminum compounds on biomolecules.

  12. A Catalase-related Hemoprotein in Coral Is Specialized for Synthesis of Short-chain Aldehydes: DISCOVERY OF P450-TYPE HYDROPEROXIDE LYASE ACTIVITY IN A CATALASE.

    Science.gov (United States)

    Teder, Tarvi; Lõhelaid, Helike; Boeglin, William E; Calcutt, Wade M; Brash, Alan R; Samel, Nigulas

    2015-08-07

    In corals a catalase-lipoxygenase fusion protein transforms arachidonic acid to the allene oxide 8R,9-epoxy-5,9,11,14-eicosatetraenoic acid from which arise cyclopentenones such as the prostanoid-related clavulones. Recently we cloned two catalase-lipoxygenase fusion protein genes (a and b) from the coral Capnella imbricata, form a being an allene oxide synthase and form b giving uncharacterized polar products (Lõhelaid, H., Teder, T., Tõldsepp, K., Ekins, M., and Samel, N. (2014) PloS ONE 9, e89215). Here, using HPLC-UV, LC-MS, and NMR methods, we identify a novel activity of fusion protein b, establishing its role in cleaving the lipoxygenase product 8R-hydroperoxy-eicosatetraenoic acid into the short-chain aldehydes (5Z)-8-oxo-octenoic acid and (3Z,6Z)-dodecadienal; these primary products readily isomerize in an aqueous medium to the corresponding 6E- and 2E,6Z derivatives. This type of enzymatic cleavage, splitting the carbon chain within the conjugated diene of the hydroperoxide substrate, is known only in plant cytochrome P450 hydroperoxide lyases. In mechanistic studies using (18)O-labeled substrate and incubations in H2(18)O, we established synthesis of the C8-oxo acid and C12 aldehyde with the retention of the hydroperoxy oxygens, consistent with synthesis of a short-lived hemiacetal intermediate that breaks down spontaneously into the two aldehydes. Taken together with our initial studies indicating differing gene regulation of the allene oxide synthase and the newly identified catalase-related hydroperoxide lyase and given the role of aldehydes in plant defense, this work uncovers a potential pathway in coral stress signaling and a novel enzymatic activity in the animal kingdom. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. X-irradiation effects on the activity of dehydrogenases in the cockroach, Periplaneta Americana L

    Energy Technology Data Exchange (ETDEWEB)

    Vijayalakshmi, S. (Sri Sathya Sai Inst. of Higher Learning, Anantpur (India))

    1984-05-01

    Sublethal dose of X-irradiation caused an early increase and subsequent normalization in succinate and lactate dehydrogenases of the cockroach, while lethal dose produced an irreversible fall in succinate dehydrogenase and a gradual elevation in lactate dehydrogenase at all post-irradiation periods studied, suggesting dose dependent impairment of aerobic and anaerobic pathways.

  14. Comparative study of the activity of lactate dehydrogenase (LDH) in different forms of disease

    International Nuclear Information System (INIS)

    Gonzalez Quesada, Jorge; Jorquera Cortez, Rodrigo; Rivera Alvarez, Sonia

    2007-01-01

    The activity of lactate dehydrogenase (LDH) was determined in the fluid gingival crevicular (FGC) from different sites of the anterior sector of the oral cavity in a clinically healthy subjects, and other with moderate gingivitis and with chronic severe generalized periodontists. Patients were treated and followed for three months, after the which has proceeded to make measurements of activity in the same sites discussed above. The results have showed statistically significant differences when comparing the activity of LDH in healthy individuals, and in other patients, treated by the pathology that presenting. On the other hand, were found without statistically significant differences between patients treated with clinically healthy subjects. The conclusion has been that the activity of LDH could be a quantitative marker for assessing cellular damage and evolution of treatment. (author) [es

  15. Synthesis, structure, and antiproliferative activity of selenophenfurin, an inosine 5'-monophosphate dehydrogenase inhibitor analogue of selenazofurin.

    Science.gov (United States)

    Franchetti, P; Cappellacci, L; Sheikha, G A; Jayaram, H N; Gurudutt, V V; Sint, T; Schneider, B P; Jones, W D; Goldstein, B M; Perra, G; De Montis, A; Loi, A G; La Colla, P; Grifantini, M

    1997-05-23

    The synthesis and biological activity of selenophenfurin (5-beta-D-ribofuranosylselenophene-3-carboxamide, 1), the selenophene analogue of selenazofurin, are described. Glycosylation of ethyl selenophene-3-carboxylate (6) under stannic chloride-catalyzed conditions gave 2- and 5-glycosylated regioisomers, as a mixture of alpha- and beta-anomers, and the beta-2,5-diglycosylated derivative. Deprotected ethyl 5-beta-D-ribofuranosylselenophene-3-carboxylate (12 beta) was converted into selenophenfurin by ammonolysis. The structure of 12 beta was determined by 1H- and 13C-NMR, crystallographic, and computational studies. Selenophenfurin proved to be antiproliferative against a number of leukemia, lymphoma, and solid tumor cell lines at concentrations similar to those of selenazofurin but was more potent than the thiophene and thiazole analogues thiophenfurin and tiazofurin. Incubation of K562 cells with selenophenfurin resulted in inhibition of IMP dehydrogenase (IMPDH) (76%) and an increase in IMP pools (14.5-fold) with a concurrent decrease in GTP levels (58%). The results obtained confirm the hypothesis that the presence of heteroatoms such as S or Se in the heterocycle in position 2 with respect to the glycosidic bond is essential for both cytotoxicity and IMP dehydrogenase inhibitory activity in this type of C-nucleosides.

  16. Peroxisomal NADP-Dependent Isocitrate Dehydrogenase. Characterization and Activity Regulation during Natural Senescence1

    Science.gov (United States)

    Corpas, Francisco J.; Barroso, Juan B.; Sandalio, Luisa M.; Palma, José M.; Lupiáñez, José A.; del Río, Luis A.

    1999-01-01

    The peroxisomal localization and characterization of NADP-dependent isocitrate dehydrogenase (perICDH) in young and senescent pea (Pisum sativum) leaves was studied by subcellular fractionation, kinetic analysis, immunoblotting, and immunoelectron microscopy. The subunit molecular mass for perICDH determined by immunoblotting was 46 kD. By isoelectric focusing (IEF) of the peroxisomal matrix fraction, the NADP-ICDH activity was resolved into four isoforms, perICDH-1 to perICDH-4, with isoelectric points (pIs) of 6.0, 5.6, 5.4, and 5.2, respectively. The kinetic properties of the NADP-ICDH in peroxisomes from young and senescent pea leaves were analyzed. The maximum initial velocity was the same in peroxisomes from young and senescent leaves, while the Michaelis constant value in senescent leaf peroxisomes was 11-fold lower than in young leaf peroxisomes. The protein levels of NADP-ICDH in peroxisomes were not altered during senescence. The kinetic behavior of this enzyme suggests a possible fine control of enzymatic activity by modulation of its Michaelis constant during the natural senescence of pea leaves. After embedding, electron microscopy immunogold labeling of NADP-ICDH confirmed that this enzyme was localized in the peroxisomal matrix. Peroxisomal NADP-ICDH represents an alternative dehydrogenase in these cell organelles and may be the main system for the reduction of NADP to NADPH for its re-utilization in the peroxisomal metabolism. PMID:10557241

  17. Characterization of human DHRS4: an inducible short-chain dehydrogenase/reductase enzyme with 3beta-hydroxysteroid dehydrogenase activity.

    Science.gov (United States)

    Matsunaga, Toshiyuki; Endo, Satoshi; Maeda, Satoshi; Ishikura, Shuhei; Tajima, Kazuo; Tanaka, Nobutada; Nakamura, Kazuo T; Imamura, Yorishige; Hara, Akira

    2008-09-15

    Human DHRS4 is a peroxisomal member of the short-chain dehydrogenase/reductase superfamily, but its enzymatic properties, except for displaying NADP(H)-dependent retinol dehydrogenase/reductase activity, are unknown. We show that the human enzyme, a tetramer composed of 27kDa subunits, is inactivated at low temperature without dissociation into subunits. The cold inactivation was prevented by a mutation of Thr177 with the corresponding residue, Asn, in cold-stable pig DHRS4, where this residue is hydrogen-bonded to Asn165 in a substrate-binding loop of other subunit. Human DHRS4 reduced various aromatic ketones and alpha-dicarbonyl compounds including cytotoxic 9,10-phenanthrenequinone. The overexpression of the peroxisomal enzyme in cultured cells did not increase the cytotoxicity of 9,10-phenanthrenequinone. While its activity towards all-trans-retinal was low, human DHRS4 efficiently reduced 3-keto-C(19)/C(21)-steroids into 3beta-hydroxysteroids. The stereospecific conversion to 3beta-hydroxysteroids was observed in endothelial cells transfected with vectors expressing the enzyme. The mRNA for the enzyme was ubiquitously expressed in human tissues and several cancer cells, and the enzyme in HepG2 cells was induced by peroxisome-proliferator-activated receptor alpha ligands. The results suggest a novel mechanism of cold inactivation and role of the inducible human DHRS4 in 3beta-hydroxysteroid synthesis and xenobiotic carbonyl metabolism.

  18. Expression of the putA gene encoding proline dehydrogenase from Rhodobacter capsulatus is independent of NtrC regulation but requires an Lrp-like activator protein.

    Science.gov (United States)

    Keuntje, B; Masepohl, B; Klipp, W

    1995-11-01

    Four Rhodobacter capsulatus mutants unable to grow with proline as the sole nitrogen source were isolated by random Tn5 mutagenesis. The Tn5 insertions were mapped within two adjacent chromosomal EcoRI fragments. DNA sequence analysis of this region revealed three open reading frames designated selD, putR, and putA. The putA gene codes for a protein of 1,127 amino acid residues which is homologous to PutA of Salmonella typhimurium and Escherichia coli. The central part of R. capsulatus PutA showed homology to proline dehydrogenase of Saccharomyces cerevisiae (Put1) and Drosophila melanogaster (SlgA). The C-terminal part of PutA exhibited homology to Put2 (pyrroline-5-carboxylate dehydrogenase) of S. cerevisiae and to aldehyde dehydrogenases from different organisms. Therefore, it seems likely that in R. capsulatus, as in enteric bacteria, both enzymatic steps for proline degradation are catalyzed by a single polypeptide (PutA). The deduced amino acid sequence of PutR (154 amino acid residues) showed homology to the small regulatory proteins Lrp, BkdR, and AsnC. The putR gene, which is divergently transcribed from putA, is essential for proline utilization and codes for an activator of putA expression. The expression of putA was induced by proline and was not affected by ammonia or other amino acids. In addition, putA expression was autoregulated by PutA itself. Mutations in glnB, nifR1 (ntrC), and NifR4 (ntrA encoding sigma 54) had no influence on put gene expression. The open reading frame located downstream of R. capsulatus putR exhibited strong homology to the E. coli selD gene, which is involved in selenium metabolism. R. capsulatus selD mutants exhibited a Put+ phenotype, demonstrating that selD is required neither for viability nor for proline utilization.

  19. Designing a highly active soluble PQQ-glucose dehydrogenase for efficient glucose biosensors and biofuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Durand, Fabien [Universite de Bordeaux, Centre de Recherche Paul Pascal (CRPP), UPR 8641, Avenue Albert Schweitzer, 33600 Pessac (France); Stines-Chaumeil, Claire [Universite de Bordeaux, CNRS, Institut de Biochimie et de Genetique Cellulaires, 1 rue Camille Saint Saens, 33077 Bordeaux Cedex (France); Flexer, Victoria [Universite de Bordeaux, Centre de Recherche Paul Pascal (CRPP), UPR 8641, Avenue Albert Schweitzer, 33600 Pessac (France); Andre, Isabelle [Universite de Toulouse, INSA, UPS, INP, LISBP, 135 Avenue de Rangueil, F-31077 Toulouse (France); CNRS, UMR5504, F-31400 Toulouse (France); INRA, UMR 792 Ingenierie des Systemes Biologiques et des Procedes, F-31400 Toulouse (France); Mano, Nicolas, E-mail: mano@crpp-bordeaux.cnrs.fr [Universite de Bordeaux, Centre de Recherche Paul Pascal (CRPP), UPR 8641, Avenue Albert Schweitzer, 33600 Pessac (France)

    2010-11-26

    Research highlights: {yields} A new mutant of PQQ-GDH designed for glucose biosensors application. {yields} First mutant of PQQ-GDH with higher activity for D-glucose than the Wild type. {yields} Position N428 is a key point to increase the enzyme activity. {yields} Molecular modeling shows that the N428 C mutant displays a better interaction for PQQ than the WT. -- Abstract: We report for the first time a soluble PQQ-glucose dehydrogenase that is twice more active than the wild type for glucose oxidation and was obtained by combining site directed mutagenesis, modelling and steady-state kinetics. The observed enhancement is attributed to a better interaction between the cofactor and the enzyme leading to a better electron transfer. Electrochemical experiments also demonstrate the superiority of the new mutant for glucose oxidation and make it a promising enzyme for the development of high-performance glucose biosensors and biofuel cells.

  20. Study of the effect of radiotreatment on the enzymatic activity alcohol dehydrogenase of yeast

    International Nuclear Information System (INIS)

    Zehlila, Amel

    2006-01-01

    Gamma irradiation, applied to alcohol dehydrogenase at low doses, increases the catalytic activity and the stability of the enzyme. on the other hand, higher dose level of irradiation, that exceed 40 Gy for the commercial enzyme and 200 Gy for yeast cell, causes activity inhibition and some perturbation (imbalance) in the conformational structure of the YADH. Moreover, immobilization in alginic beads ameliorates the effect of ionising radiations since the enhancement in the thermo-resistance of the enzyme and the higher stability according to use number of entrapped ADH. Kinetics parameters, investigated in reveal a significant effect on the increase of the commercial enzyme affinity. Enhanced activity and stability of the enzyme prove the efficacy of gamma rays application in order to ameliorate the rate of some reactions. (author)

  1. Effect of different mulch materials on the soil dehydrogenase activity (DHA) in an organic pepper crop

    Science.gov (United States)

    Moreno, Marta M.; Peco, Jesús; Campos, Juan; Villena, Jaime; González, Sara; Moreno, Carmen

    2016-04-01

    The use biodegradable materials (biopolymers of different composition and papers) as an alternative to conventional mulches has increased considerably during the last years mainly for environmental reason. In order to assess the effect of these materials on the soil microbial activity during the season of a pepper crop organically grown in Central Spain, the soil dehydrogenase activity (DHA) was measured in laboratory. The mulch materials tested were: 1) black polyethylene (PE, 15 μm); black biopolymers (15 μm): 2) Mater-Bi® (corn starch based), 3) Sphere 4® (potato starch based), 4) Sphere 6® (potato starch based), 5) Bioflex® (polylactic acid based), 6) Ecovio® (polylactic acid based), 7) Mimgreen® (black paper, 85 g/m2). A randomized complete block design with four replications was adopted. The crop was drip irrigated following the water demand of each treatment. Soil samples (5-10 cm depth) under the different mulches were taken at different dates (at the beginning of the crop cycle and at different dates throughout the crop season). Additionally, samples of bare soil in a manual weeding and in an untreated control were taken. The results obtained show the negative effect of black PE on the DHA activity, mainly as result of the higher temperature reached under the mulch and the reduction in the gas interchange between the soil and the atmosphere. The values corresponding to the biodegradable materials were variable, although highlighting the low DHA activity observed under Bioflex®. In general, the uncovered treatments showed higher values than those reached under mulches, especially in the untreated control. Keywords: mulch, biodegradable, biopolymer, paper, dehydrogenase activity (DHA). Acknowledgements: the research was funded by Project RTA2011-00104-C04-03 from the INIA (Spanish Ministry of Economy and Competitiveness).

  2. Diabetes Impairs the Aldehyde Detoxifying Capacity of the Retina.

    Science.gov (United States)

    McDowell, Rosemary E; McGahon, Mary K; Augustine, Josy; Chen, Mei; McGeown, J Graham; Curtis, Tim M

    2016-09-01

    We studied whether the accumulation of advanced lipoxidation end-products (ALEs) in the diabetic retina is linked to the impairment of lipid aldehyde detoxification mechanisms. Retinas were collected from nondiabetic and diabetic rats and processed for conventional and quantitative RT-PCR (qRT-PCR), Western blotting, immunohistochemistry, and aldehyde dehydrogenase (ALDH) activity assays. The effect of the ALDH1a1 inhibitor, NCT-501, on ALE accumulation and cell viability in cultured Müller glia also was investigated. The rat retina expressed a range of lipid aldehyde detoxifying ALDH and aldo-keto reductase (AKR) genes. In diabetes, mRNA levels were reduced for 5 of 9 transcripts tested. These findings contrasted with those in the lens and cornea where many of these enzymes were upregulated. We have reported previously accumulation of the acrolein (ACR)-derived ALE, FDP-lysine, in retinal Müller glia during diabetes. In the present study, we show that the main ACR-detoxifying ALDH and AKR genes expressed in the retina, namely, ALDH1a1, ALDH2, and AKR1b1, are principally localized to Müller glia. Diabetes-induced FDP-lysine accumulation in Müller glia was associated with a reduction in ALDH1a1 mRNA and protein expression in whole retina and a decrease in ALDH1a1-immunoreactivity specifically within these cells. No such changes were detected for ALDH2 or AKR1b1. Activity of ALDH was suppressed in the diabetic retina and blockade of ALDH1a1 in cultured Müller glia triggered FDP-lysine accumulation and reduced cell viability. These findings suggest that downregulation of ALDH and AKR enzymes, particularly ALDH1a1, may contribute ALE accumulation in the diabetic retina.

  3. Sulfide dehydrogenase activity of the monomeric flavoprotein SoxF of Paracoccus pantotrophus.

    Science.gov (United States)

    Quentmeier, Armin; Hellwig, Petra; Bardischewsky, Frank; Wichmann, Rolf; Friedrich, Cornelius G

    2004-11-23

    Flavocytochrome c-sulfide dehydrogenases (FCSDs) are complexes of a flavoprotein with a c-type cytochrome performing hydrogen sulfide-dependent cytochrome c reduction in vitro. The amino acid sequence analysis revealed that the phylogenetic relationship of different flavoproteins reflected the relationship of sulfur-oxidizing bacteria. The flavoprotein SoxF of Paracoccus pantotrophus is 29-67% identical to the flavoprotein subunit of FCSD of phototrophic sulfur-oxidizing bacteria. Purification of SoxF yielded a homogeneous emerald-green monomeric protein of 42 797 Da. SoxF catalyzed sulfide-dependent horse heart cytochrome c reduction at the optimum pH of 6.0 with a k(cat) of 3.9 s(-1), a K(m) of 2.3 microM for sulfide, and a K(m) of 116 microM for cytochrome c, as determined by nonlinear regression analysis. The yield of 1.9 mol of cytochrome c reduced per mole of sulfide suggests sulfur or polysulfide as the product. Sulfide dehydrogenase activity of SoxF was inhibited by sulfur (K(i) = 1.3 microM) and inactivated by sulfite. Cyanide (1 mM) inhibited SoxF activity at pH 6.0 by 25% and at pH 8.0 by 92%. Redox titrations in the infrared spectral range from 1800 to 1200 cm(-1) and in the visible spectral range from 400 to 700 nm both yielded a midpoint potential for SoxF of -555 +/- 10 mV versus Ag/AgCl at pH 7.5 and -440 +/- 20 mV versus Ag/AgCl at pH 6.0 (-232 mV versus SHE') and a transfer of 1.9 electrons. Electrochemically induced FTIR difference spectra of SoxF as compared to those of free flavin in solution suggested a strong cofactor interaction with the apoprotein. Furthermore, an activation/variation of SoxF during the redox cycles is observed. This is the first report of a monomeric flavoprotein with sulfide dehydrogenase activity.

  4. Mechanisms of activation of muscle branched-chain alpha-keto acid dehydrogenase during exercise in man

    DEFF Research Database (Denmark)

    Van Hall, Gerrit; MacLean, D A; Saltin, B

    1996-01-01

    1. Exercise leads to activation (dephosphorylation) of the branched-chain alpha-keto acid dehydrogenase (BCKADH). Here we investigate the effect of low pre-exercise muscle glycogen content and of branched-chain amino acid (BCAA) ingestion on the activity of BCKADH at rest and after 90 min of one...

  5. Loss of succinate dehydrogenase activity results in dependency on pyruvate carboxylation for cellular anabolism

    Science.gov (United States)

    Lussey-Lepoutre, Charlotte; Hollinshead, Kate E. R.; Ludwig, Christian; Menara, Mélanie; Morin, Aurélie; Castro-Vega, Luis-Jaime; Parker, Seth J.; Janin, Maxime; Martinelli, Cosimo; Ottolenghi, Chris; Metallo, Christian; Gimenez-Roqueplo, Anne-Paule; Favier, Judith; Tennant, Daniel A.

    2015-01-01

    The tricarboxylic acid (TCA) cycle is a central metabolic pathway responsible for supplying reducing potential for oxidative phosphorylation and anabolic substrates for cell growth, repair and proliferation. As such it thought to be essential for cell proliferation and tissue homeostasis. However, since the initial report of an inactivating mutation in the TCA cycle enzyme complex, succinate dehydrogenase (SDH) in paraganglioma (PGL), it has become clear that some cells and tissues are not only able to survive with a truncated TCA cycle, but that they are also able of supporting proliferative phenotype observed in tumours. Here, we show that loss of SDH activity leads to changes in the metabolism of non-essential amino acids. In particular, we demonstrate that pyruvate carboxylase is essential to re-supply the depleted pool of aspartate in SDH-deficient cells. Our results demonstrate that the loss of SDH reduces the metabolic plasticity of cells, suggesting vulnerabilities that can be targeted therapeutically. PMID:26522426

  6. EFFECTS OF AMARANTHS’ SEEDS ON DEHYDROGENASE ACTIVITY AND GASES EMISSION IN METHANOGENIC BIOREACTORS

    Directory of Open Access Journals (Sweden)

    Victor COVALIOV

    2015-12-01

    Full Text Available The influence of amaranths‘ seeds as the source of squalene on the dehydrogenase activity and efficiency of methane production were investigated in methanogenic bench-scale (5000 ml bioreactors used to treat the mixture of distillery wastes and farmyard manure. The adding of amaranth seeds to the methanogenic bioreactor has an inhibitory effect on the dehydrogenase activity and stimulates the process of methanogenesis. Dehydrogenase activity decreased with the increase of doses of squalene and its trend had a close connection with doses (R2=0.77-0.78. The methane content in the total amount of gases is 65.3-71.3% in a bioreactor with the additive of amaranth seeds in a dose of 50 mg l-1, which is 22.1% higher than in the the control bioreactor without additives. The increase in squalene concentration higher than 0.0005% is not rational because its stimulating effect on the methanogenic process decreases. Anaerobic digestion of alcohol distillery industry wastes with manure is a complex nonlinear time-varying microbiological process. Dehydrogenase activity trends in the experiment are described by the power function for 5 hours observations and by the logarithmic function for 120 hours of observations. Trends of CH4 are described by the polynomial function in all periods of testing. Correlation coefficients are 0.37 and 0.70 for CH4 after 5 and 120 hours of the anaerobic digestion. Dehydrogenase activity is in the close negative connection with the amount of gases, including methane. Correlation analysis between dehydrogenase activity and the release of gases has revealed the moderate and strongly negative link during 24 hours after the start of the experiment.EFECTUL SEMINŢELOR DE AMARANT ASUPRA ACTIVITĂŢII DEHIDROGENAZEI ŞI EMISIEI GAZELOR ÎN BIOREACTOARELE METANOGENEÎn bioreactoare metanogene unite consecutiv, cu volum de 5000 ml, utilizate pentru tratarea amestecului de borhot de la distilarea alcoolului cu gunoi de grajd, a fost

  7. Lewis base activation of Lewis acids: catalytic, enantioselective addition of glycolate-derived silyl ketene acetals to aldehydes.

    Science.gov (United States)

    Denmark, Scott E; Chung, Won-Jin

    2008-06-20

    A catalytic system involving silicon tetrachloride and a chiral, Lewis basic bisphosphoramide catalyst is effective for the addition of glycolate-derived silyl ketene acetals to aldehydes. It was found that the sense of diastereoselectivity could be modulated by changing the size of the substituents on the silyl ketene acetals. In general, the trimethylsilyl ketene acetals derived from methyl glycolates with a large protecting group on the alpha-oxygen provide enantiomerically enriched alpha,beta-dihydroxy esters with high syn-diastereoselectivity, whereas the tert-butyldimethylsilyl ketene acetals derived from bulky esters of alpha-methoxyacetic acid provide enantiomerically enriched alpha,beta-dihydroxy esters with high anti-diastereoselecitvity.

  8. Vitality Improvement of the Mediterranean Fruit Fly, Ceratitis capitata Wied 1- Measured by using dehydrogenase Enzyme Activities

    International Nuclear Information System (INIS)

    Salama, M.S.; Shoman, A.A.; Elbermawy, S.M.; Abul Yazid, I.

    2000-01-01

    The present study searches for the improvement vitality of the Mediterranean fruit fly, Ceratitis capitata Wied. Through the induction of a specific variance (mutation) in the genetic material. Several types of treatments that were thought to cause this mutation were used, as IGR's, temperature, formaldehyde, colchicine, alcohols, several types of larval rearing media and gamma-rays. Generally, the activities of the energy enzymes alpha-glycerophosphate dehydrogenase (alpha-GPDH) enzyme lactate dehydrogenase (LDH) enzyme and malate dehydrogenase (MDH) enzyme, when used as a direct measure for the fly vitality, increased due to treatments of the egg stage by the previously mentioned treatments specially by the usage of rice hulls in the larval rearing medium alone or followed by irradiation of the pupal stage with 90 Gy

  9. Functional Role of Lanthanides in Enzymatic Activity and Transcriptional Regulation of Pyrroloquinoline Quinone-Dependent Alcohol Dehydrogenases inPseudomonas putidaKT2440.

    Science.gov (United States)

    Wehrmann, Matthias; Billard, Patrick; Martin-Meriadec, Audrey; Zegeye, Asfaw; Klebensberger, Janosch

    2017-06-27

    The oxidation of alcohols and aldehydes is crucial for detoxification and efficient catabolism of various volatile organic compounds (VOCs). Thus, many Gram-negative bacteria have evolved periplasmic oxidation systems based on pyrroloquinoline quinone-dependent alcohol dehydrogenases (PQQ-ADHs) that are often functionally redundant. Here we report the first description and characterization of a lanthanide-dependent PQQ-ADH (PedH) in a nonmethylotrophic bacterium based on the use of purified enzymes from the soil-dwelling model organism Pseudomonas putida KT2440. PedH (PP_2679) exhibits enzyme activity on a range of substrates similar to that of its Ca 2+ -dependent counterpart PedE (PP_2674), including linear and aromatic primary and secondary alcohols, as well as aldehydes, but only in the presence of lanthanide ions, including La 3+ , Ce 3+ , Pr 3+ , Sm 3+ , or Nd 3+ Reporter assays revealed that PedH not only has a catalytic function but is also involved in the transcriptional regulation of pedE and pedH , most likely acting as a sensory module. Notably, the underlying regulatory network is responsive to as little as 1 to 10 nM lanthanum, a concentration assumed to be of ecological relevance. The present study further demonstrates that the PQQ-dependent oxidation system is crucial for efficient growth with a variety of volatile alcohols. From these results, we conclude that functional redundancy and inverse regulation of PedE and PedH represent an adaptive strategy of P. putida KT2440 to optimize growth with volatile alcohols in response to the availability of different lanthanides. IMPORTANCE Because of their low bioavailability, lanthanides have long been considered biologically inert. In recent years, however, the identification of lanthanides as a cofactor in methylotrophic bacteria has attracted tremendous interest among various biological fields. The present study reveals that one of the two PQQ-ADHs produced by the model organism P. putida KT2440 also

  10. Synthesis and antifungal activity of nicotinamide derivatives as succinate dehydrogenase inhibitors.

    Science.gov (United States)

    Ye, Yong-Hao; Ma, Liang; Dai, Zhi-Cheng; Xiao, Yu; Zhang, Ying-Ying; Li, Dong-Dong; Wang, Jian-Xin; Zhu, Hai-Liang

    2014-05-07

    Thirty-eight nicotinamide derivatives were designed and synthesized as potential succinate dehydrogenase inhibitors (SDHI) and precisely characterized by (1)H NMR, ESI-MS, and elemental analysis. The compounds were evaluated against two phytopathogenic fungi, Rhizoctonia solani and Sclerotinia sclerotiorum, by mycelia growth inhibition assay in vitro. Most of the compounds displayed moderate activity, in which, 3a-17 exhibited the most potent antifungal activity against R. solani and S. sclerotiorum with IC50 values of 15.8 and 20.3 μM, respectively, comparable to those of the commonly used fungicides boscalid and carbendazim. The structure-activity relationship (SAR) of nicotinamide derivatives demonstrated that the meta-position of aniline was a key position contributing to the antifungal activity. Inhibition activities against two fungal SDHs were tested and achieved the same tendency with the data acquired from in vitro antifungal assay. Significantly, 3a-17 was demonstrated to successfully suppress disease development in S. sclerotiorum infected cole in vivo. In the molecular docking simulation, sulfur and chlorine of 3a-17 were bound with PHE291 and PRO150 of the SDH homology model, respectively, which could explain the probable mechanism of action between the inhibitory and target protein.

  11. Low intensity microwave radiation as modulator of the L-lactate dehydrogenase activity.

    Science.gov (United States)

    Vojisavljevic, Vuk; Pirogova, Elena; Cosic, Irena

    2011-07-01

    In this study, we investigated experimentally the possibility of modulating protein activity by low intensity microwaves by measuring alternations of L: -Lactate Dehydrogenase enzyme (LDH) activity. The LDH enzyme solutions were irradiated by microwaves of the selected frequencies and powers using the Transverse Electro-Magnetic (TEM) cell. The kinetics of the irradiated LDH was measured by continuous monitoring of nicotine adenine dinucleotide, reduced (NADH) absorbance at 340 nm. A comparative analysis of changes in the activity of the irradiated LDH enzyme versus the non-radiated enzyme was performed for the selected frequencies and powers. It was found that LDH activity can be selectively increased only by irradiation at the particular frequencies of 500 MHz [electric field: 0.02 V/m (1.2 × 10⁻⁶ W/m²)-2.1 V/m (1.2 × 10⁻² W/m²)] and 900 MHz [electric field: 0.021-0.21 V/m (1.2 × 10⁻⁴ W/m²)]. Based on results obtained it was concluded that LDH enzyme activity can be modulated by specific frequencies of low power microwave radiation. This finding can serve to support the hypothesis that low intensity microwaves can induce non-thermal effects in bio-molecules.

  12. Exercise training induces similar elevations in the activity of oxoglutarate dehydrogenase and peak oxygen uptake in the human quadriceps muscle

    DEFF Research Database (Denmark)

    Blomstrand, Eva; Krustrup, Peter; Søndergaard, Hans

    2011-01-01

    During exercise involving a small muscle mass, peak oxygen uptake is thought to be limited by peripheral factors, such as the degree of oxygen extraction from the blood and/or mitochondrial oxidative capacity. Previously, the maximal activity of the Krebs cycle enzyme oxoglutarate dehydrogenase h...

  13. Decreased succinate dehydrogenase activity of gamma and alpha motoneurons in mouse spinal cords following 13 weeks of exposure to microgravity.

    Science.gov (United States)

    Ishihara, Akihiko; Nagatomo, Fumiko; Fujino, Hidemi; Kondo, Hiroyo; Ohira, Yoshinobu

    2013-10-01

    Cell body size and succinate dehydrogenase activity of motoneurons in the dorsolateral region of the ventral horn in the lumbar and cervical segments of the mouse spinal cord were assessed after long-term exposure to microgravity and compared with those of ground-based controls. Mice were housed in a mouse drawer system on the International Space Station for 13 weeks. The mice were transported to the International Space Station by the Space Shuttle Discovery and returned to Earth by the Space Shuttle Atlantis. No changes in the cell body size of motoneurons were observed in either segment after exposure to microgravity, but succinate dehydrogenase activity of small-sized (700 μm(2)) alpha motoneurons, in both segments was lower than that of ground-based controls. We concluded that exposure to microgravity for longer than 3 months induced decreased succinate dehydrogenase activity of both gamma and slow-type alpha motoneurons. In particular, the decreased succinate dehydrogenase activity of gamma motoneurons was observed only after long-term exposure to microgravity.

  14. Reaction rate studies of glucose-6-phosphate dehydrogenase activity in sections of rat liver using four tetrazolium salts

    NARCIS (Netherlands)

    Butcher, R. G.; van Noorden, C. J.

    1985-01-01

    The reaction rate of glucose-6-phosphate dehydrogenase activity in liver sections from fed and starved rats has been monitored by the continuous measurement at 37 degrees C of the reaction product as it is formed using scanning and integrating microdensitometry. Control media lacked either substrate

  15. A sensitive cytochemical staining method for glucose-6-phosphate dehydrogenase activity in individual erythrocytes. I. Optimalization of the staining procedure

    NARCIS (Netherlands)

    van Noorden, C. J.; Vogels, I. M.; James, J.; Tas, J.

    1982-01-01

    A sensitive cytochemical staining method for glucose-6-phosphate dehydrogenase activity in individual human erythrocytes is described. This staining method can be used for the rapid routine discrimination of patients with a deficiency of the enzyme in its homozygote or heterozygote form, but also

  16. 15-hydroxyprostaglandin dehydrogenase activity in vitro in lung and kidney of essential fatty acid-deficient rats

    DEFF Research Database (Denmark)

    Hansen, Harald S.; Toft, B.S.

    1978-01-01

    of formation of H-labelled 15-keto-dihydro-prostaglandin E plus 15-keto-prostaglandin E in high speed supernatants of lung and kidney from each of the groups of rats. Dehydrogenase activity (expressed as either pmol/min per mg soluble protein, or as nmol/min per g tissue) was decreased 30-40% in the lungs...

  17. Determination of glutamate dehydrogenase activity and its kinetics in mouse tissues using metabolic mapping (quantitative enzyme histochemistry)

    NARCIS (Netherlands)

    Botman, Dennis; Tigchelaar, Wikky; van Noorden, Cornelis J. F.

    2014-01-01

    Glutamate dehydrogenase (GDH) catalyses the reversible conversion of glutamate into α-ketoglutarate with the concomitant reduction of NAD(P)(+) to NAD(P)H or vice versa. GDH activity is subject to complex allosteric regulation including substrate inhibition. To determine GDH kinetics in situ, we

  18. Kinetic and Structural Studies of Aldehyde Oxidoreductase from Desulfovibrio gigas Reveal a Dithiolene-Based Chemistry for Enzyme Activation and Inhibition by H2O2

    Science.gov (United States)

    Brondino, Carlos D.; Moura, José J. G.; Romão, Maria J.; González, Pablo J.; Santos-Silva, Teresa

    2013-01-01

    Mononuclear Mo-containing enzymes of the xanthine oxidase (XO) family catalyze the oxidative hydroxylation of aldehydes and heterocyclic compounds. The molybdenum active site shows a distorted square-pyramidal geometry in which two ligands, a hydroxyl/water molecule (the catalytic labile site) and a sulfido ligand, have been shown to be essential for catalysis. The XO family member aldehyde oxidoreductase from Desulfovibrio gigas (DgAOR) is an exception as presents in its catalytically competent form an equatorial oxo ligand instead of the sulfido ligand. Despite this structural difference, inactive samples of DgAOR can be activated upon incubation with dithionite plus sulfide, a procedure similar to that used for activation of desulfo-XO. The fact that DgAOR does not need a sulfido ligand for catalysis indicates that the process leading to the activation of inactive DgAOR samples is different to that of desulfo-XO. We now report a combined kinetic and X-ray crystallographic study to unveil the enzyme modification responsible for the inactivation and the chemistry that occurs at the Mo site when DgAOR is activated. In contrast to XO, which is activated by resulfuration of the Mo site, DgAOR activation/inactivation is governed by the oxidation state of the dithiolene moiety of the pyranopterin cofactor, which demonstrates the non-innocent behavior of the pyranopterin in enzyme activity. We also showed that DgAOR incubation with dithionite plus sulfide in the presence of dioxygen produces hydrogen peroxide not associated with the enzyme activation. The peroxide molecule coordinates to molybdenum in a η2 fashion inhibiting the enzyme activity. PMID:24391748

  19. Kinetic and structural studies of aldehyde oxidoreductase from Desulfovibrio gigas reveal a dithiolene-based chemistry for enzyme activation and inhibition by H(2O(2.

    Directory of Open Access Journals (Sweden)

    Jacopo Marangon

    Full Text Available Mononuclear Mo-containing enzymes of the xanthine oxidase (XO family catalyze the oxidative hydroxylation of aldehydes and heterocyclic compounds. The molybdenum active site shows a distorted square-pyramidal geometry in which two ligands, a hydroxyl/water molecule (the catalytic labile site and a sulfido ligand, have been shown to be essential for catalysis. The XO family member aldehyde oxidoreductase from Desulfovibrio gigas (DgAOR is an exception as presents in its catalytically competent form an equatorial oxo ligand instead of the sulfido ligand. Despite this structural difference, inactive samples of DgAOR can be activated upon incubation with dithionite plus sulfide, a procedure similar to that used for activation of desulfo-XO. The fact that DgAOR does not need a sulfido ligand for catalysis indicates that the process leading to the activation of inactive DgAOR samples is different to that of desulfo-XO. We now report a combined kinetic and X-ray crystallographic study to unveil the enzyme modification responsible for the inactivation and the chemistry that occurs at the Mo site when DgAOR is activated. In contrast to XO, which is activated by resulfuration of the Mo site, DgAOR activation/inactivation is governed by the oxidation state of the dithiolene moiety of the pyranopterin cofactor, which demonstrates the non-innocent behavior of the pyranopterin in enzyme activity. We also showed that DgAOR incubation with dithionite plus sulfide in the presence of dioxygen produces hydrogen peroxide not associated with the enzyme activation. The peroxide molecule coordinates to molybdenum in a η(2 fashion inhibiting the enzyme activity.

  20. Kinetic and structural studies of aldehyde oxidoreductase from Desulfovibrio gigas reveal a dithiolene-based chemistry for enzyme activation and inhibition by H(2)O(2).

    Science.gov (United States)

    Marangon, Jacopo; Correia, Hugo D; Brondino, Carlos D; Moura, José J G; Romão, Maria J; González, Pablo J; Santos-Silva, Teresa

    2013-01-01

    Mononuclear Mo-containing enzymes of the xanthine oxidase (XO) family catalyze the oxidative hydroxylation of aldehydes and heterocyclic compounds. The molybdenum active site shows a distorted square-pyramidal geometry in which two ligands, a hydroxyl/water molecule (the catalytic labile site) and a sulfido ligand, have been shown to be essential for catalysis. The XO family member aldehyde oxidoreductase from Desulfovibrio gigas (DgAOR) is an exception as presents in its catalytically competent form an equatorial oxo ligand instead of the sulfido ligand. Despite this structural difference, inactive samples of DgAOR can be activated upon incubation with dithionite plus sulfide, a procedure similar to that used for activation of desulfo-XO. The fact that DgAOR does not need a sulfido ligand for catalysis indicates that the process leading to the activation of inactive DgAOR samples is different to that of desulfo-XO. We now report a combined kinetic and X-ray crystallographic study to unveil the enzyme modification responsible for the inactivation and the chemistry that occurs at the Mo site when DgAOR is activated. In contrast to XO, which is activated by resulfuration of the Mo site, DgAOR activation/inactivation is governed by the oxidation state of the dithiolene moiety of the pyranopterin cofactor, which demonstrates the non-innocent behavior of the pyranopterin in enzyme activity. We also showed that DgAOR incubation with dithionite plus sulfide in the presence of dioxygen produces hydrogen peroxide not associated with the enzyme activation. The peroxide molecule coordinates to molybdenum in a η(2) fashion inhibiting the enzyme activity.

  1. Potential active-site residues in polyneuridine aldehyde esterase, a central enzyme of indole alkaloid biosynthesis, by modelling and site-directed mutagenesis.

    Science.gov (United States)

    Mattern-Dogru, Emine; Ma, Xueyan; Hartmann, Joachim; Decker, Heinz; Stöckigt, Joachim

    2002-06-01

    In the biosynthesis of the antiarrhythmic alkaloid ajmaline, polyneuridine aldehyde esterase (PNAE) catalyses a central reaction by transforming polyneuridine aldehyde into epi-vellosimine, which is the immediate precursor for the synthesis of the ajmalane skeleton. The PNAE cDNA was previously heterologously expressed in E. coli. Sequence alignments indicated that PNAE has a 43% identity to a hydroxynitrile lyase from Hevea brasiliensis, which is a member of the alpha/beta hydrolase superfamily. The catalytic triad, which is typical for this family, is conserved. By site-directed mutagenesis, the members of the catalytic triad were identified. For further detection of the active residues, a model of PNAE was constructed based on the X-ray crystallographic structure of hydroxynitrile lyase. The potential active site residues were selected on this model, and were mutated in order to better understand the relationship of PNAE with the alpha/beta hydrolases, and as well its mechanism of action. The results showed that PNAE is a novel member of the alpha/beta hydrolase enzyme superfamily.

  2. Lactate Dehydrogenase and Oxidative Stress Activity in Primary Open-Angle Glaucoma Aqueous Humour

    Directory of Open Access Journals (Sweden)

    Predrag Jovanović

    2010-02-01

    Full Text Available Lactate dehydrogenase (LDH and lactate are some of the hypoxy biochemical parameters. Extracellular activity of this enzyme increases under the condition of oxidative stress, since the cell integrity can be disrupted during the lipid peroxidation process. Subsequently that leads to the increase level of the lactic acid and lactic acid salts. The objective of this investigation is establishing the level of LDH, LDH1 (HBDH and the lactate concentration in aqueous humour in patients with primary open-angle glaucoma.Biochemical analysis have been made by enzymatic-colometric method (lactate and UV-kinetic method (LDH and HBDH in aqueous humour of 30 patients (42 eyes with primary open-angle glaucoma (POAG and 30 patients (40 eyes with cataract (the control group.The increased values of lactate and the activity of LDH and HBDH enzyme in aqueous humour of POAG patients in correlation with the control group are the results not only of oxidative stress but also of hypoxy and the mitochondry oxidative function (p<0,001.The increased activity of the examined biochemical parameters in the aqueous humour of the POAG patients points to the fact that other mechanisms, besides IOP, have a role in glaucoma pathogenesis.

  3. Structural insights into the enzymatic activity and potential substrate promiscuity of human 3-phosphoglycerate dehydrogenase (PHGDH).

    Science.gov (United States)

    Unterlass, Judith E; Wood, Robert J; Baslé, Arnaud; Tucker, Julie; Cano, Céline; Noble, Martin M E; Curtin, Nicola J

    2017-11-28

    Cancer cells reprogram their metabolism and energy production to sustain increased growth, enable metastasis and overcome resistance to cancer treatments. Although primary roles for many metabolic proteins have been identified, some are promiscuous in regards to the reaction they catalyze. To efficiently target these enzymes, a good understanding of their enzymatic function and structure, as well as knowledge regarding any substrate or catalytic promiscuity is required. Here we focus on the characterization of human 3-phosphoglycerate dehydrogenase (PHGDH). PHGDH catalyzes the NAD + -dependent conversion of 3-phosphoglycerate to phosphohydroxypyruvate, which is the first step in the de novo synthesis pathway of serine, a critical amino acid for protein and nucleic acid biosynthesis. We have investigated substrate analogues to assess whether PHGDH might possess other enzymatic roles that could explain its occasional over-expression in cancer, as well as to help with the design of specific inhibitors. We also report the crystal structure of the catalytic subunit of human PHGDH, a dimer, solved with bound cofactor in one monomer and both cofactor and L -tartrate in the second monomer. In vitro enzyme activity measurements show that the catalytic subunit of PHGDH is still active and that PHGDH activity could be significantly inhibited with adenosine 5'-diphosphoribose.

  4. Antimalarial activity of potential inhibitors of Plasmodium falciparum lactate dehydrogenase enzyme selected by docking studies.

    Directory of Open Access Journals (Sweden)

    Julia Penna-Coutinho

    Full Text Available The Plasmodium falciparum lactate dehydrogenase enzyme (PfLDH has been considered as a potential molecular target for antimalarials due to this parasite's dependence on glycolysis for energy production. Because the LDH enzymes found in P. vivax, P. malariae and P. ovale (pLDH all exhibit ∼90% identity to PfLDH, it would be desirable to have new anti-pLDH drugs, particularly ones that are effective against P. falciparum, the most virulent species of human malaria. Our present work used docking studies to select potential inhibitors of pLDH, which were then tested for antimalarial activity against P. falciparum in vitro and P. berghei malaria in mice. A virtual screening in DrugBank for analogs of NADH (an essential cofactor to pLDH and computational studies were undertaken, and the potential binding of the selected compounds to the PfLDH active site was analyzed using Molegro Virtual Docker software. Fifty compounds were selected based on their similarity to NADH. The compounds with the best binding energies (itraconazole, atorvastatin and posaconazole were tested against P. falciparum chloroquine-resistant blood parasites. All three compounds proved to be active in two immunoenzymatic assays performed in parallel using monoclonals specific to PfLDH or a histidine rich protein (HRP2. The IC(50 values for each drug in both tests were similar, were lowest for posaconazole (<5 µM and were 40- and 100-fold less active than chloroquine. The compounds reduced P. berghei parasitemia in treated mice, in comparison to untreated controls; itraconazole was the least active compound. The results of these activity trials confirmed that molecular docking studies are an important strategy for discovering new antimalarial drugs. This approach is more practical and less expensive than discovering novel compounds that require studies on human toxicology, since these compounds are already commercially available and thus approved for human use.

  5. Human xanthine oxidase changes its substrate specificity to aldehyde oxidase type upon mutation of amino acid residues in the active site: roles of active site residues in binding and activation of purine substrate.

    Science.gov (United States)

    Yamaguchi, Yuichiro; Matsumura, Tomohiro; Ichida, Kimiyoshi; Okamoto, Ken; Nishino, Takeshi

    2007-04-01

    Xanthine oxidase (oxidoreductase; XOR) and aldehyde oxidase (AO) are similar in protein structure and prosthetic group composition, but differ in substrate preference. Here we show that mutation of two amino acid residues in the active site of human XOR for purine substrates results in conversion of the substrate preference to AO type. Human XOR and its Glu803-to-valine (E803V) and Arg881-to-methionine (R881M) mutants were expressed in an Escherichia coli system. The E803V mutation almost completely abrogated the activity towards hypoxanthine as a substrate, but very weak activity towards xanthine remained. On the other hand, the R881M mutant lacked activity towards xanthine, but retained slight activity towards hypoxanthine. Both mutants, however, exhibited significant aldehyde oxidase activity. The crystal structure of E803V mutant of human XOR was determined at 2.6 A resolution. The overall molybdopterin domain structure of this mutant closely resembles that of bovine milk XOR; amino acid residues in the active centre pocket are situated at very similar positions and in similar orientations, except that Glu803 was replaced by valine, indicating that the decrease in activity towards purine substrate is not due to large conformational change in the mutant enzyme. Unlike wild-type XOR, the mutants were not subject to time-dependent inhibition by allopurinol.

  6. Further insights into the isoenzyme composition and activity of glutamate dehydrogenase in Arabidopsis thaliana.

    Science.gov (United States)

    Fontaine, Jean-Xavier; Tercé-Laforgue, Thérèse; Bouton, Sophie; Pageau, Karine; Lea, Peter J; Dubois, Frédéric; Hirel, Bertrand

    2013-03-01

    Following the discovery that in Arabidopsis, a third isoenzyme of NADH-dependent glutamate dehydrogenase (GDH) is expressed in the mitochondria of the root companion cells, we have re-examined the GDH isoenzyme composition. By analyzing the NADH-GDH isoenzyme composition of single, double and triple mutants deficient in the expression of the three genes encoding the enzyme, we have found that the α, β and γ polypeptides that comprise the enzyme can be assembled into a complex combination of heterohexamers in roots. Moreover, we observed that when one or two of the three root isoenzymes were missing from the mutants, the remaining isoenzymes compensated for this deficiency. The significance of such complexity is discussed in relation to the metabolic and signaling function of the NADH-GDH enzyme. Although it has been shown that a fourth gene encoding a NADPH-dependent enzyme is present in Arabidopsis, we were not able to detect corresponding enzyme activity, even in the triple mutant totally lacking NADH-GDH activity.

  7. Estrogen modification of human glutamate dehydrogenases is linked to enzyme activation state.

    Science.gov (United States)

    Borompokas, Nikolas; Papachatzaki, Maria-Martha; Kanavouras, Konstantinos; Mastorodemos, Vasileios; Zaganas, Ioannis; Spanaki, Cleanthe; Plaitakis, Andreas

    2010-10-08

    Mammalian glutamate dehydrogenase (GDH) is a housekeeping enzyme central to the metabolism of glutamate. Its activity is potently inhibited by GTP (IC(50) = 0.1-0.3 μM) and thought to be controlled by the need of the cell in ATP. Estrogens are also known to inhibit mammalian GDH, but at relatively high concentrations. Because, in addition to this housekeeping human (h) GDH1, humans have acquired via a duplication event an hGDH2 isoform expressed in human cortical astrocytes, we tested here the interaction of estrogens with the two human isoenzymes. The results showed that, under base-line conditions, diethylstilbestrol potently inhibited hGDH2 (IC(50) = 0.08 ± 0.01 μM) and with ∼18-fold lower affinity hGDH1 (IC(50) = 1.67 ± 0.06 μM; p Structure/function analyses revealed that the evolutionary R443S substitution, which confers low basal activity, was largely responsible for sensitivity of hGDH2 to estrogens. Inhibition of both human GDHs by estrogens was inversely related to their state of activation induced by ADP, with the slope of this correlation being steeper for hGDH2 than for hGDH1. Also, the study of hGDH1 and hGDH2 mutants displaying different states of activation revealed that the affinity of estrogen for these enzymes correlated inversely (R = 0.99; p = 0.0001) with basal catalytic activity. Because astrocytes are known to synthesize estrogens, these hormones, by interacting potently with hGDH2 in its closed state, may contribute to regulation of glutamate metabolism in brain.

  8. Subcellular Characterization of Porcine Oocytes with Different Glucose-6-phosphate Dehydrogenase Activities.

    Science.gov (United States)

    Fu, Bo; Ren, Liang; Liu, Di; Ma, Jian-Zhang; An, Tie-Zhu; Yang, Xiu-Qin; Ma, Hong; Zhang, Dong-Jie; Guo, Zhen-Hua; Guo, Yun-Yun; Zhu, Meng; Bai, Jing

    2015-12-01

    The in vitro maturation (IVM) efficiency of porcine embryos is still low because of poor oocyte quality. Although brilliant cresyl blue positive (BCB+) oocytes with low glucose-6-phosphate dehydrogenase (G6PDH) activity have shown superior quality than BCB negative (-) oocytes with high G6PDH activity, the use of a BCB staining test before IVM is still controversial. This study aimed to shed more light on the subcellular characteristics of porcine oocytes after selection using BCB staining. We assessed germinal vesicle chromatin configuration, cortical granule (CG) migration, mitochondrial distribution, the levels of acetylated lysine 9 of histone H3 (AcH3K9) and nuclear apoptosis features to investigate the correlation between G6PDH activity and these developmentally related features. A pattern of chromatin surrounding the nucleoli was seen in 53.0% of BCB+ oocytes and 77.6% of BCB+ oocytes showed peripherally distributed CGs. After IVM, 48.7% of BCB+ oocytes had a diffused mitochondrial distribution pattern. However, there were no significant differences in the levels of AcH3K9 in the nuclei of blastocysts derived from BCB+ and BCB- oocytes; at the same time, we observed a similar incidence of apoptosis in the BCB+ and control groups. Although this study indicated that G6PDH activity in porcine oocytes was correlated with several subcellular characteristics such as germinal vesicle chromatin configuration, CG migration and mitochondrial distribution, other features such as AcH3K9 level and nuclear apoptotic features were not associated with G6PDH activity and did not validate the BCB staining test. In using this test for selecting porcine oocytes, subcellular characteristics such as the AcH3K9 level and apoptotic nuclear features should also be considered. Adding histone deacetylase inhibitors or apoptosis inhibitors into the culture medium used might improve the efficiency of IVM of BCB+ oocytes.

  9. Subcellular Characterization of Porcine Oocytes with Different Glucose-6-phosphate Dehydrogenase Activities

    Directory of Open Access Journals (Sweden)

    Bo Fu

    2015-12-01

    Full Text Available The in vitro maturation (IVM efficiency of porcine embryos is still low because of poor oocyte quality. Although brilliant cresyl blue positive (BCB+ oocytes with low glucose-6-phosphate dehydrogenase (G6PDH activity have shown superior quality than BCB negative (− oocytes with high G6PDH activity, the use of a BCB staining test before IVM is still controversial. This study aimed to shed more light on the subcellular characteristics of porcine oocytes after selection using BCB staining. We assessed germinal vesicle chromatin configuration, cortical granule (CG migration, mitochondrial distribution, the levels of acetylated lysine 9 of histone H3 (AcH3K9 and nuclear apoptosis features to investigate the correlation between G6PDH activity and these developmentally related features. A pattern of chromatin surrounding the nucleoli was seen in 53.0% of BCB+ oocytes and 77.6% of BCB+ oocytes showed peripherally distributed CGs. After IVM, 48.7% of BCB+ oocytes had a diffused mitochondrial distribution pattern. However, there were no significant differences in the levels of AcH3K9 in the nuclei of blastocysts derived from BCB+ and BCB− oocytes; at the same time, we observed a similar incidence of apoptosis in the BCB+ and control groups. Although this study indicated that G6PDH activity in porcine oocytes was correlated with several subcellular characteristics such as germinal vesicle chromatin configuration, CG migration and mitochondrial distribution, other features such as AcH3K9 level and nuclear apoptotic features were not associated with G6PDH activity and did not validate the BCB staining test. In using this test for selecting porcine oocytes, subcellular characteristics such as the AcH3K9 level and apoptotic nuclear features should also be considered. Adding histone deacetylase inhibitors or apoptosis inhibitors into the culture medium used might improve the efficiency of IVM of BCB+ oocytes.

  10. A deletion of the gene encoding amino aldehyde dehydrogenase enhances the "pandan-like" aroma of winter melon (Benincasa hispida) and is a functional marker for the development of the aroma.

    Science.gov (United States)

    Ruangnam, Saowalak; Wanchana, Samart; Phoka, Nongnat; Saeansuk, Chatree; Mahatheeranont, Sugunya; de Hoop, Simon Jan; Toojinda, Theerayut; Vanavichit, Apichart; Arikit, Siwaret

    2017-12-01

    The gene conferring a "pandan-like" aroma of winter melon was identified. The sequence variation (804-bp deletion) found in the gene was used as the target for functional marker development. Winter melon (Benincasa hispida), a member of the Cucurbitaceae family, is a commonly consumed vegetable in Asian countries that is popular for its nutritional and medicinal value. A "pandan-like" aroma, which is economically important in crops including rice and soybean, is rarely found in most commercial varieties of winter melon, but is present in some landraces. This aroma is a value-added potential trait in breeding winter melon with a higher economic value. In this study, we confirmed that the aroma of winter melon is due to the potent volatile compound 2-acetyl-1-pyrroline (2AP) as previously identified in other plants. Based on an analysis of public transcriptome data, BhAMADH encoding an aminoaldehyde dehydrogenase (AMADH) was identified as a candidate gene conferring aroma of winter melon. A sequence comparison of BhAMADH between the aromatic and non-aromatic accessions revealed an 804-bp deletion encompassing exons 11-13 in the aromatic accession. The deletion caused several premature stop codons and could result in a truncated protein with a length of only 208 amino acids compared with 503 amino acids in the normal protein. A functional marker was successfully developed based on the 804-bp deletion and validated in 237 F 2 progenies. A perfect association of the marker genotypes and aroma phenotypes indicates that BhAMADH is the major gene conferring the aroma. The recently developed functional marker could be efficiently used in breeding programs for the aroma trait in winter melon.

  11. Hypoxia and anoxia effects on alcohol dehydrogenase activity and hemoglobin content in Chironomus riparius Meigen, 1804

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    Valentina Grazioli

    2016-02-01

    Full Text Available The metabolic effects of low oxygen content on alcohol-dehydrogenase (ADH activity and hemoglobin (Hb concentration were investigated in IV-instar larvae of Chironomus riparius (Diptera: Chironomidae from an Italian stream. Two series of short-term (48 h experiments were carried out: exposure to (1 progressive hypoxia (95 to 5% of oxygen saturation and (2 anoxia (at <5% of oxygen saturation. In (1, Hb amount increased with increasing oxygen depletion up to a critical value of oxygenation (about 70% of oxygen saturation. Below this percentage, the Hb amount declined to values comparable with those present in the control. The respiration rate (R remained almost constant at oxygen saturation >50% and decreased significantly only after 48 h of treatment (= <5% of oxygen saturation reaching values <100 mmolO2 gAFDW-1 h-1. ADH activity showed two phases of growth, within the first 14 h and over 18 h of exposure. Overall, we inferred that i Hb might function as short-term oxygen storage, enabling animals to delay the on-set of anaerobiosis; and ii alcoholic fermentation co-occurs for a short time with aerobic respiration, becoming the prevalent metabolic pathway below 5% of oxygen saturation (<1 mg L-1. These considerations were supported also by results from anoxia exposure (2. In such condition, larvae were visibly stressed, becoming immobile after few minutes of incubation, and ADH reached higher values than in the hypoxia treatment (2.03±0.15 UADH mg prot-1. Overall, this study showed a shift from aerobic to anaerobic activity in C. riparius larvae exposed to poorly oxygenated water with an associated alteration of ADH activity and the Hb amount. Such metabolites might be valid candidate biomarkers for the environmental monitoring of running waters.

  12. A new bianthron glycoside as inhibitor of Trypanosoma cruzi glyceraldehyde 3-phosphate dehydrogenase activity

    Energy Technology Data Exchange (ETDEWEB)

    Macedo, Edangelo M.S. de; Silva, Maria G.V. [Universidade Federal do Ceara (UFC), Fortaleza, CE (Brazil). Dept. de Quimica Analitica e Fisico-Quimica; Wiggers, Helton J.; Montanari, Carlos A. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica; Braz-Filho, Raimundo [Universidade Estadual do Norte Fluminense, Campos dos Goytacazes, RJ, (Brazil). Setor de Quimica de Produtos Naturais; Andricopulo, Adriano D. [Universidade de Sao Paulo (USP), Sao Carlos SP (Brazil). Inst. de Fisica

    2009-07-01

    A phytochemical investigation of the ethanolic extract of stalks of Senna martiana Benth. (Leguminoseae), native specie of northeast Brazil, resulted in the isolation and spectroscopic characterization of a new bianthrone glycoside, martianine 1 (10,10'-il-chrysophanol-10-oxi- 10,10'-bi-glucosyl). Its identification was established by HRMS, IR and 2D NMR experiments. The evaluation of martianine trypanocidal activity was carried out against gliceraldehyde 3-phosphate dehydrogenase enzyme from Trypanosoma cruzi. Its inhibitory constant (K{sub i}) is in the low micromolar concentration and it was determined by isothermal titration calorimetry to be 27.3 +-2.47 {mu}mol L{sup -1}. The non-competitive mechanism is asserted to be putative of the mode of action martianine displays against T. cruzi GAPDH. Results show that martianine has a great potential to become new lead molecule by inhibiting this key enzyme and for the development of new drugs against Chagas disease. (author)

  13. Effects of silver nanoparticle on lactate dehydrogenase activity and histological changes of heart tissue in male wistar rats

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    Noushin Naghsh

    2013-03-01

    Full Text Available Background & Objective: The silver nanoparticles are important in many applications of nanoparticles on human health . The toxicity of silver nanoparticles are not well documented yet. The aim of this study was to investigate the effect of silver nanoparticles on lactate dehydrogenase activity and histological changes in heart tissue.   Materials &Methods: In this study, 40 adult male wistar rats of 220±20gr were divided in to five groups including control and four experimental groups. The latter groups were injected intraperitoneally spherical nano silver particles of 50, 100, 200 and 400 ppm respectively for five consecutive days. Then three, eight and twelve days after the last injection, blood samples were collected and lactate dehydrogenase (LDH activity was assayed . Also, tissue samples from the heart muscle were prepared and studied after staining with Hematoxiline-Eosine. Data of LDH activity was analyzed by One way- ANOVA- test and P-value of ≤ 0.05 were considered as significant.   Results : The result showed that different concentrations of silver nanoparticles have no significant effect on the lactate dehydrogenase (p=0.192 . T he histological study of the tissue after exposure to 400 ppm concentration of silver nanoparticles showed the start of primary apoptosis in heart tissue.   Conclusion: The LDH activity was not changed significantly after exposure to different concentration of silver nanoparticles, which shows the safety of these particles on LDH activity.

  14. Purification and characterization of xylitol dehydrogenase with l-arabitol dehydrogenase activity from the newly isolated pentose-fermenting yeast Meyerozyma caribbica 5XY2.

    Science.gov (United States)

    Sukpipat, Wiphat; Komeda, Hidenobu; Prasertsan, Poonsuk; Asano, Yasuhisa

    2017-01-01

    Meyerozyma caribbica strain 5XY2, which was isolated from an alcohol fermentation starter in Thailand, was found to catabolize l-arabinose as well as d-glucose and d-xylose. The highest production amounts of ethanol from d-glucose, xylitol from d-xylose, and l-arabitol from l-arabinose were 0.45 g/g d-glucose, 0.60 g/g d-xylose, and 0.61 g/g l-arabinose with 21.7 g/L ethanol, 20.2 g/L xylitol, and 30.3 g/l l-arabitol, respectively. The enzyme with l-arabitol dehydrogenase (LAD) activity was purified from the strain and found to exhibit broad specificity to polyols, such as xylitol, d-sorbitol, ribitol, and l-arabitol. Xylitol was the preferred substrate with K m =16.1 mM and k cat /K m =67.0 min -1 mM -1 , while l-arabitol was also a substrate for the enzyme with K m =31.1 mM and k cat /K m =6.5 min -1  mM -1 . Therefore, this enzyme from M. caribbica was named xylitol dehydrogenase (McXDH). McXDH had an optimum temperature and pH at 40°C and 9.5, respectively. The McXDH gene included a coding sequence of 1086 bp encoding a putative 362 amino acid protein of 39 kDa with an apparent homopentamer structure. Native McXDH and recombinant McXDH exhibited relative activities toward l-arabitol of approximately 20% that toward xylitol, suggesting the applicability of this enzyme with the functions of XDH and LAD to the development of pentose-fermenting Saccharomyces cerevisiae. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  15. Structural basis for leucine-induced allosteric activation of glutamate dehydrogenase.

    Science.gov (United States)

    Tomita, Takeo; Kuzuyama, Tomohisa; Nishiyama, Makoto

    2011-10-28

    Glutamate dehydrogenase (GDH) catalyzes reversible conversion between glutamate and 2-oxoglutarate using NAD(P)(H) as a coenzyme. Although mammalian GDH is regulated by GTP through the antenna domain, little is known about the mechanism of allosteric activation by leucine. An extremely thermophilic bacterium, Thermus thermophilus, possesses GDH with a unique subunit configuration composed of two different subunits, GdhA (regulatory subunit) and GdhB (catalytic subunit). T. thermophilus GDH is unique in that the enzyme is subject to allosteric activation by leucine. To elucidate the structural basis for leucine-induced allosteric activation of GDH, we determined the crystal structures of the GdhB-Glu and GdhA-GdhB-Leu complexes at 2.1 and 2.6 Å resolution, respectively. The GdhB-Glu complex is a hexamer that binds 12 glutamate molecules: six molecules are bound at the substrate-binding sites, and the remaining six are bound at subunit interfaces, each composed of three subunits. The GdhA-GdhB-Leu complex is crystallized as a heterohexamer composed of four GdhA subunits and two GdhB subunits. In this complex, six leucine molecules are bound at subunit interfaces identified as glutamate-binding sites in the GdhB-Glu complex. Consistent with the structure, replacement of the amino acid residues of T. thermophilus GDH responsible for leucine binding made T. thermophilus GDH insensitive to leucine. Equivalent amino acid replacement caused a similar loss of sensitivity to leucine in human GDH2, suggesting that human GDH2 also uses the same allosteric site for regulation by leucine.

  16. Enhanced pyruvate dehydrogenase activity improves cardiac outcomes in a murine model of cardiac arrest.

    Directory of Open Access Journals (Sweden)

    Lin Piao

    Full Text Available Post-ischemic changes in cellular metabolism alter myocardial and neurological function. Pyruvate dehydrogenase (PDH, the limiting step in mitochondrial glucose oxidation, is inhibited by increased expression of PDH kinase (PDK during ischemia/reperfusion injury. This results in decreased utilization of glucose to generate cellular ATP. Post-cardiac arrest (CA hypothermia improves outcomes and alters metabolism, but its influence on PDH and PDK activity following CA are unknown. We hypothesized that therapeutic hypothermia (TH following CA is associated with the inhibition of PDK activity and increased PDH activity. We further hypothesized that an inhibitor of PDK activity, dichloroacetate (DCA, would improve PDH activity and post-CA outcomes.Anesthetized and ventilated adult female C57BL/6 wild-type mice underwent a 12-minute KCl-induced CA followed by cardiopulmonary resuscitation. Compared to normothermic (37°C CA controls, administering TH (30°C improved overall survival (72-hour survival rate: 62.5% vs. 28.6%, P<0.001, post-resuscitation myocardial function (ejection fraction: 50.9±3.1% vs. 27.2±2.0%, P<0.001; aorta systolic pressure: 132.7±7.3 vs. 72.3±3.0 mmHg, P<0.001, and neurological scores at 72-hour post CA (9.5±1.3 vs. 5.4±1.3, P<0.05. In both heart and brain, CA increased lactate concentrations (1.9-fold and 3.1-fold increase, respectively, P<0.01, decreased PDH enzyme activity (24% and 50% reduction, respectively, P<0.01, and increased PDK protein expressions (1.2-fold and 1.9-fold, respectively, P<0.01. In contrast, post-CA treatment with TH normalized lactate concentrations (P<0.01 and P<0.05 and PDK expressions (P<0.001 and P<0.05, while increasing PDH activity (P<0.01 and P<0.01 in both the heart and brain. Additionally, treatment with DCA (0.2 mg/g body weight 30 min prior to CA improved both myocardial hemodynamics 2 hours post-CA (aortic systolic pressure: 123±3 vs. 96±4 mmHg, P<0.001 and 72-hour survival rates

  17. Dose-dependent regulation of microbial activity on sinking particles by polyunsaturated aldehydes: Implications for the carbon cycle.

    Science.gov (United States)

    Edwards, Bethanie R; Bidle, Kay D; Van Mooy, Benjamin A S

    2015-05-12

    Diatoms and other phytoplankton play a crucial role in the global carbon cycle, fixing CO2 into organic carbon, which may then be exported to depth via sinking particles. The molecular diversity of this organic carbon is vast and many highly bioactive molecules have been identified. Polyunsaturated aldehydes (PUAs) are bioactive on various levels of the marine food web, and yet the potential for these molecules to affect the fate of organic carbon produced by diatoms remains an open question. In this study, the effects of PUAs on the natural microbial assemblages associated with sinking particles were investigated. Sinking particles were collected from 150 m in the water column and exposed to varying concentrations of PUAs in dark incubations over 24 h. PUA doses ranging from 1 to 10 µM stimulated respiration, organic matter hydrolysis, and cell growth by bacteria associated with sinking particles. PUA dosages near 100 µM appeared to be toxic, resulting in decreased bacterial cell abundance and metabolism, as well as pronounced shifts in bacterial community composition. Sinking particles were hot spots for PUA production that contained concentrations within the stimulatory micromolar range in contrast to previously reported picomolar concentrations of these compounds in bulk seawater. This suggests PUAs produced in situ stimulate the remineralization of phytoplankton-derived sinking organic matter, decreasing carbon export efficiency, and shoaling the average depths of nutrient regeneration. Our results are consistent with a "bioactivity hypothesis" for explaining variations in carbon export efficiency in the oceans.

  18. A substrate in pieces: allosteric activation of glycerol 3-phosphate dehydrogenase (NAD+) by phosphite dianion.

    Science.gov (United States)

    Tsang, Wing-Yin; Amyes, Tina L; Richard, John P

    2008-04-22

    The ratio of the second-order rate constants for reduction of dihydroxyacetone phosphate (DHAP) and of the neutral truncated substrate glycolaldehyde (GLY) by glycerol 3-phosphate dehydrogenase (NAD (+), GPDH) saturated with NADH is (1.0 x 10 (6) M (-1) s (-1))/(8.7 x 10 (-3) M (-1) s (-1)) = 1.1 x 10 (8), which was used to calculate an intrinsic phosphate binding energy of at least 11.0 kcal/mol. Phosphite dianion binds very weakly to GPDH ( K d > 0.1 M), but the bound dianion strongly activates GLY toward enzyme-catalyzed reduction by NADH. Thus, the large intrinsic phosphite binding energy is expressed only at the transition state for the GPDH-catalyzed reaction. The ratio of rate constants for the phosphite-activated and the unactivated GPDH-catalyzed reduction of GLY by NADH is (4300 M (-2) s (-1))/(8.7 x 10 (-3) M (-1) s (-1)) = 5 x 10 (5) M (-1), which was used to calculate an intrinsic phosphite binding energy of -7.7 kcal/mol for the association of phosphite dianion with the transition state complex for the GPDH-catalyzed reduction of GLY. Phosphite dianion has now been shown to activate bound substrates for enzyme-catalyzed proton transfer, decarboxylation, hydride transfer, and phosphoryl transfer reactions. Structural data provide strong evidence that enzymic activation by the binding of phosphite dianion occurs at a modular active site featuring (1) a binding pocket complementary to the reactive substrate fragment which contains all the active site residues needed to catalyze the reaction of the substrate piece or of the whole substrate and (2) a phosphate/phosphite dianion binding pocket that is completed by the movement of flexible protein loop(s) to surround the nonreacting oxydianion. We propose that loop motion and associated protein conformational changes that accompany the binding of phosphite dianion and/or phosphodianion substrates lead to encapsulation of the substrate and/or its pieces in the protein interior, and to placement of the active

  19. Inhibition of Calcineurin or IMP Dehydrogenase Exerts Moderate to Potent Antiviral Activity against Norovirus Replication.

    Science.gov (United States)

    Dang, Wen; Yin, Yuebang; Wang, Yijin; Wang, Wenshi; Su, Junhong; Sprengers, Dave; van der Laan, Luc J W; Felczak, Krzysztof; Pankiewicz, Krzysztof W; Chang, Kyeong-Ok; Koopmans, Marion P G; Metselaar, Herold J; Peppelenbosch, Maikel P; Pan, Qiuwei

    2017-11-01

    Norovirus is a major cause of acute gastroenteritis worldwide and has emerged as an important issue of chronic infection in transplantation patients. Since no approved antiviral is available, we evaluated the effects of different immunosuppressants and ribavirin on norovirus and explored their mechanisms of action by using a human norovirus (HuNV) replicon-harboring model and a surrogate murine norovirus (MNV) infectious model. The roles of the corresponding drug targets were investigated by gain- or loss-of-function approaches. We found that the calcineurin inhibitors cyclosporine (CsA) and tacrolimus (FK506) moderately inhibited HuNV replication. Gene silencing of their cellular targets, cyclophilin A, FKBP12, and calcineurin, significantly inhibited HuNV replication. A low concentration, therapeutically speaking, of mycophenolic acid (MPA), an uncompetitive IMP dehydrogenase (IMPDH) inhibitor, potently and rapidly inhibited norovirus replication and ultimately cleared HuNV replicons without inducible resistance following long-term drug exposure. Knockdown of the MPA cellular targets IMPDH1 and IMPDH2 suppressed HuNV replication. Consistent with the nucleotide-synthesizing function of IMPDH, exogenous guanosine counteracted the antinorovirus effects of MPA. Furthermore, the competitive IMPDH inhibitor ribavirin efficiently inhibited norovirus and resulted in an additive effect when combined with immunosuppressants. The results from this study demonstrate that calcineurin phosphatase activity and IMPDH guanine synthase activity are crucial in sustaining norovirus infection; thus, they can be therapeutically targeted. Our results suggest that MPA shall be preferentially considered immunosuppressive medication for transplantation patients at risk of norovirus infection, whereas ribavirin represents as a potential antiviral for both immunocompromised and immunocompetent patients with norovirus gastroenteritis. Copyright © 2017 American Society for Microbiology.

  20. H3 clipping activity of glutamate dehydrogenase is regulated by stefin B and chromatin structure.

    Science.gov (United States)

    Mandal, Papita; Chauhan, Sakshi; Tomar, Raghuvir S

    2014-12-01

    Glutamate dehydrogenase has been recently identified as a tissue-specific histone H3-specific clipping enzyme. We have previously shown that it cleaves free as well as chromatin-bound histone H3. However, the physiological significance of this enzyme is still not clear. The present study aimed to improve our understanding of its significance in vivo. Using biochemical and cell biological approaches, we show that glutamate dehydrogenase is primarily associated with euchromatin, and it re-localizes from the nuclear periphery to the nucleolus upon DNA damage. The cysteine protease inhibitor stefin B regulates the H3 clipping activity of the enzyme. Chromatin structure and certain histone modifications influence H3 clipping activity. Interestingly, we also observed that an in vivo truncated form of H3 lacks H3K56 acetylation, which is a code for the DNA damage response. Together, these results suggest that glutamate dehydrogenase is a euchromatin-associated enzyme, and its H3 clipping activity is regulated by chromatin structure, histone modifications and an in vivo inhibitor. In response to DNA damage, it re-localizes to the nuclei, and hence may be involved in regulation of gene expression in vivo. © 2014 FEBS.

  1. Redox Balance in Lactobacillus reuteri DSM20016: Roles of Iron-Dependent Alcohol Dehydrogenases in Glucose/ Glycerol Metabolism.

    Science.gov (United States)

    Chen, Lu; Bromberger, Paul David; Nieuwenhuiys, Gavin; Hatti-Kaul, Rajni

    2016-01-01

    Lactobacillus reuteri, a heterofermentative bacterium, metabolizes glycerol via a Pdu (propanediol-utilization) pathway involving dehydration to 3-hydroxypropionaldehyde (3-HPA) followed by reduction to 1,3-propandiol (1,3-PDO) with concomitant generation of an oxidized cofactor, NAD+ that is utilized to maintain cofactor balance required for glucose metabolism and even for oxidation of 3-HPA by a Pdu oxidative branch to 3-hydroxypropionic acid (3-HP). The Pdu pathway is operative inside Pdu microcompartment that encapsulates different enzymes and cofactors involved in metabolizing glycerol or 1,2-propanediol, and protects the cells from the toxic effect of the aldehyde intermediate. Since L. reuteri excretes high amounts of 3-HPA outside the microcompartment, the organism is likely to have alternative alcohol dehydrogenase(s) in the cytoplasm for transformation of the aldehyde. In this study, diversity of alcohol dehydrogenases in Lactobacillus species was investigated with a focus on L. reuteri. Nine ADH enzymes were found in L. reuteri DSM20016, out of which 3 (PduQ, ADH6 and ADH7) belong to the group of iron-dependent enzymes that are known to transform aldehydes/ketones to alcohols. L. reuteri mutants were generated in which the three ADHs were deleted individually. The lagging growth phenotype of these deletion mutants revealed that limited NAD+/NADH recycling could be restricting their growth in the absence of ADHs. Notably, it was demonstrated that PduQ is more active in generating NAD+ during glycerol metabolism within the microcompartment by resting cells, while ADH7 functions to balance NAD+/NADH by converting 3-HPA to 1,3-PDO outside the microcompartment in the growing cells. Moreover, evaluation of ADH6 deletion mutant showed strong decrease in ethanol level, supporting the role of this bifuctional alcohol/aldehyde dehydrogenase in ethanol production. To the best of our knowledge, this is the first report revealing both internal and external recycling

  2. Lactate dehydrogenase activity of rat epididymis and spermatozoa: Effect of constant light

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    RH Ponce

    2009-12-01

    Full Text Available During its passage through the epididymis, the gamete undergoes a process of “maturation” leading to the acquisition of its fertilizing ability. The epididymis displays regional variations in the morphology and metabolic properties of its epithelium which are relevant for the progressive development of mature sperm characteristics. The epididymis has spontaneous peristaltic contractions and receives sympathetic innervation that is modulated by melatonin, a hormone synthesized and released by the pineal gland. Constant lighting disrupts melatonin synthesis and secretion. We have studied the effect of constant light on lactate dehydrogenase (LDH; EC 1.1.1.27 and its isozyme C4 activities and protein content in whole epididymis, epididymal tissue and in spermatozoa from caput and cauda segments. Animals were exposed from birth to an illumination schedule of 14 h light: 10 h dark (group L:D. At 60 days of age one group of animals was submitted to constant light over 50 days (group L:L. In order to test the fertilizing ability, the rats of each group were mated with soliciting estrous females. The percentage of pregnancies in females mated with males maintained in L:L was remarkably lower than those in females mated with males maintained in the L:D photoperiod (44% and 88% respectively. Constant light increased protein concentration and LDH activity in caput as well as in cauda of total epididymis. On the contrary, in epididymal tissue, the protein content decreased in both epididymal sections compared with controls. When enzymatic activity was expressed in Units per spermatozoa, constant light induced a significant reduction of total LDH and LDHC4 in caput and cauda spermatozoa while LDH activity of epididymal tissue was not affected. In spite of the decrease in LDH per sperm cell when rats were exposed to constant light, in total epididymis (epididymis tissue plus sperm cells content and in spermatozoa, values of enzyme activities expressed per

  3. Exploring the clonal evolution of CD133/aldehyde-dehydrogenase-1 (ALDH1)-positive cancer stem-like cells from primary to recurrent high-grade serous ovarian cancer (HGSOC). A study of the Ovarian Cancer Therapy-Innovative Models Prolong Survival (OCTIPS) Consortium.

    Science.gov (United States)

    Ruscito, Ilary; Cacsire Castillo-Tong, Dan; Vergote, Ignace; Ignat, Iulia; Stanske, Mandy; Vanderstichele, Adriaan; Ganapathi, Ram N; Glajzer, Jacek; Kulbe, Hagen; Trillsch, Fabian; Mustea, Alexander; Kreuzinger, Caroline; Benedetti Panici, Pierluigi; Gourley, Charlie; Gabra, Hani; Kessler, Mirjana; Sehouli, Jalid; Darb-Esfahani, Silvia; Braicu, Elena Ioana

    2017-07-01

    High-grade serous ovarian cancer (HGSOC) causes 80% of all ovarian cancer (OC) deaths. In this setting, the role of cancer stem-like cells (CSCs) is still unclear. In particular, the evolution of CSC biomarkers from primary (pOC) to recurrent (rOC) HGSOCs is unknown. Aim of this study was to investigate changes in CD133 and aldehyde dehydrogenase-1 (ALDH1) CSC biomarker expression in pOC and rOC HGSOCs. Two-hundred and twenty-four pOC and rOC intrapatient paired tissue samples derived from 112 HGSOC patients were evaluated for CD133 and ALDH1 expression using immunohistochemistry (IHC); pOCs and rOCs were compared for CD133 and/or ALDH1 levels. Expression profiles were also correlated with patients' clinicopathological and survival data. Some 49.1% of the patient population (55/112) and 37.5% (42/112) pOCs were CD133+ and ALDH1+ respectively. CD133+ and ALDH1+ samples were detected in 33.9% (38/112) and 36.6% (41/112) rOCs. CD133/ALDH1 coexpression was observed in 23.2% (26/112) and 15.2% (17/112) of pOCs and rOCs respectively. Pairwise analysis showed a significant shift of CD133 staining from higher (pOCs) to lower expression levels (rOCs) (p cancer cells, providing also a first evidence that there is no correlation between CSCs and BRCA status. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Electrochemical Studies of the Inhibition and Activation Effects of Al (III on the Activity of Bovine Liver Glutamate Dehydrogenase

    Directory of Open Access Journals (Sweden)

    Shuping Bi

    2005-04-01

    Full Text Available Since the study of Al3+ ion on the enzyme activity by using of electrochemical techniques was rarely found in available literatures, the differential-pulse polarography (DPP technique was applied to study the effects of Al3+ ion on the glutamate dehydrogenase (GDH activity in the catalytical reaction of α-KG +NADH+NH4 + ⇔ L-Glu+NAD++H2O by monitoring the DPP reduction current of NAD+. At the plant and animal physiologically relevant pH values (pH=6.5 and 7.5, the GDH enzyme activities were strongly depended on the concentrations of the metal ion in the assay mixture solutions. In the lower Al (III concentration solutions (80μM, the inhibition effects of Al (III were shown again. The cyclic voltammetry of NAD+ and NAD+-GDH in the presence of Al (III can help to explain some biological phenomena. According to the differential-pulse polarography and cyclic voltammetry experiments, the present research confirmed that the electrochemical technique is a convenient and reliable sensor for accurate determination of enzyme activity in biological and environmental samples.

  5. Active site of Zn2+-dependent sn-glycerol-1-phosphate dehydrogenase from Aeropyrum pernix K1

    Directory of Open Access Journals (Sweden)

    Jin-Suk Han

    2005-01-01

    Full Text Available The enzyme sn-glycerol-1-phosphate dehydrogenase (Gro1PDH, EC 1.1.1.261 is key to the formation of the enantiomeric configuration of the glycerophosphate backbone (sn-glycerol-1-phosphate of archaeal ether lipids. This enzyme catalyzes the reversible conversion between dihydroxyacetone phosphate and glycerol-1-phosphate. To date, no information about the active site and catalytic mechanism of this enzyme has been reported. Using the sequence and structural information for glycerol dehydrogenase, we constructed six mutants (D144N, D144A, D191N, H271A, H287A and D191N/H271A of Gro1PDH from Aeropyrum pernix K1 and examined their characteristics to clarify the active site of this enzyme. The enzyme was found to be a zinc-dependent metalloenzyme, containing one zinc ion for every monomer protein that was essential for activity. Site-directed mutagenesis of D144 increased the activity of the enzyme. Mutants D144N and D144A exhibited low affinity for the substrates and higher activity than the wild type, but their affinity for the zinc ion was the same as that of the wild type. Mutants D191N, H271A and H287A had a low affinity for the zinc ion and a low activity compared with the wild type. The double mutation, D191N/ H271A, had no enzyme activity and bound no zinc. From these results, it was clarified that residues D191, H271 and H287 participate in the catalytic activity of the enzyme by binding the zinc ion, and that D144 has an effect on substrate binding. The structure of the active site of Gro1PDH from A. pernix K1 seems to be similar to that of glycerol dehydrogenase, despite the differences in substrate specificity and biological role.

  6. A sensitive cytochemical staining method for glucose-6-phosphate dehydrogenase activity in individual erythrocytes. II. Further improvements of the staining procedure and some observations with glucose-6-phosphate dehydrogenase deficiency

    NARCIS (Netherlands)

    van Noorden, C. J.; Vogels, I. M.

    1985-01-01

    A cytochemical method for staining glucose-6-phosphate dehydrogenase (G6PD) activity in individual erythrocytes as reported previously has been optimized further by the incorporation of a number of technical improvements. Analysis of the enzyme content in erythrocytes of normal individuals as well

  7. Regulation of the activity of lactate dehydrogenases from four lactic acid bacteria

    NARCIS (Netherlands)

    Feldman-Salit, A.; Hering, S.; Messiha, H.L.; Veith, N.; Cojocaru, V.; Sieg, A.; Westerhoff, H.V.; Kreikemeyer, B.; Wade, R.C.; Fiedler, T.

    2013-01-01

    Despite high similarity in sequence and catalytic properties, the l-lactate dehydrogenases (LDHs) in lactic acid bacteria (LAB) display differences in their regulation that may arise from their adaptation to different habitats. We combined experimental and computational approaches to investigate the

  8. Properties of an NAD(H)-containing methanol dehydrogenase and its activator protein from Bacillus methanolicus

    NARCIS (Netherlands)

    Arfman, Nico; Hektor, Harm J.; Bystrykh, Leonid V.; Govorukhina, Natalya I.; Dijkhuizen, Lubbert; Frank, Johannes

    1997-01-01

    Oxidation of C1-C4 primary alcohols in thermotolerant Bacillus methanolicus strains is catalyzed by an NAD-dependent methanol dehydrogenase (MDH), composed of ten identical 43000-Mr subunits. Each MDH subunit contains a tightly, but non-covalently, bound NAD(H) molecule, in addition to 1 Zn2+ and

  9. Redox Status, Procoagulant Activity, and Metabolome of Fresh Frozen Plasma in Glucose 6-Phosphate Dehydrogenase Deficiency

    Directory of Open Access Journals (Sweden)

    Vassilis L. Tzounakas

    2018-02-01

    Full Text Available ObjectiveTransfusion of fresh frozen plasma (FFP helps in maintaining the coagulation parameters in patients with acquired multiple coagulation factor deficiencies and severe bleeding. However, along with coagulation factors and procoagulant extracellular vesicles (EVs, numerous bioactive and probably donor-related factors (metabolites, oxidized components, etc. are also carried to the recipient. The X-linked glucose 6-phosphate dehydrogenase deficiency (G6PD−, the most common human enzyme genetic defect, mainly affects males. By undermining the redox metabolism, the G6PD− cells are susceptible to the deleterious effects of oxidants. Considering the preferential transfusion of FFP from male donors, this study aimed at the assessment of FFP units derived from G6PD− males compared with control, to show whether they are comparable at physiological, metabolic and redox homeostasis levels.MethodsThe quality of n = 12 G6PD− and control FFP units was tested after 12 months of storage, by using hemolysis, redox, and procoagulant activity-targeted biochemical assays, flow cytometry for EV enumeration and phenotyping, untargeted metabolomics, in addition to statistical and bioinformatics tools.ResultsHigher procoagulant activity, phosphatidylserine positive EVs, RBC-vesiculation, and antioxidant capacity but lower oxidative modifications in lipids and proteins were detected in G6PD− FFP compared with controls. The FFP EVs varied in number, cell origin, and lipid/protein composition. Pathway analysis highlighted the riboflavin, purine, and glycerolipid/glycerophospholipid metabolisms as the most altered pathways with high impact in G6PD−. Multivariate and univariate analysis of FFP metabolomes showed excess of diacylglycerols, glycerophosphoinositol, aconitate, and ornithine but a deficiency in riboflavin, flavin mononucleotide, adenine, and arginine, among others, levels in G6PD− FFPs compared with control.ConclusionOur results point

  10. Electrophilic aldehyde products of lipid peroxidation selectively adduct to heat shock protein 90 and arylsulfatase A in stallion spermatozoa.

    Science.gov (United States)

    Hall, Sally E; Aitken, R John; Nixon, Brett; Smith, Nathan D; Gibb, Zamira

    2017-01-01

    Oxidative stress is a major determinant of mammalian sperm function stimulating lipid peroxidation cascades that culminate in the generation of potentially cytotoxic aldehydes. The aim of this study was to assess the impact of such aldehydes on the functionality of stallion spermatozoa. The impact of exposure to exogenous acrolein (ACR) and 4-hydroxynonenal (4HNE) was manifested in a highly significant dose- and time-dependent increase in mitochondrial reactive oxygen species (ROS), total cellular ROS, a decrease in sperm motility, and a time-dependent increase in lipid peroxidation. Notably, low doses of ACR and 4HNE also caused a significant decrease in zona binding. In contrast, exogenous malondialdehyde, a commonly used marker of oxidative stress, had little impact on the various sperm parameters assessed. In accounting for the negative physiological impact of ACR and 4HNE, it was noted that both aldehydes readily adducted to sperm proteins located predominantly within the head, proximal centriole, and tail. The detoxifying activity of mitochondrial aldehyde dehydrogenase 2 appeared responsible for a lack of adduction in the midpiece; however, this activity was overwhelmed by 24 h of electrophilic aldehyde exposure. Sequencing of the dominant proteins targeted for ACR and 4HNE covalent modification identified heat shock protein 90 alpha (cytosolic) class A member 1 and arylsulfatase A, respectively. These collective findings may prove useful in the identification of diagnostic biomarkers of stallion fertility and resolving the mechanistic basis of sperm dysfunction in this species. © The Authors 2016. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please journals.permissions@oup.com.

  11. Lactic acid production by Rhizopus oryzae transformants with modified lactate dehydrogenase activity.

    Science.gov (United States)

    Skory, C D

    2004-04-01

    Rhizopus oryzae is capable of producing high levels of lactic acid by the fermentation of glucose. Yields typically vary over 60-80%, with the remaining glucose diverted primarily into ethanol fermentation. The goal of this work was to increase lactate dehydrogenase (LDH) activity, so lactic acid fermentation could more effectively compete for available pyruvate. Three different constructs, pLdhA71X, pLdhA48XI, and pLdhA89VII, containing various lengths of the ldhA gene fragment, were transformed into R. oryzae. This fungus rarely integrates DNA used for transformation, but instead relies on extra-chromosomal replication in a high-copy number. Plasmid pLdhA48XI was linearized prior to transformation in order to facilitate integration into the pyrG gene used for selection. Isolates transformed with ldhA containing plasmid were compared with both the wild-type parent strain and the auxotrophic recipient strain containing vector only. All isolates transformed with pLdhA71X or pLdhA48XI had multiple copies of the ldhA gene that resulted in ldhA transcript accumulation, LDH specific activity, and lactic acid production higher than the controls. Integration of plasmid pLdhA48XI increased the stability of the strain, but did not seem to offer any benefit for increasing lactic acid production. Since lactic acid fermentation competes with ethanol and fumaric acid production, it was not unexpected that increased lactic acid production was always concomitant with decreased ethanol and fumaric acid. Plasmid pLdhA71X, containing a large ldhA fragment (6.1 kb), routinely yielded higher levels of lactic acid than the smaller region (3.3 kb) used to construct plasmid pLdhA48XI. The greatest levels of ldhA transcript and enzyme production occurred with isolates transformed with plasmid pLdhA89VII. However, these transformants always produced less lactic acid and higher amounts of ethanol, fumaric, and glycerol compared with the control.

  12. Characterization of retinaldehyde dehydrogenase 3

    OpenAIRE

    Graham, Caroline E.; Brocklehurst, Keith; Pickersgill, Richard W.; Warren, Martin J.

    2006-01-01

    RALDH3 (retinal dehydrogenase 3) was characterized by kinetic and binding studies, protein engineering, homology modelling, ligand docking and electrostatic-potential calculations. The major recognition determinant of an RALDH3 substrate was shown to be an eight-carbon chain bonded to the aldehyde group whose kinetic influence (kcat/Km at pH 8.5) decreases when shortened or lengthened. Surprisingly, the β-ionone ring of all-trans-retinal is not a major recognition site. The dissociation const...

  13. Succinate Dehydrogenase Activity Assay in situ with Blue Tetrazolium Salt in Crabtree-Positive Saccharomyces cerevisiae Strain

    Directory of Open Access Journals (Sweden)

    Joanna Berlowska

    2008-01-01

    Full Text Available A spectrophotometric method for determining succinate dehydrogenase (SDH activity assay in azide-sensitive yeast Saccharomyces cerevisiae has been developed. The permeabilization of yeast cells by 0.05 % digitonin permitted to study yeast enzymatic activity in situ. The reduction of blue tetrazolium salt (BT to blue tetrazolium formazan (BTf was conducted in the presence of phenazine methosulphate (PMS as an exogenous electron carrier, and sodium azide (SA as an inhibitor of cytochrome oxidase (Cyt pathway. Various factors such as type of substrate, BT concentration, cell number, temperature and time of incubation, and different Cyt pathway blockers were optimized. In earlier studies, dimethyl sulfoxide (DMSO had been selected as the best solvent for extraction of BTf from yeast cells. The linear correlation between permeabilized yeast cell density and amount of formed formazan was evidenced in the range from 9·10^7 to 5·10^8 cells per sample solution. Below the yeast cell concentration of 10^7 the absorbance values were too low to detect formazans with good precision. This standarized procedure allows the estimation of SDH activity in whole cells, depending on vitality level of yeast populations. Significant increases of succinate dehydrogenase activities were observed in sequential passages as the result of the increase of activity of the strain and adaptation to cultivation conditions.

  14. [Malate dehydrogenase and lactate dehydrogenase in trematodes and turbellarians].

    Science.gov (United States)

    Vykhrestiuk, N P; Burenina, E A; Iarygina, G V

    1986-01-01

    Studies have been made on the activity and properties of malate and lactate dehydrogenases from the cattle rumen trematodes Eurytrema pancreaticum, Calicophoron ijimai and the turbellarian Phagocata sibirica which has a common free-living ancestor with the trematodes. All the species studied have a highly active malate dehydrogenase, its activity in the reaction of reducing oxaloacetate being 6-14 times higher than in the reaction of malate oxidation. The affinity of malate dehydrogenase to oxaloacetate was found to be higher than that to malate. The activity of lactate dehydrogenase (reducing the pyruvate) was lower than the activity of malate dehydrogenase, the difference being 50 times for C. ijimai, 4 times for E. pancreaticum and 10 times for P. sibirica.

  15. Aldehyde Dehydrogenase 1 and Raf Kinase Inhibitor Protein ...

    African Journals Online (AJOL)

    Immunohistochemical and Western blotting techniques were employed to study the expression profiles of ALDH1 and RKIP. The specificity of Sox2 that determines cancer stem cells served as control to validate ALDH1 and RKIP expressions. Results: Histological data helped to differentiate low from high grade cervical ...

  16. Engineering a thermostable highly active glucose 6-phosphate dehydrogenase and its application to hydrogen production in vitro.

    Science.gov (United States)

    Huang, Rui; Chen, Hui; Zhou, Wei; Ma, Chunling; Zhang, Y-H Percival

    2018-04-01

    Glucose 6-phosphate dehydrogenase (G6PDH) is one of the most important dehydrogenases responsible for generating reduced NADPH for anabolism and is also the rate-limiting enzyme in the Entner-Doudoroff pathway. For in vitro biocatalysis, G6PDH must possess both high activity and good thermostability due to requirements of efficient use and low expense of biocatalyst. Here, we used directed evolution to improve thermostability of the highly active G6PDH from Zymomonas mobilis. Four generations of random mutagenesis and Petri-dish-based double-layer screening evolved the thermolabile wild-type enzyme to the thermostable mutant Mut 4-1, which showed a more than 124-fold increase in half-life time (t 1/2 ) at 60 °C, a 3.4 °C increase in melting temperature (T m ), and a 5 °C increase in optimal temperature (T opt ), without compromising the specific activity. In addition, the thermostable mutant was conducted to generate hydrogen from maltodextrin via in vitro synthetic biosystems (ivSB), gaining a more than 8-fold improvement of productivity rate with 76% of theoretical yield at 60 °C. Thus, the engineered G6PDH has been shown to effectively regenerate NADPH at high temperatures and will be applicable for NAD(P)H regeneration in numerous in vitro biocatalysis applications.

  17. Glutamate 270 plays an essential role in K(+)-activation and domain closure of Thermus thermophilus isopropylmalate dehydrogenase.

    Science.gov (United States)

    Gráczer, Éva; Palló, Anna; Oláh, Julianna; Szimler, Tamás; Konarev, Petr V; Svergun, Dmitri I; Merli, Angelo; Závodszky, Péter; Weiss, Manfred S; Vas, Mária

    2015-01-16

    The mutant E270A of Thermus thermophilus 3-isopropylmalate dehydrogenase exhibits largely reduced (∼1%) catalytic activity and negligible activation by K(+) compared to the wild-type enzyme. A 3-4 kcal/mol increase in the activation energy of the catalysed reaction upon this mutation could also be predicted by QM/MM calculations. In the X-ray structure of the E270A mutant a water molecule was observed to take the place of K(+). SAXS and FRET experiments revealed the essential role of E270 in stabilisation of the active domain-closed conformation of the enzyme. In addition, E270 seems to position K(+) into close proximity of the nicotinamide ring of NAD(+) and the electron-withdrawing effect of K(+) may help to polarise the aromatic ring in order to aid the hydride-transfer. Copyright © 2014. Published by Elsevier B.V.

  18. NAD(P-DEPENDENT DEHYDROGENASE ACTIVITY IN PERIPHERAL BLOOD LYMPHOCYTES OF INFANTS WITH ENLARGEMENT OF PHARYNGEAL TONSILS

    Directory of Open Access Journals (Sweden)

    L. M. Kurtasova

    2014-01-01

    Full Text Available We have observed and examined 57 children 1 to 3 years old diagnosed with enlargement of pharyngeal tonsils. A control group was presented by 35 healthy children. Bioluminescence technique was applied for studying NAD(P-dependent dehydrogenase activity in peripheral blood lymphocytes. Activation of aerobic respiration and increasing activity of pentose phosphate cycle-dependent plastic processes were registered in blood lymphocytes of children with hypertrophic pharyngeal tonsils; along with decreased function of malate-aspartate shunt in energy metabolism of the cells, diminished anaerobic reaction of NADHdependent LDH, lower interaction between Krebs cycle and reactions of amino acid metabolism, and reduced activity of glutathione reductase.

  19. Formyl-d aromatic aldehydes

    International Nuclear Information System (INIS)

    Chancellor, T.; Quill, M.; Bergbreiter, D.E.; Newcomb, M.

    1978-01-01

    A simple exchange reaction for preparation of aldehydes labeled with deuterium at the formyl carbon is described. It can be successfully accomplished with several aromatic aldehydes, a catalytic or stoichiometric amount of either potassium cyanide or a thiazolium salt, a weak Lewis base, and deuterium oxide as the deuterium source

  20. First general methods toward aldehyde enolphosphates.

    Science.gov (United States)

    Barthes, Nicolas; Grison, Claude

    2012-02-01

    We herein report two innovative methods toward aldehyde enolphosphates and the first saccharidic aldehyde enolphosphates. Aldehyde enolphosphate function is worthwhile to be considered as a good phosphoenolpyruvate analogue. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Alcohol dehydrogenases in Acinetobacter sp. strain HO1-N: role in hexadecanse and hexadecanol metabolism

    International Nuclear Information System (INIS)

    Singer, M.E.; Finnerty, W.R.

    1985-01-01

    Multiple alcohol dehydrogenases (ADH) were demonstrated in Acinetobacter sp. strain HO1-N. ADH-A and ADH-B were distinguished on the basis of electrophoretic mobility, pyridine nucleotide cofactor requirement, and substrate specificity. ADH-A is a soluble, NAD-linked, inducible ethanol dehydrogenase (EDH). An ethanol-negative mutant (Eth1) was isolated which contained 6.5% of wild-type EDH activity and was deficient in ADH-A. Eth1 exhibited normal growth on hexadecane and hexadecanol. A second ethanol-negative mutant (Eth3) was acetaldehyde dehydrogenase (ALDH) deficient, having 12.5% of wild-type ALDH activity. Eth3 had threefold-higher EDH activity than the wild-type strain. ALDH is a soluble, NAD-linked, ethanol-inducible enzyme. Eth3 exhibited normal growth on hexadecane, hexadecanol, and fatty aldehyde. ADH-B is soluble, constitutive, NADP-linked ADH which was active with medium-chain-length alcohols. Hexadecanol dehydrogenase (HDH), a soluble and membrane-bound, NAD-linked ADH, was induced 5- to 11-fold by growth on hexadecane or hexadecanol. HDH was distinct from ADH-A and ADH-B. NAD-linked HDH appears to possess a functional role in hexadecane and hexadecanol dissimilation

  2. Diglycolic acid inhibits succinate dehydrogenase activity in human proximal tubule cells leading to mitochondrial dysfunction and cell death.

    Science.gov (United States)

    Landry, Greg M; Dunning, Cody L; Conrad, Taylor; Hitt, Mallory J; McMartin, Kenneth E

    2013-08-29

    Diethylene glycol (DEG) is a solvent used in consumer products allowing the increased risk for consumer exposure. DEG metabolism produces two primary metabolites, 2-hydroxyethoxyacetic acid (2-HEAA) and diglycolic acid (DGA). DGA has been shown to be the toxic metabolite responsible for the proximal tubule cell necrosis seen in DEG poisoning. The mechanism of DGA toxicity in the proximal tubule cell is not yet known. The chemical structure of DGA is very similar to citric acid cycle intermediates. Studies were designed to assess whether its mechanism of toxicity involves disruption of cellular metabolic pathways resulting in mitochondrial dysfunction. First, DGA preferentially inhibited succinate dehydrogenase, including human kidney cell enzyme, but had no effect on other citric acid cycle enzyme activities. DGA produces a cellular ATP depletion that precedes cell death. Human proximal tubule (HPT) cells, pre-treated with increasing DGA concentrations, showed significantly decreased oxygen consumption. DGA did not increase lactate levels, indicating no effect on glycolytic activity. DGA increased reactive oxygen species (ROS) production in HPT cells in a concentration and time dependent manner. These results indicate that DGA produced proximal tubule cell dysfunction by specific inhibition of succinate dehydrogenase and oxygen consumption. Disruption of these processes results in decreased energy production and proximal tubule cell death. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  3. Novel NAD+-Farnesal Dehydrogenase from Polygonum minus Leaves. Purification and Characterization of Enzyme in Juvenile Hormone III Biosynthetic Pathway in Plant.

    Directory of Open Access Journals (Sweden)

    Ahmad-Faris Seman-Kamarulzaman

    Full Text Available Juvenile Hormone III is of great concern due to negative effects on major developmental and reproductive maturation in insect pests. Thus, the elucidation of enzymes involved JH III biosynthetic pathway has become increasing important in recent years. One of the enzymes in the JH III biosynthetic pathway that remains to be isolated and characterized is farnesal dehydrogenase, an enzyme responsible to catalyze the oxidation of farnesal into farnesoic acid. A novel NAD+-farnesal dehydrogenase of Polygonum minus was purified (315-fold to apparent homogeneity in five chromatographic steps. The purification procedures included Gigacap S-Toyopearl 650M, Gigacap Q-Toyopearl 650M, and AF-Blue Toyopearl 650ML, followed by TSK Gel G3000SW chromatographies. The enzyme, with isoelectric point of 6.6 is a monomeric enzyme with a molecular mass of 70 kDa. The enzyme was relatively active at 40°C, but was rapidly inactivated above 45°C. The optimal temperature and pH of the enzyme were found to be 35°C and 9.5, respectively. The enzyme activity was inhibited by sulfhydryl agent, chelating agent, and metal ion. The enzyme was highly specific for farnesal and NAD+. Other terpene aldehydes such as trans- cinnamaldehyde, citral and α- methyl cinnamaldehyde were also oxidized but in lower activity. The Km values for farnesal, citral, trans- cinnamaldehyde, α- methyl cinnamaldehyde and NAD+ were 0.13, 0.69, 0.86, 1.28 and 0.31 mM, respectively. The putative P. minus farnesal dehydrogenase that's highly specific towards farnesal but not to aliphatic aldehydes substrates suggested that the enzyme is significantly different from other aldehyde dehydrogenases that have been reported. The MALDI-TOF/TOF-MS/MS spectrometry further identified two peptides that share similarity to those of previously reported aldehyde dehydrogenases. In conclusion, the P. minus farnesal dehydrogenase may represent a novel plant farnesal dehydrogenase that exhibits distinctive substrate

  4. Aldehyde PEGylation of laccase from Trametes versicolor in route to increase its stability: effect on enzymatic activity.

    Science.gov (United States)

    Mayolo-Deloisa, Karla; González-González, Mirna; Simental-Martínez, Jesús; Rito-Palomares, Marco

    2015-03-01

    Laccase is a multicopper oxidase that catalyzes the oxidation of phenolic compounds. Laccase can be used in bioremediation, beverage (wine, fruit juice, and beer) processing, ascorbic acid determination, sugar beet pectin gelation baking, and as a biosensor. Recently, the antiproliferative activity of laccase toward tumor cells has been reported. Because of the potential applications of this enzyme, the efforts for enhancing and stabilizing its activity have increased. Thus, the PEGylation of laccase can be an alternative. PEGylation is the covalent attachment of one or more molecules of methoxy poly(ethylene glycol) (mPEG) to a protein. Normally, during the PEGylation reaction, the activity is reduced but the stability increases; thus, it is important to minimize the loss of activity. In this work, the effects of molar ratio (1:4, 1:8, and 1:12), concentration of laccase (6 and 12 mg/ml), reaction time (4 and 17 h), molecular weight, and type of mPEG (20, 30, 40 kDa and 40 kDa-branched) were analyzed. The activity was measured using three substrates: ABTS, 2,6-dimethoxyphenol, and syringaldazine. The best conditions for laccase PEGylation were 12 mg/ml of laccase, molar ratio 1:4, and 4 h reaction time. Under these conditions, the enzyme was able to maintain nearly 100% of its enzymatic activity with ABTS. The PEGylation of laccase has not been extensively explored, so it is important to analyze the effects of this bioconjugation in route to produce a robust modified enzyme. Copyright © 2015 John Wiley & Sons, Ltd.

  5. Inhibitory effects of drugs on the metabolic activity of mouse and human aldehyde oxidases and influence on drug-drug interactions.

    Science.gov (United States)

    Takaoka, Naoki; Sanoh, Seigo; Okuda, Katsuhiro; Kotake, Yaichiro; Sugahara, Go; Yanagi, Ami; Ishida, Yuji; Tateno, Chise; Tayama, Yoshitaka; Sugihara, Kazumi; Kitamura, Shigeyuki; Kurosaki, Mami; Terao, Mineko; Garattini, Enrico; Ohta, Shigeru

    2018-04-17

    As aldehyde oxidase (AOX) plays an emerging role in drug metabolism, understanding its significance for drug-drug interactions (DDI) is important. Therefore, we tested 10 compounds for species-specific and substrate-dependent differences in the inhibitory effect of AOX activity using genetically engineered HEK293 cells over-expressing human AOX1, mouse AOX1 or mouse AOX3. The IC 50 values of 10 potential inhibitors of the three AOX enzymes were determined using phthalazine and O 6 -benzylguanine as substrates. 17β-Estradiol, menadione, norharmane and raloxifene exhibited marked differences in inhibitory effects between the human and mouse AOX isoforms when the phthalazine substrate was used. Some of the compounds tested exhibited substrate-dependent differences in their inhibitory effects. Docking simulations with human AOX1 and mouse AOX3 were conducted for six representative inhibitors. The rank order of the minimum binding energy reflected the order of the corresponding IC 50 values. We also evaluated the potential DDI between an AOX substrate (O 6 -benzylguanine) and an inhibitor (hydralazine) using chimeric mice with humanized livers. Pretreatment of hydralazine increased the maximum plasma concentration (C max ) and the area under the plasma concentration-time curve (AUC 0-24 ) of O 6 -benzylguanine compared to single administration. Our in vitro data indicate species-specific and substrate-dependent differences in the inhibitory effects on AOX activity. Our in vivo data demonstrate the existence of a DDI which may be of relevance in the clinical context. Copyright © 2018 Elsevier Inc. All rights reserved.

  6. Glutamate production from ammonia via glutamate dehydrogenase 2 activity supports cancer cell proliferation under glutamine depletion.

    Science.gov (United States)

    Takeuchi, Yukiko; Nakayama, Yasumune; Fukusaki, Eiichiro; Irino, Yasuhiro

    2018-01-01

    Cancer cells rapidly consume glutamine as a carbon and nitrogen source to support proliferation, but the cell number continues to increase exponentially after glutamine is nearly depleted from the medium. In contrast, cell proliferation rates are strongly depressed when cells are cultured in glutamine-free medium. How cancer cells survive in response to nutrient limitation and cellular stress remains poorly understood. In addition, rapid glutamine catabolism yields ammonia, which is a potentially toxic metabolite that is secreted into the extracellular space. Here, we show that ammonia can be utilized for glutamate production, leading to cell proliferation under glutamine-depleted conditions. This proliferation requires glutamate dehydrogenase 2, which synthesizes glutamate from ammonia and α-ketoglutarate and is expressed in MCF7 and T47D cells. Our findings provide insight into how cancer cells survive under glutamine deprivation conditions and thus contribute to elucidating the mechanisms of tumor growth. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Bioinformatics based structural characterization of glucose dehydrogenase (gdh gene and growth promoting activity of Leclercia sp. QAU-66

    Directory of Open Access Journals (Sweden)

    Muhammad Naveed

    2014-06-01

    Full Text Available Glucose dehydrogenase (GDH; EC 1.1. 5.2 is the member of quinoproteins group that use the redox cofactor pyrroloquinoline quinoine, calcium ions and glucose as substrate for its activity. In present study, Leclercia sp. QAU-66, isolated from rhizosphere of Vigna mungo, was characterized for phosphate solubilization and the role of GDH in plant growth promotion of Phaseolus vulgaris. The strain QAU-66 had ability to solubilize phosphorus and significantly (p < 0.05 promoted the shoot and root lengths of Phaseolus vulgaris. The structural determination of GDH protein was carried out using bioinformatics tools like Pfam, InterProScan, I-TASSER and COFACTOR. These tools predicted the structural based functional homology of pyrroloquinoline quinone domains in GDH. GDH of Leclercia sp. QAU-66 is one of the main factor that involved in plant growth promotion and provides a solid background for further research in plant growth promoting activities.

  8. Bioinformatics based structural characterization of glucose dehydrogenase (gdh) gene and growth promoting activity of Leclercia sp. QAU-66.

    Science.gov (United States)

    Naveed, Muhammad; Ahmed, Iftikhar; Khalid, Nauman; Mumtaz, Abdul Samad

    2014-01-01

    Glucose dehydrogenase (GDH; EC 1.1. 5.2) is the member of quinoproteins group that use the redox cofactor pyrroloquinoline quinoine, calcium ions and glucose as substrate for its activity. In present study, Leclercia sp. QAU-66, isolated from rhizosphere of Vigna mungo, was characterized for phosphate solubilization and the role of GDH in plant growth promotion of Phaseolus vulgaris. The strain QAU-66 had ability to solubilize phosphorus and significantly (p ≤ 0.05) promoted the shoot and root lengths of Phaseolus vulgaris. The structural determination of GDH protein was carried out using bioinformatics tools like Pfam, InterProScan, I-TASSER and COFACTOR. These tools predicted the structural based functional homology of pyrroloquinoline quinone domains in GDH. GDH of Leclercia sp. QAU-66 is one of the main factor that involved in plant growth promotion and provides a solid background for further research in plant growth promoting activities.

  9. Inhibition of catalase by aminotriazole in vivo results in reduction of glucose-6-phosphate dehydrogenase activity in Saccharomyces cerevisiae cells.

    Science.gov (United States)

    Bayliak, M; Gospodaryov, D; Semchyshyn, H; Lushchak, V

    2008-04-01

    The inhibitor of catalase 3-amino-1,2,4-triazole (AMT) was used to study the physiological role of catalase in the yeast Saccharomyces cerevisiae under starvation. It was shown that AMT at the concentration of 10 mM did not affect the growth of the yeast. In vivo and in vitro the degree of catalase inhibition by AMT was concentration- and time-dependent. Peroxisomal catalase in bakers' yeast was more sensitive to AMT than the cytosolic one. In vivo inhibition of catalase by AMT in S. cerevisiae caused a simultaneous decrease in glucose-6-phosphate dehydrogenase activity and an increase in glutathione reductase activity. At the same time, the level of protein carbonyls, a marker of oxidative modification, was not affected. Possible mechanisms compensating the negative effects caused by AMT inhibition of catalase are discussed.

  10. Effects of synthetic detergents on in vivo activity of tissue phosphatases and succinic dehydrogenase from Mystus vittatus

    Energy Technology Data Exchange (ETDEWEB)

    Mohan, D.; Verma, S.R.

    1981-05-01

    African catfish (Mystus vittatus) were exposed to three sub-lethal concentrations of Swascofix E45 (13.8, 9.2 and 4.6 mg/l) and Swascol 3L (69.3, 46.2 and 23.1 mg/l) for 15 and 30 days, and their effects on alkaline and acid phosphatase, and succinic dehydrogenase in liver, kidney and intestine were measured. The enzymes were found to be inhibited in all the tissues. Maximum inhibition (38.44%) was observed in liver alkaline phosphatase activity after 30 days with the highest concentration of Swascofix E45 and the lowest inhibition (0.118%) was found in kidney acid phosphatase activity with the lowest concentration of Swascol 3L after 15 days. Insignificant enzyme stimulation in some cases was also observed.

  11. Palladium alpha-lipoic acid complex formulation enhances activities of Krebs cycle dehydrogenases and respiratory complexes I-IV in the heart of aged rats.

    Science.gov (United States)

    Sudheesh, N P; Ajith, T A; Janardhanan, K K; Krishnan, C V

    2009-08-01

    Age-related decline in the capacity to withstand stress, such as ischemia and reperfusion, results in congestive heart failure. Though the mechanisms underlying cardiac decay are not clear, age dependent somatic damages to mitochondrial DNA (mtDNA), loss of mitochondrial function, and a resultant increase in oxidative stress in heart muscle cells may be responsible for the increased risk for cardiovascular diseases. The effect of a safe nutritional supplement, POLY-MVA, containing the active ingredient palladium alpha-lipoic acid complex, was evaluated on the activities of the Krebs cycle enzymes such as isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, and malate dehydrogenase as well as mitochondrial complexes I, II, III, and IV in heart mitochondria of aged male albino rats of Wistar strain. Administration of 0.05 ml/kg of POLY-MVA (which is equivalent to 0.38 mg complexed alpha-lipoic acid/kg, p.o), once daily for 30 days, was significantly (pKrebs cycle dehydrogenases, and mitochondrial electron transport chain complexes. The unique electronic and redox properties of palladium alpha-lipoic acid complex appear to be a key to this physiological effectiveness. The results strongly suggest that this formulation might be effective to protect the aging associated risk of cardiovascular and neurodegenerative diseases.

  12. STUDIES ON THE DYNAMICS OF DEHYDROGENASES KREBS CYCLE ACTIVITY AT MONILINIA LAXA (ADERH. & RUHL. HONEY FUNGUS GROWN ON MEDIA WITH DIFFERENT CARBOHYDRATES

    Directory of Open Access Journals (Sweden)

    Elena Ciornea

    2010-09-01

    Full Text Available As ubiquitous organisms, fungi grow on a large number of organic substrate, alive or dead, confronting therefore with a wide variety of carbohydrates and various physical factors, and their versatility to adapt and be able to use a large number of these compounds could provide them the chance to survive. Given that, these fungi have a rich enzyme equipment that allows them to operate on different metabolic pathways, this study aims to monitor the dynamics activity of some Krebs cycle dehydrogenases in Monilinia laxa (Aderh & Ruhl. Honey species parasitic on various species of plum trees. To this end, the fungus was cultivated in vitro on media enriched with different carbohydrates and the isocitrate dehydrogenase, �-cetoglutarate dehydrogenase, succinate dehydrogenase and malate dehydrogenase activity in the fungus mycelium was followed, at 7, respectively, 14 days after the inoculation of the culture medium and determined using the spectrophotometric Sîsoev and Krasna method (Cojocaru, D.C., 2009. Data revealed obvious differences depending on the type of carbohydrate introduced into the medium and the age of the culture mycelia.

  13. STUDIES ON THE DYNAMICS OF DEHYDROGENASES KREBS CYCLE ACTIVITY AT MONILINIA LAXA (ADERH. & RUHL. HONEY FUNGUS GROWN ON MEDIA WITH DIFFERENT CARBOHYDRATES

    Directory of Open Access Journals (Sweden)

    Elena Ciornea

    2011-11-01

    Full Text Available As ubiquitous organisms, fungi grow on a large number of organic substrate, alive or dead, confronting therefore with a wide variety of carbohydrates and various physical factors, and their versatility to adapt and be able to use a large number of these compounds could provide them the chance to survive. Given that, these fungi have a rich enzyme equipment that allows them to operate on different metabolic pathways, this study aims to monitor the dynamics activity of some Krebs cycle dehydrogenases in Monilinia laxa (Aderh & Ruhl. Honey species parasitic on various species of plum trees. To this end, the fungus was cultivated in vitro on media enriched with different carbohydrates and the isocitrate dehydrogenase, �-cetoglutarate dehydrogenase, succinate dehydrogenase and malate dehydrogenase activity in the fungus mycelium was followed, at 7, respectively, 14 days after the inoculation of the culture medium and determined using the spectrophotometric Sîsoev and Krasna method (Cojocaru, D.C., 2009. Data revealed obvious differences depending on the type of carbohydrate introduced into the medium and the age of the culture mycelia.

  14. Determination of glutamate dehydrogenase activity and its kinetics in mouse tissues using metabolic mapping (quantitative enzyme histochemistry).

    Science.gov (United States)

    Botman, Dennis; Tigchelaar, Wikky; Van Noorden, Cornelis J F

    2014-11-01

    Glutamate dehydrogenase (GDH) catalyses the reversible conversion of glutamate into α-ketoglutarate with the concomitant reduction of NAD(P)(+) to NAD(P)H or vice versa. GDH activity is subject to complex allosteric regulation including substrate inhibition. To determine GDH kinetics in situ, we assessed the effects of various glutamate concentrations in combination with either the coenzyme NAD(+) or NADP(+) on GDH activity in mouse liver cryostat sections using metabolic mapping. NAD(+)-dependent GDH V(max) was 2.5-fold higher than NADP(+)-dependent V(max), whereas the K(m) was similar, 1.92 mM versus 1.66 mM, when NAD(+) or NADP(+) was used, respectively. With either coenzyme, V(max) was determined at 10 mM glutamate and substrate inhibition was observed at higher glutamate concentrations with a K(i) of 12.2 and 3.95 for NAD(+) and NADP(+) used as coenzyme, respectively. NAD(+)- and NADP(+)-dependent GDH activities were examined in various mouse tissues. GDH activity was highest in liver and much lower in other tissues. In all tissues, the highest activity was found when NAD(+) was used as a coenzyme. In conclusion, GDH activity in mice is highest in the liver with NAD(+) as a coenzyme and highest GDH activity was determined at a glutamate concentration of 10 mM. © The Author(s) 2014.

  15. Towards a systematic analysis of human short-chain dehydrogenases/reductases (SDR): Ligand identification and structure-activity relationships.

    Science.gov (United States)

    Bhatia, Chitra; Oerum, Stephanie; Bray, James; Kavanagh, Kathryn L; Shafqat, Naeem; Yue, Wyatt; Oppermann, Udo

    2015-06-05

    Short-chain dehydrogenases/reductases (SDRs) constitute a large, functionally diverse branch of enzymes within the class of NAD(P)(H) dependent oxidoreductases. In humans, over 80 genes have been identified with distinct metabolic roles in carbohydrate, amino acid, lipid, retinoid and steroid hormone metabolism, frequently associated with inherited genetic defects. Besides metabolic functions, a subset of atypical SDR proteins appears to play critical roles in adapting to redox status or RNA processing, and thereby controlling metabolic pathways. Here we present an update on the human SDR superfamily and a ligand identification strategy using differential scanning fluorimetry (DSF) with a focused library of oxidoreductase and metabolic ligands to identify substrate classes and inhibitor chemotypes. This method is applicable to investigate structure-activity relationships of oxidoreductases and ultimately to better understand their physiological roles. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  16. The level of glucose-6-phosphate dehydrogenase activity strongly influences xylose fermentation and inhibitor sensitivity in recombinant Saccharomyces cerevisiae strains

    DEFF Research Database (Denmark)

    Jeppsson, M.; Johansson, B.; Jensen, Peter Ruhdal

    2003-01-01

    Disruption of the ZWF1 gene encoding glucose-6-phosphate dehydrogenase (G6PDH) has been shown to reduce the xylitol yield and the xylose consumption in the xylose-utilizing recombinant Saccharomyces cerevisiae strain TMB3255. In the present investigation we have studied the influence of different...... consumption, respectively, compared with the ZWF1-disrupted strain. Both strains exhibited decreased xylitol yields (0.13 and 0.19 g/g xylose) and enhanced ethanol yields (0.36 and 0.34 g/g xylose) compared with the control strain TMB3001 (0.29 g xylitol/g xylose, 0.31 g ethanol/g xylose). Cytoplasmic...... transhydrogenase (TH) from Azotobacter vinelandii has previously been shown to transfer NADPH and NAD(+) into NADP(+) and NADH, and TH-overproduction resulted in lower xylitol yield and enhanced glycerol yield during xylose utilization. Strains with low G6PDH-activity grew slower in a lignocellulose hydrolysate...

  17. Alcohol, Aldehydes, Adducts and Airways

    Directory of Open Access Journals (Sweden)

    Muna Sapkota

    2015-11-01

    Full Text Available Drinking alcohol and smoking cigarettes results in the formation of reactive aldehydes in the lung, which are capable of forming adducts with several proteins and DNA. Acetaldehyde and malondialdehyde are the major aldehydes generated in high levels in the lung of subjects with alcohol use disorder who smoke cigarettes. In addition to the above aldehydes, several other aldehydes like 4-hydroxynonenal, formaldehyde and acrolein are also detected in the lung due to exposure to toxic gases, vapors and chemicals. These aldehydes react with nucleophilic targets in cells such as DNA, lipids and proteins to form both stable and unstable adducts. This adduction may disturb cellular functions as well as damage proteins, nucleic acids and lipids. Among several adducts formed in the lung, malondialdehyde DNA (MDA-DNA adduct and hybrid malondialdehyde-acetaldehyde (MAA protein adducts have been shown to initiate several pathological conditions in the lung. MDA-DNA adducts are pre-mutagenic in mammalian cells and induce frame shift and base-pair substitution mutations, whereas MAA protein adducts have been shown to induce inflammation and inhibit wound healing. This review provides an insight into different reactive aldehyde adducts and their role in the pathogenesis of lung disease.

  18. Alcohol dehydrogenase gene ADH3 activates glucose alcoholic fermentation in genetically engineered Dekkera bruxellensis yeast.

    Science.gov (United States)

    Schifferdecker, Anna Judith; Siurkus, Juozas; Andersen, Mikael Rørdam; Joerck-Ramberg, Dorte; Ling, Zhihao; Zhou, Nerve; Blevins, James E; Sibirny, Andriy A; Piškur, Jure; Ishchuk, Olena P

    2016-04-01

    Dekkera bruxellensis is a non-conventional Crabtree-positive yeast with a good ethanol production capability. Compared to Saccharomyces cerevisiae, its tolerance to acidic pH and its utilization of alternative carbon sources make it a promising organism for producing biofuel. In this study, we developed an auxotrophic transformation system and an expression vector, which enabled the manipulation of D. bruxellensis, thereby improving its fermentative performance. Its gene ADH3, coding for alcohol dehydrogenase, was cloned and overexpressed under the control of the strong and constitutive promoter TEF1. Our recombinant D. bruxellensis strain displayed 1.4 and 1.7 times faster specific glucose consumption rate during aerobic and anaerobic glucose fermentations, respectively; it yielded 1.2 times and 1.5 times more ethanol than did the parental strain under aerobic and anaerobic conditions, respectively. The overexpression of ADH3 in D. bruxellensis also reduced the inhibition of fermentation by anaerobiosis, the "Custer effect". Thus, the fermentative capacity of D. bruxellensis could be further improved by metabolic engineering.

  19. Green Tea Polyphenols Control Dysregulated Glutamate Dehydrogenase in Transgenic Mice by Hijacking the ADP Activation Site

    Energy Technology Data Exchange (ETDEWEB)

    Li, Changhong; Li, Ming; Chen, Pan; Narayan, Srinivas; Matschinsky, Franz M.; Bennett, Michael J.; Stanley, Charles A.; Smith, Thomas J. (CH-PA); (UPENN); (Danforth)

    2012-05-09

    Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and, in animals, is extensively regulated by a number of metabolites. Gain of function mutations in GDH that abrogate GTP inhibition cause the hyperinsulinism/hyperammonemia syndrome (HHS), resulting in increased pancreatic {beta}-cell responsiveness to leucine and susceptibility to hypoglycemia following high protein meals. We have previously shown that two of the polyphenols from green tea (epigallocatechin gallate (EGCG) and epicatechin gallate (ECG)) inhibit GDH in vitro and that EGCG blocks GDH-mediated insulin secretion in wild type rat islets. Using structural and site-directed mutagenesis studies, we demonstrate that ECG binds to the same site as the allosteric regulator, ADP. Perifusion assays using pancreatic islets from transgenic mice expressing a human HHS form of GDH demonstrate that the hyperresponse to glutamine caused by dysregulated GDH is blocked by the addition of EGCG. As observed in HHS patients, these transgenic mice are hypersensitive to amino acid feeding, and this is abrogated by oral administration of EGCG prior to challenge. Finally, the low basal blood glucose level in the HHS mouse model is improved upon chronic administration of EGCG. These results suggest that this common natural product or some derivative thereof may prove useful in controlling this genetic disorder. Of broader clinical implication is that other groups have shown that restriction of glutamine catabolism via these GDH inhibitors can be useful in treating various tumors. This HHS transgenic mouse model offers a highly useful means to test these agents in vivo.

  20. Succinate-dependent energy generation and pyruvate dehydrogenase complex activity in isolated Ascaris suum mitochondria

    Energy Technology Data Exchange (ETDEWEB)

    Campbell, T.A.

    1988-01-01

    Body wall muscle from the parasitic nematode, Ascaris suum, contain unique anaerobic mitochondria that preferentially utilize fumarate and branched-chain enoyl CoA's as terminal electron acceptors instead of oxygen. While electron transport in these organelles is well characterized, the role of oxygen in succinate-dependent phosphorylation is still not clearly defined. Therefore, the present study was designed to more fully characterize succinate metabolism in these organelles as well as the in vitro regulation of a key mitochondrial enzyme, the pyruvate dehydrogenase complex (PDC). In the absence of added adenine nucleotides, incubations in succinate resulted in substantial elevations in intramitochrondrial ATP levels, but ATP/ADP ratios were considerably higher in incubations with malate. The stimulation of phosphorylation in aerobic incubations with succinate was rotenone sensitive and appears to be Site I dependent. Increase substrate level phosphorylation, coupled to propionate formation, or additional sites of electron-transport associated ATP synthesis were not significant. Under aerobic conditions, {sup 14}CO{sub 2} evolution from 1,4-({sup 14}C)succinate was stimulated and NADH/NAD{sup +} ratios were elevated, but the formation of {sup 14}C propionate was unchanged.

  1. Overexpression of a GmCnx1 gene enhanced activity of nitrate reductase and aldehyde oxidase, and boosted mosaic virus resistance in soybean.

    Directory of Open Access Journals (Sweden)

    Zheng Zhou

    Full Text Available Molybdenum cofactor (Moco is required for the activities of Moco-dependant enzymes. Cofactor for nitrate reductase and xanthine dehydrogenase (Cnx1 is known to be involved in the biosynthesis of Moco in plants. In this work, a soybean (Glycine max L. Cnx1 gene (GmCnx1 was transferred into soybean using Agrobacterium tumefaciens-mediated transformation method. Twenty seven positive transgenic soybean plants were identified by coating leaves with phosphinothricin, bar protein quick dip stick and PCR analysis. Moreover, Southern blot analysis was carried out to confirm the insertion of GmCnx1 gene. Furthermore, expression of GmCnx1 gene in leaf and root of all transgenic lines increased 1.04-2.12 and 1.55-3.89 folds, respectively, as compared to wild type with GmCnx1 gene and in line 10 , 22 showing the highest expression. The activities of Moco-related enzymes viz nitrate reductase (NR and aldehydeoxidase (AO of T1 generation plants revealed that the best line among the GmCnx1 transgenic plants accumulated 4.25 μg g(-1 h(-1 and 30 pmol L(-1, respectively (approximately 2.6-fold and 3.9-fold higher than non-transgenic control plants.In addition, overexpression ofGmCnx1boosted the resistance to various strains of soybean mosaic virus (SMV. DAS-ELISA analysis further revealed that infection rate of GmCnx1 transgenic plants were generally lower than those of non-transgenic plants among two different virus strains tested. Taken together, this study showed that overexpression of a GmCnx1 gene enhanced NR and AO activities and SMV resistance, suggesting its important role in soybean genetic improvement.

  2. Novel dehydrogenase catalyzes oxidative hydrolysis of carbon-nitrogen double bonds for hydrazone degradation.

    Science.gov (United States)

    Itoh, Hideomi; Suzuta, Tetsuya; Hoshino, Takayuki; Takaya, Naoki

    2008-02-29

    Hydrazines and their derivatives are versatile artificial and natural compounds that are metabolized by elusive biological systems. Here we identified microorganisms that assimilate hydrazones and isolated the yeast, Candida palmioleophila MK883. When cultured with adipic acid bis(ethylidene hydrazide) as the sole source of carbon, C. palmioleophila MK883 degraded hydrazones and accumulated adipic acid dihydrazide. Cytosolic NAD+- or NADP+-dependent hydrazone dehydrogenase (Hdh) activity was detectable under these conditions. The production of Hdh was inducible by adipic acid bis(ethylidene hydrazide) and the hydrazone, varelic acid ethylidene hydrazide, under the control of carbon catabolite repression. Purified Hdh oxidized and hydrated the C=N double bond of acetaldehyde hydrazones by reducing NAD+ or NADP+ to produce relevant hydrazides and acetate, the latter of which the yeast assimilated. The deduced amino acid sequence revealed that Hdh belongs to the aldehyde dehydrogenase (Aldh) superfamily. Kinetic and mutagenesis studies showed that Hdh formed a ternary complex with the substrates and that conserved Cys is essential for the activity. The mechanism of Hdh is similar to that of Aldh, except that it catalyzed oxidative hydrolysis of hydrazones that requires adding a water molecule to the reaction catalyzed by conventional Aldh. Surprisingly, both Hdh and Aldh from baker's yeast (Ald4p) catalyzed the Hdh reaction as well as aldehyde oxidation. Our findings are unique in that we discovered a biological mechanism for hydrazone utilization and a novel function of proteins in the Aldh family that act on C=N compounds.

  3. In Silico Identification and in Vitro Activity of Novel Natural Inhibitors of Trypanosoma brucei Glyceraldehyde-3-phosphate-dehydrogenase

    Directory of Open Access Journals (Sweden)

    Fabian C. Herrmann

    2015-09-01

    Full Text Available As part of our ongoing efforts to identify natural products with activity against pathogens causing neglected tropical diseases, we are currently performing an extensive screening of natural product (NP databases against a multitude of protozoan parasite proteins. Within this project, we screened a database of NPs from a commercial supplier, AnalytiCon Discovery (Potsdam, Germany, against Trypanosoma brucei glyceraldehyde-3-phosphate dehydrogenase (TbGAPDH, a glycolytic enzyme whose inhibition deprives the parasite of energy supply. NPs acting as potential inhibitors of the mentioned enzyme were identified using a pharmacophore-based virtual screening and subsequent docking of the identified hits into the active site of interest. In a set of 700 structures chosen for the screening, 13 (1.9% were predicted to possess significant affinity towards the enzyme and were therefore tested in an in vitro enzyme assay using recombinant TbGAPDH. Nine of these in silico hits (69% showed significant inhibitory activity at 50 µM, of which two geranylated benzophenone derivatives proved to be particularly active with IC50 values below 10 µM. These compounds also showed moderate in vitro activity against T. brucei rhodesiense and may thus represent interesting starting points for further optimization.

  4. In Silico Identification and in Vitro Activity of Novel Natural Inhibitors of Trypanosoma brucei Glyceraldehyde-3-phosphate-dehydrogenase.

    Science.gov (United States)

    Herrmann, Fabian C; Lenz, Mairin; Jose, Joachim; Kaiser, Marcel; Brun, Reto; Schmidt, Thomas J

    2015-09-03

    As part of our ongoing efforts to identify natural products with activity against pathogens causing neglected tropical diseases, we are currently performing an extensive screening of natural product (NP) databases against a multitude of protozoan parasite proteins. Within this project, we screened a database of NPs from a commercial supplier, AnalytiCon Discovery (Potsdam, Germany), against Trypanosoma brucei glyceraldehyde-3-phosphate dehydrogenase (TbGAPDH), a glycolytic enzyme whose inhibition deprives the parasite of energy supply. NPs acting as potential inhibitors of the mentioned enzyme were identified using a pharmacophore-based virtual screening and subsequent docking of the identified hits into the active site of interest. In a set of 700 structures chosen for the screening, 13 (1.9%) were predicted to possess significant affinity towards the enzyme and were therefore tested in an in vitro enzyme assay using recombinant TbGAPDH. Nine of these in silico hits (69%) showed significant inhibitory activity at 50 µM, of which two geranylated benzophenone derivatives proved to be particularly active with IC50 values below 10 µM. These compounds also showed moderate in vitro activity against T. brucei rhodesiense and may thus represent interesting starting points for further optimization.

  5. Chronic Inhibition of 11β-Hydroxysteroid Dehydrogenase Type 1 Activity Decreases Hypertension, Insulin Resistance, and Hypertriglyceridemia in Metabolic Syndrome

    Directory of Open Access Journals (Sweden)

    Christine G. Schnackenberg

    2013-01-01

    Full Text Available Metabolic syndrome is a constellation of risk factors including hypertension, dyslipidemia, insulin resistance, and obesity that promote the development of cardiovascular disease. Metabolic syndrome has been associated with changes in the secretion or metabolism of glucocorticoids, which have important functions in adipose, liver, kidney, and vasculature. Tissue concentrations of the active glucocorticoid cortisol are controlled by the conversion of cortisone to cortisol by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1. Because of the various cardiovascular and metabolic activities of glucocorticoids, we tested the hypothesis that 11β-HSD1 is a common mechanism in the hypertension, dyslipidemia, and insulin resistance in metabolic syndrome. In obese and lean SHR/NDmcr-cp (SHR-cp, cardiovascular, metabolic, and renal functions were measured before and during four weeks of administration of vehicle or compound 11 (10 mg/kg/d, a selective inhibitor of 11β-HSD1. Compound 11 significantly decreased 11β-HSD1 activity in adipose tissue and liver of SHR-cp. In obese SHR-cp, compound 11 significantly decreased mean arterial pressure, glucose intolerance, insulin resistance, hypertriglyceridemia, and plasma renin activity with no effect on heart rate, body weight gain, or microalbuminuria. These results suggest that 11β-HSD1 activity in liver and adipose tissue is a common mediator of hypertension, hypertriglyceridemia, glucose intolerance, and insulin resistance in metabolic syndrome.

  6. Elevation of Glucose 6-Phosphate Dehydrogenase Activity Induced by Amplified Insulin Response in Low Glutathione Levels in Rat Liver

    Directory of Open Access Journals (Sweden)

    Misako Taniguchi

    2016-01-01

    Full Text Available Weanling male Wistar rats were fed on a 10% soybean protein isolate (SPI diet for 3 weeks with or without supplementing 0.3% sulfur-containing amino acids (SAA; methionine or cystine to examine relationship between glutathione (GSH levels and activities of NADPH-producing enzymes, glucose 6-phosphate dehydrogenase (G6PD and malic enzyme (ME, in the liver. Of rats on the 10% SPI diet, GSH levels were lower and the enzyme activities were higher than of those fed on an SAA-supplemented diet. Despite the lower GSH level, γ-glutamylcysteine synthetase (γ-GCS activity was higher in the 10% SPI group than other groups. Examination of mRNAs of G6PD and ME suggested that the GSH-suppressing effect on enzyme induction occurred prior to and/or at transcriptional levels. Gel electrophoresis of G6PD indicated that low GSH status caused a decrease in reduced form and an increase in oxidized form of the enzyme, suggesting an accelerated turnover rate of the enzyme. In primary cultured hepatocytes, insulin response to induce G6PD activity was augmented in low GSH levels manipulated in the presence of buthionine sulfoximine. These findings indicated that elevation of the G6PD activity in low GSH levels was caused by amplified insulin response for expression of the enzyme and accelerated turnover rate of the enzyme molecule.

  7. The Arabidopsis KS-type dehydrin recovers lactate dehydrogenase activity inhibited by copper with the contribution of His residues.

    Science.gov (United States)

    Hara, Masakazu; Monna, Shuhei; Murata, Takae; Nakano, Taiyo; Amano, Shono; Nachbar, Markus; Wätzig, Hermann

    2016-04-01

    Dehydrin, which is one of the late embryogenesis abundant (LEA) proteins, is involved in the ability of plants to tolerate the lack of water. Although many reports have indicated that dehydrins bind heavy metals, the physiological role of this metal binding has not been well understood. Here, we report that the Arabidopsis KS-type dehydrin (AtHIRD11) recovered the lactate dehydrogenase (LDH) activity denatured by Cu(2+). The LDH activity was partially inhibited by 0.93 μM Cu(2+) but totally inactivated by 9.3 μM Cu(2+). AtHIRD11 recovered the activity of LDH treated with 9.3 μM Cu(2+) in a dose-dependent manner. The recovery activity of AtHIRD11 was significantly higher than those of serum albumin and lysozyme. The conversion of His residues to Ala in AtHIRD11 resulted in the loss of the Cu(2+) binding of the protein as well as the disappearance of the conformational change induced by Cu(2+) that is observed by circular dichroism spectroscopy. The mutant protein showed lower recovery activity than the original AtHIRD11. These results indicate that AtHIRD11 can reactivate LDH inhibited by Cu(2+) via the His residues. This function may prevent physiological damage to plants due to heavy-metal stress. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  8. Effects of 14 days of spaceflight and nine days of recovery on cell body size and succinate dehydrogenase activity of rat dorsal root ganglion neurons

    Science.gov (United States)

    Ishihara, A.; Ohira, Y.; Roy, R. R.; Nagaoka, S.; Sekiguchi, C.; Hinds, W. E.; Edgerton, V. R.

    1997-01-01

    The cross-sectional areas and succinate dehydrogenase activities of L5 dorsal root ganglion neurons in rats were determined after 14 days of spaceflight and after nine days of recovery. The mean and distribution of the cross-sectional areas were similar to age-matched, ground-based controls for both the spaceflight and for the spaceflight plus recovery groups. The mean succinate dehydrogenase activity was significantly lower in spaceflight compared to aged-matched control rats, whereas the mean succinate dehydrogenase activity was similar in age-matched control and spaceflight plus recovery rats. The mean succinate dehydrogenase activity of neurons with cross-sectional areas between 1000 and 2000 microns2 was lower (between 7 and 10%) in both the spaceflight and the spaceflight plus recovery groups compared to the appropriate control groups. The reduction in the oxidative capacity of a subpopulation of sensory neurons having relatively large cross-sectional areas immediately following spaceflight and the sustained depression for nine days after returning to 1 g suggest that the 0 g environment induced significant alterations in proprioceptive function.

  9. A Conserved Active Site Tyrosine Residue of Proline Dehydrogenase Helps Enforce the Preference for Proline over Hydroxyproline as the Substrate

    Energy Technology Data Exchange (ETDEWEB)

    Ostrander, E.L.; Larson, J.D.; Schuermann, J.P.; Tanner, J.J.; (Cornell); (Missouri)

    2009-03-02

    Proline dehydrogenase (PRODH) catalyzes the oxidation of L-proline to {Delta}-1-pyrroline-5-carboxylate. PRODHs exhibit a pronounced preference for proline over hydroxyproline (trans-4-hydroxy-L-proline) as the substrate, but the basis for specificity is unknown. The goal of this study, therefore, is to gain insight into the structural determinants of substrate specificity of this class of enzyme, with a focus on understanding how PRODHs discriminate between the two closely related molecules, proline and hydroxyproline. Two site-directed mutants of the PRODH domain of Escherichia coli PutA were created: Y540A and Y540S. Kinetics measurements were performed with both mutants. Crystal structures of Y540S complexed with hydroxyproline, proline, and the proline analogue L-tetrahydro-2-furoic acid were determined at resolutions of 1.75, 1.90, and 1.85 {angstrom}, respectively. Mutation of Tyr540 increases the catalytic efficiency for hydroxyproline 3-fold and decreases the specificity for proline by factors of 20 (Y540S) and 50 (Y540A). The structures show that removal of the large phenol side chain increases the volume of the substrate-binding pocket, allowing sufficient room for the 4-hydroxyl of hydroxyproline. Furthermore, the introduced serine residue participates in recognition of hydroxyproline by forming a hydrogen bond with the 4-hydroxyl. This result has implications for understanding the substrate specificity of the related enzyme human hydroxyproline dehydrogenase, which has serine in place of tyrosine at this key active site position. The kinetic and structural results suggest that Tyr540 is an important determinant of specificity. Structurally, it serves as a negative filter for hydroxyproline by clashing with the 4-hydroxyl group of this potential substrate.

  10. Heat stress prevents the decrease in succinate dehydrogenase activity in the extensor digitorum longus of streptozotocin-induced diabetic rats.

    Science.gov (United States)

    Nonaka, K; Une, S; Komatsu, M; Yamaji, R; Akiyama, J

    2018-03-16

    This study aimed to investigate whether heat stress (HS) prevents a decrease in succinate dehydrogenase (SDH) activity and heat shock protein 60 (HSP60) and superoxide dismutase 2 (SOD2) contents in the extensor digitorum longus of streptozotocin (STZ)-induced diabetic rats. Twelve-week-old male Wistar rats were assigned to one of the four groups (n=6/group): control (Con), HS, diabetes mellitus (DM), and diabetes mellitus and heat stress (DM+HS). Diabetes was induced by the administration of STZ (50 mg/kg). HS was initiated 7 days after STZ treatment and performed at 42 °C for 30 min 5 times a week for 3 weeks. SDH activity was decreased in the DM and DM+HS groups. However, SDH activity was greater in the DM+HS group than in the DM group. Although HSP60 content was lower in the DM group than in the Con group, it was maintained in the DM+HS groups and was higher than that in the DM group. SOD2 content was decreased only in the DM group. These findings suggest that HS prevents the decrease in SDH activity in the skeletal muscle induced by DM. According to this mechanism, the maintenance of SOD2 and HSP60 by HS may suppress the increase in oxidative stress.

  11. Genetic analysis of central carbon metabolism unveils an amino acid substitution that alters maize NAD-dependent isocitrate dehydrogenase activity.

    Directory of Open Access Journals (Sweden)

    Nengyi Zhang

    2010-04-01

    Full Text Available Central carbon metabolism (CCM is a fundamental component of life. The participating genes and enzymes are thought to be structurally and functionally conserved across and within species. Association mapping utilizes a rich history of mutation and recombination to achieve high resolution mapping. Therefore, applying association mapping in maize (Zea mays ssp. mays, the most diverse model crop species, to study the genetics of CCM is a particularly attractive system.We used a maize diversity panel to test the CCM functional conservation. We found heritable variation in enzyme activity for every enzyme tested. One of these enzymes was the NAD-dependent isocitrate dehydrogenase (IDH, E.C. 1.1.1.41, in which we identified a novel amino-acid substitution in a phylogenetically conserved site. Using candidate gene association mapping, we identified that this non-synonymous polymorphism was associated with IDH activity variation. The proposed mechanism for the IDH activity variation includes additional components regulating protein level. With the comparison of sequences from maize and teosinte (Zea mays ssp. Parviglumis, the maize wild ancestor, we found that some CCM genes had also been targeted for selection during maize domestication.Our results demonstrate the efficacy of association mapping for dissecting natural variation in primary metabolic pathways. The considerable genetic diversity observed in maize CCM genes underlies heritable phenotypic variation in enzyme activities and can be useful to identify putative functional sites.

  12. 21 CFR 862.1500 - Malic dehydrogenase test system.

    Science.gov (United States)

    2010-04-01

    ... Systems § 862.1500 Malic dehydrogenase test system. (a) Identification. A malic dehydrogenase test system is a device that is intended to measure the activity of the enzyme malic dehydrogenase in serum and... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Malic dehydrogenase test system. 862.1500 Section...

  13. Spaceflight exposure effects on transcription, activity, and localization of alcohol dehydrogenase in the roots of Arabidopsis thaliana

    Science.gov (United States)

    Porterfield, D. M.; Matthews, S. W.; Daugherty, C. J.; Musgrave, M. E.

    1997-01-01

    Although considerable research and speculation have been directed toward understanding a plant's perception of gravity and the resulting gravitropic responses, little is known about the role of gravity-dependent physical processes in normal physiological function. These studies were conducted to determine whether the roots of plants exposed to spaceflight conditions may be experiencing hypoxia. Arabidopsis thaliana (L.) Heynh. plants were grown in agar medium during 6 or 11 d of spaceflight exposure on shuttle missions STS-54 (CHROMEX-03) and STS-68 (CHROMEX-05), respectively. The analysis included measurement of agar redox potential and root alcohol dehydrogenase (ADH) activity, localization, and expression. ADH activity increased by 89% as a result of spaceflight exposure for both CHROMEX-03 and -05 experiments, and ADH RNase protection assays revealed a 136% increase in ADH mRNA. The increase in ADH activity associated with the spaceflight roots was realized by a 28% decrease in oxygen availability in a ground-based study; however, no reduction in redox potential was observed in measurements of the spaceflight bulk agar. Spaceflight exposure appears to effect a hypoxic response in the roots of agar-grown plants that may be caused by changes in gravity-mediated fluid and/or gas behavior.

  14. Field Trial of the CareStart Biosensor Analyzer for the Determination of Glucose-6-Phosphate Dehydrogenase Activity in Haiti.

    Science.gov (United States)

    Weppelmann, Thomas A; von Fricken, Michael E; Wilfong, Tara D; Aguenza, Elisa; Philippe, Taina T; Okech, Bernard A

    2017-10-01

    Throughout many developing and tropical countries around the world, malaria remains a significant threat to human health. One barrier to malaria elimination is the ability to safely administer primaquine chemotherapy for the radical cure of malaria infections in populations with a high prevalence of glucose-6-phosphate dehydrogenase (G6PD) deficiency. In the current study, a field trial of the world's first quantitative, point-of-care assay for measuring G6PD activity was conducted in Haiti. The performance of the CareStart Biosensor Analyzer was compared with the gold standard spectrophotometric assay and genotyping of the G6PD allele in schoolchildren ( N = 343) from the Ouest Department of Haiti. In this population, 19.5% of participants (67/343) had some form of G6PD deficiency (< 60% residual activity) and 9.9% (34/343) had moderate-to-severe G6PD deficiency (< 30% residual activity). Overall, 18.95% of participants had the presence of the A-allele (65/343) with 7.87% (27/343) considered at high risk for drug-induced hemolysis (hemizygous males and homozygous females). Compared with the spectrophotometric assay, the sensitivity and specificity to determine participants with < 60% residual activity were 53.7% and 94.6%, respectively; for participants with 30% residual activity, the sensitivity and specificity were 5.9% and 99.7%, respectively. The biosensor overestimated the activity in deficient individuals and underestimated it in participants with normal G6PD activity, indicating the potential for a systematic measurement error. Thus, we suggest that the current version of the biosensor lacks adequate sensitivity and should be improved prior to its use as a point-of-care diagnostic for G6PD deficiency.

  15. Structure-oriented substrate specificity engineering of aldehyde-deformylating oxygenase towards aldehydes carbon chain length.

    Science.gov (United States)

    Bao, Luyao; Li, Jian-Jun; Jia, Chenjun; Li, Mei; Lu, Xuefeng

    2016-01-01

    Aldehyde-deformylating oxygenase (ADO) is an important enzyme involved in the biosynthetic pathway of fatty alk(a/e)nes in cyanobacteria. However, ADO exhibits quite low chain-length specificity with respect to the substrates ranging from C4 to C18 aldehydes, which is not suitable for producing fuels with different properties or different chain lengths. Based on the crystal structures of cADOs (cyanobacterial ADO) with substrate analogs bound, some amino acids affecting the substrate specificity of cADO were identified, including the amino acids close to the aldehyde group and the hydrophobic tail of the substrate and those along the substrate channel. Using site-directed mutagenesis, selected amino acids were replaced with bulky ones introducing steric hindrance to the binding pocket via large functional groups. All mutants were overexpressed, purified and kinetically characterized. All mutants, except F87Y, displayed dramatically reduced activity towards C14,16,18 aldehydes. Notably, the substrate preferences of some mutants towards different chain-length substrates were enhanced: I24Y for n-heptanal, I27F for n-decanal and n-dodecanal, V28F for n-dodecanal, F87Y for n-decanal, C70F for n-hexanal, A118F for n-butanal, A121F for C4,6,7 aldehydes, V184F for n-dodecanal and n-decanal, M193Y for C6-10 aldehydes and L198F for C7-10 aldehydes. The impact of the engineered cADO mutants on the change of the hydrocarbon profile was demonstrated by co-expressing acyl-ACP thioesterase BTE, fadD and V184F in E. coli, showing that n-undecane was the main fatty alkane. Some amino acids, which can control the chain-length selectivity of substrates of cADO, were identified. The substrate specificities of cADO were successfully changed through structure-guided protein engineering, and some mutants displayed different chain-length preference. The in vivo experiments of V184F in genetically engineered E. coli proved the importance of engineered cADOs on the distribution of the

  16. Age-dependent decrease in the activity of succinic dehydrogenase in rat CA1 pyramidal cells: a quantitative cytochemical study.

    Science.gov (United States)

    Bertoni-Freddari, C; Fattoretti, P; Caselli, U; Paoloni, R; Meier-Ruge, W

    1996-09-09

    A computer-assisted morphometric study has been carried out on the ultrastructure of perikaryal CA1 pyramidal cell mitochondria positive to the copper ferricyanide cytochemical reaction for succinic dehydrogenase (SDH) in rats of 3, 12 and 23 months of age. The cytoplasmic volume fraction occupied by the positive mitochondria (Volume density: Vv), the number of organelles/micron 3 of CA1 pyramidal cell cytoplasm (Numerical density: Nv) and the average mitochondrial volume (V) were automatically calculated by means of computer-assisted morphometry. Vv was significantly decreased in 23-month-old animals versus the other age groups. Nv was unchanged between 3 and 12 months of age, but was decreased to a significant extent in old animals. V did not undergo significant changes in the three age groups taken into account. In the old animals the percent of organelles smaller than 0.16 micron 3 is above 20%, while in the young and adult groups the same size of mitochondria accounts for 7 and 3%, respectively. Thus, a reduction in the number of medium sized organelles appears to be responsible for the decrease in Vv due to age. Since SDH activity is known to support maximum rates of respiration, quantitative estimation of the active mitochondria provides information on the metabolic competence of the cells investigated when energy demand is high. In this context, our present findings document that a significant impairment in the efficiency to match actual energy provisions occurs in old CA1 pyramidal cells.

  17. Furanfurin and thiophenfurin: two novel tiazofurin analogues. Synthesis, structure, antitumor activity, and interactions with inosine monophosphate dehydrogenase.

    Science.gov (United States)

    Franchetti, P; Cappellacci, L; Grifantini, M; Barzi, A; Nocentini, G; Yang, H; O'Connor, A; Jayaram, H N; Carrell, C; Goldstein, B M

    1995-09-15

    The syntheses of furan and thiophene analogues of tiazofurin (furanfurin and thiophenfurin, respectively) are described. Direct stannic chloride-catalyzed C-glycosylation of ethyl 3-furan-carboxylate (6) or ethyl 3-thiophencarboxylate (18) with 1,2,3,5-tetra-O-acetyl-D-ribofuranose gave 2- and 5-glycosylated regioisomers, as a mixture of alpha- and beta-anomers, and the beta-2,5-diglycosylated derivatives. Deprotection of ethyl 5-(2,3,5-tri-O-acetyl-beta-D-ribofuranosyl)furan-3-carboxylate (9 beta) and ethyl 5-(2,3,5-tri-O-acetyl-beta-D-ribofuranosyl)thiophene-3-carboxylate (20 beta) with sodium ethoxide afforded ethyl 5-beta-D-ribofuranosylfuran-3-carboxylate (12 beta) and ethyl 5-beta-D-ribofuranosylthiophene-3-carboxylate (23 beta) which were converted into 5-beta-D-ribofuranosylfuran-3-carboxamide (furanfurin, 4) and 5-beta-D-ribofuranosylthiophene-3-carboxamide (thiophenfurin, 5) by reaction with ammonium hydroxide. The anomeric configuration and the site of glycosylation were established by 1H-NMR and proton-proton nuclear Overhauser effect difference spectroscopy. The structure of compound 23 beta was confirmed by X-ray crystallography. Thiophenfurin was found to be cytotoxic in vitro toward murine lymphocytic leukemia P388 and L1210, human myelogenous leukemia K562, human promyelocytic leukemia HL-60, human colon adenocarcinoma LoVo, and B16 melanoma at concentrations similar to that of tiazofurin. In the same test furanfurin proved to be inactive. Thiophenfurin was found active in vivo in BD2F1 mice inoculated with L1210 cells with a % T/C of 168 at 25 mg/kg. K562 cells incubation with thiophenfurin resulted in inhibition of inosine monophosphate (IMP) dehydrogenase (63%) and an increase in IMP pools (6-fold) with a concurrent decrease in GTP levels (42%). Incubation of adenosine-labeled K562 cells with tiazofurin, thiophenfurin, and furanfurin resulted in a 2-fold higher NAD analogue formulation by thiophenfurin than by tiazofurin. Furanfurin was

  18. The promoter activity of isovaleryl-CoA dehydrogenase-encoding gene (ivdA) from Aspergillus oryzae is strictly repressed by glutamic acid.

    Science.gov (United States)

    Yamashita, Nobuo; Sakamoto, Kazutoshi; Yamada, Osamu; Akita, Osamu; Nishimura, Akira

    2007-06-01

    We cloned the isovaleryl-CoA dehydrogenase (IVD)-encoding gene from Aspergillus oryzae. The promoter of ivdA was subjected to beta-glucuronidase (GUS) reporter assays in which certain amino acids were used as a major carbon source. L-leucine most strongly induced GUS-activity, while in the case of L-glutamate, significantly low activity was found, indicating that ivdA transcription was strongly repressed by glutamic acid.

  19. In vitro assessment of human airway toxicity from major aldehydes in automotive emissions

    Energy Technology Data Exchange (ETDEWEB)

    Grafstroem, R.C. [Karolinska Inst., Stockholm (Sweden). Inst. of Environmental Medicine

    1997-09-01

    Automotive exhausts can significantly contribute to the levels of reactive aldehydes, including formaldehyde, acetaldehyde and acrolein, in urban air. The use of alcohols as an alternative fuel for gasoline or diesel may further increase these emissions. Since it is unclear if aldehyde inhalation may induce pathological states, including cancer, in human airways, the toxic properties of the above-mentioned aldehydes were studied in cultured target cell types. Each aldehyde modified vital cellular functions in a dose-dependent manner, and invariably inhibited growth and induced abnormal terminal differentiation. Decreases of cellular thiols and increases of intracellular Ca{sup 2+} were observed, and moreover, variable types and amounts of short-lived or persistent genetic damage were induced. The concentrations required for specified levels of a particular type of injury varied up to 10000-fold among the aldehydes. Overall, distinctive patterns of cytopathological activity were observed, which differed both qualitatively and quantitatively among the aldehydes. Finally, aldehydes inhibited DNA repair processes and increased cytotoxicity and mutagenesis in synergy with other known toxicants, indicating that aldehydes may also enhance damage by other constituents in automotive exhausts. In summary, the aldehydes, notably {sup m}u{sup M}-mM formaldehyde, caused pathological effects and induced mechanisms that relate to acute toxicity and cancer development in airway epithelial cells. Since `no-effect` levels may not exist for carcinogenic agents, the overall results support a need for elimination of aldehydes in automotive exhausts. 41 refs

  20. Mild reduction in the activity of the α-ketoglutarate dehydrogenase complex elevates GABA shunt and glycolysis

    Science.gov (United States)

    Shi, Qingli; Risa, Øystein; Sonnewald, Ursula; Gibson, Gary E.

    2009-01-01

    Diminished energy metabolism and reduced activity of brain α-ketoglutarate dehydrogenase complex (KGDHC) occur in a number of neurodegenerative diseases. The relation between diminished KGDHC activity and altered energy metabolism is unknown. The present study tested whether a reduction in the KGDHC activity would alter cellular metabolism by comparing metabolism of [U-13C]glucose in a human embryonic kidney (HEK) cell line (E2k100) to one in which the KGDHC activity was about 70% of control (E2k67). After a two hours incubation of the cells with [U-13C]glucose, the E2k67 cells showed a greater increase in 13C labeling of alanine compared to the E2k100 cells. This suggested an increase in glycolysis. Furthermore, an increase in labeled lactate after 12 hr incubation supported the suggestion of an increased glycolysis in the E2k67 cells. Increased GABA shunt in the E2k67 cells was indicated by increased 13C labeling of GABA at both 2 and 12 hr compared to the control cells. GABA concentration as determined by HPLC was also increased in the E2k67 cells compared to the control cells. However, the GABA shunt was not sufficient to normalize metabolism in the E2k67 cells compared to control at 2 or 12 hours. However, by 24 hr metabolism had normalized (ie. labeling was similar in E2k67 and E2k100). Thus, the data are consistent with an enhanced glycolysis and GABA shunt in response to a mild reduction in KGDHC. Our findings indicate that a mild change in KGDHC activity can lead to large changes in metabolism. The changes may maintain normal energy metabolism but make the cells more vulnerable to perturbations such as occur with oxidants. PMID:19393030

  1. In vitro effects of metals and pesticides on dehydrogenase activity in ...

    African Journals Online (AJOL)

    AJB SERVER

    2007-01-04

    Jan 4, 2007 ... microbial activity and anaerobic conditions (Chapman et al., 1981). Application of pesticides to agricultural soils may affect soil biological activity in a variety of ways (Shetty and. Magu, 1997, 1998). Herbicides may have negative eff- ects on the growth of rhizobia (Clark and Mahanty, 1991;. Mårtensson ...

  2. The effects of chemical and radioactive properties of Tl-201 on human erythrocyte glucose 6-phosphate dehydrogenase activity

    International Nuclear Information System (INIS)

    Sahin, Ali; Senturk, Murat; Ciftci, Mehmet; Varoglu, Erhan; Kufrevioglu, Omer Irfan

    2010-01-01

    Aim: The inhibitory effects of thallium-201 ( 201 Tl) solution on human erythrocyte glucose 6-phosphate dehydrogenase (G6PD) activity were investigated. Methods: For this purpose, erythrocyte G6PD was initially purified 835-fold at a yield of 41.7% using 2',5'-Adenosine diphosphate sepharose 4B affinity gel chromatography. The purification was monitored by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, which showed a single band for the final enzyme preparation. The in vitro and in vivo effects of the 201 Tl solution including Tl + , Fe +3 and Cu +2 metals and the in vitro effects of the radiation effect of the 201 Tl solution and non-radioactive Tl + , Fe +3 and Cu +2 metals on human erythrocyte G6PD enzyme were studied. Enzyme activity was determined with the Beutler method at 340 nm using a spectrophotometer. All purification procedures were carried out at +4 deg. C. Results: 201 Tl solution and radiation exposure had inhibitory effects on the enzyme activity. IC 50 value of 201 Tl solution was 36.86 μl ([Tl + ]: 0.0036 μM, [Cu +2 ]: 0.0116 μM, [Fe +3 ]: 0.0132 μM), of human erythrocytes G6PD. Seven human patients were also used for in vivo studies of 201 Tl solution. Furthermore, non-radioactive Tl + , Fe +3 and Cu +2 were found not to have influenced the enzyme in vitro. Conclusion: Human erythrocyte G6PD activity was inhibited by exposure for up to 10 minutes to 0.057 mCi/kg 201 Tl solution. It was detected in in vitro and in vivo studies that the human erythrocyte G6PD enzyme is inhibited due to the radiation effect of 201 Tl solution.

  3. Impact of cyclometalated ruthenium(II) complexes on lactate dehydrogenase activity and cytotoxicity in gastric and colon cancer cells.

    Science.gov (United States)

    Rico Bautista, Hugo; Saavedra Díaz, Rafael Omar; Shen, Longzhu Q; Orvain, Christophe; Gaiddon, Christian; Le Lagadec, Ronan; Ryabov, Alexander D

    2016-10-01

    Lactate dehydrogenase (LDH) is a redox enzyme often overexpressed in cancer cells allowing their survival in stressful metabolic tumor environment. Ruthenium(II) complexes have been shown to impact on the activity of purified horseradish peroxidase and glucose oxidase but the physiological relevance remains unclear. In this study we investigated how ruthenium complexes impact on the activity of LDH in vitro and in cancer cells and performed a comparative study using polypyridine ruthenium(II) complex [Ru(bpy) 3 ] 2+ (1) and its structurally related cyclometalated 2-phenylpyridinato counterpart [Ru(phpy)(bpy) 2 ] + (2) (bpy=2,2'-bipyridine, phpyH=2-phenylpyridine). We show that the cytotoxicity in gastric and colon cancer cells induced by 2 is significantly higher compared to 1. The kinetic inhibition mechanisms on purified LDH and the corresponding inhibition constants K i or i 0.5 values were calculated. Though complexes 1 and 2 are structurally very similar (one Ru-C bond in 2 replaces one Ru-N bond in 1), their inhibition modes are different. Cyclometalated complex 2 behaves exclusively as a non-competitive inhibitor of LDH from rabbit muscle (LDH rm) , strongly suggesting that 2 does not interact with LDH in the vicinities of either lactate/pyruvate or NAD + /NADH binding sites. Sites of interaction of 1 and 2 with LDH rm were revealed theoretically through computational molecular docking. Inhibition of LDH activity by 2 was confirmed in cancer cells. Altogether, these results revealed an inhibition of LDH activity by ruthenium complex through a direct interaction structurally tuned by a Ru-C bond. Copyright © 2016 Elsevier Inc. All rights reserved.

  4. The Spatial Variability of Soil Dehydrogenase Activity: A Survey in Urban Soils

    OpenAIRE

    Kizilkaya, Ridvan; Aşkin, Tayfun

    2007-01-01

    Information on soil microorganisms and their activity used to determine microbiological characteristics are very important for soil quality and productivity. Studies of enzyme activities provide information on the biochemical processes occurring in soil. There is growing evidence that soil biological parameters may be potential and sensitive indicators of soil ecological conditions and soil management. Soil microbiological parameters may be evaluated statistically due to application of geosta...

  5. Synthesis, conformational analysis, and biological activity of new analogues of thiazole-4-carboxamide adenine dinucleotide (TAD) as IMP dehydrogenase inhibitors.

    Science.gov (United States)

    Franchetti, Palmarisa; Cappellacci, Loredana; Pasqualini, Michela; Petrelli, Riccardo; Jayaprakasan, Vetrichelvan; Jayaram, Hiremagalur N; Boyd, Donald B; Jain, Manojkumar D; Grifantini, Mario

    2005-03-15

    Thiazole-4-carboxamide adenine dinucleotide (TAD) analogues T-2'-MeAD (1) and T-3'-MeAD (2) containing, respectively, a methyl group at the ribose 2'-C-, and 3'-C-position of the adenosine moiety, were prepared as potential selective human inosine monophosphate dehydrogenase (IMPDH) type II inhibitors. The synthesis of heterodinucleotides was carried out by CDI-catalyzed coupling reaction of unprotected 2'-C-methyl- or 3'-C-methyl-adenosine 5'-monophosphate with 2',3'-O-isopropylidene-tiazofurin 5'-monophosphate, and then deisopropylidenation. Biological evaluation of dinucleotides 1 and 2 as inhibitors of recombinant human IMPDH type I and type II resulted in a good activity. Inhibition of both isoenzymes by T-2'-MeAD and T-3'-MeAD was noncompetitive with respect to NAD substrate. Binding of T-3'-MeAD was comparable to that of parent compound TAD, while T-2'-MeAD proved to be a weaker inhibitor. However, no significant difference was found in inhibition of the IMPDH isoenzymes. T-2'-MeAD and T-3'-MeAD were found to inhibit the growth of K562 cells (IC(50) 30.7 and 65.0muM, respectively).

  6. Preclinical activity of FF-10501-01, a novel inosine-5'-monophosphate dehydrogenase inhibitor, in acute myeloid leukemia.

    Science.gov (United States)

    Yang, Hui; Fang, Zhihong; Wei, Yue; Bohannan, Zachary S; Gañán-Gómez, Irene; Pierola, Ana Alfonso; Paradiso, Linda J; Iwamura, Hiroyuki; Garcia-Manero, Guillermo

    2017-08-01

    FF-10501-01 is a selective inosine monophosphate dehydrogenase (IMPDH) inhibitor that has shown activity in cancer cell lines. We studied whether FF-10501-01 is effective in targeting a variety of hypomethylating agent (HMA)-sensitive and -resistant acute myelogenous leukemia (AML) cell lines. We treated multiple cell lines (including HMA-resistant cells) with FF-10501-01 and analyzed proliferation, apoptosis, and cell cycle status. We also assessed HMA-FF-10501-01 combinations and the ability of extracellular guanosine to rescue cell proliferation in FF-10501-01-treated cells. We performed high-performance liquid chromatography (HPLC) to study guanine nucleotide levels in treated and untreated cells. Finally, we studied the effects of FF-10501-01 in fresh peripheral blood cells taken from AML patients. FF-10501-01 showed a strong dose-dependent effect on proliferation and induced apoptosis at approximately 30μM. The effects of FF-10501-01 treatment on cell cycle status were variable, with no statistically significant trends. Guanosine rescued proliferation in FF-10501-01-treated cells, and HPLC results showed significant decreases in phosphorylated guanosine levels in MOLM13 cells. FF-10501-01 effectively reduced proliferation at concentrations of 300μM and above in 3 primary AML samples. FF-10501-01 effectively induces AML cell death and reduces AML peripheral blood cell proliferation by targeting guanine nucleotide biosynthesis regardless of HMA resistance status. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. High activity of mitochondrial glycerophosphate dehydrogenase and glycerophosphate-dependent ROS production in prostate cancer cell lines

    International Nuclear Information System (INIS)

    Chowdhury, Subir K.R.; Gemin, Adam; Singh, Gurmit

    2005-01-01

    Most malignant cells are highly glycolytic and produce high levels of reactive oxygen species (ROS) compared to normal cells. Mitochondrial glycerophosphate dehydrogenase (mGPDH) participates in the reoxidation of cytosolic NADH by delivering reducing equivalents from this molecule into the electron transport chain, thus sustaining glycolysis. Here, we investigate the role of mGPDH in maintaining an increased rate of glycolysis and evaluate glycerophosphate-dependent ROS production in prostate cancer cell lines (LNCaP, DU145, PC3, and CL1). Immunoblot, polarographic, and spectrophotometric analyses revealed that mGPDH abundance and activity was significantly elevated in prostate cancer cell lines when compared to the normal prostate epithelial cell line PNT1A. Furthermore, both the glycolytic capacity and glycerophosphate-dependent ROS production was increased 1.68- to 4.44-fold and 5- to 7-fold, respectively, in prostate cancer cell lines when compared to PNT1A cells. Overall, these data demonstrate that mGPDH is involved in maintaining a high rate of glycolysis and is an important site of electron leakage leading to ROS production in prostate cancer cells

  8. Redox Balance in Lactobacillus reuteri DSM20016: Roles of Iron-Dependent Alcohol Dehydrogenases in Glucose/ Glycerol Metabolism.

    Directory of Open Access Journals (Sweden)

    Lu Chen

    Full Text Available Lactobacillus reuteri, a heterofermentative bacterium, metabolizes glycerol via a Pdu (propanediol-utilization pathway involving dehydration to 3-hydroxypropionaldehyde (3-HPA followed by reduction to 1,3-propandiol (1,3-PDO with concomitant generation of an oxidized cofactor, NAD+ that is utilized to maintain cofactor balance required for glucose metabolism and even for oxidation of 3-HPA by a Pdu oxidative branch to 3-hydroxypropionic acid (3-HP. The Pdu pathway is operative inside Pdu microcompartment that encapsulates different enzymes and cofactors involved in metabolizing glycerol or 1,2-propanediol, and protects the cells from the toxic effect of the aldehyde intermediate. Since L. reuteri excretes high amounts of 3-HPA outside the microcompartment, the organism is likely to have alternative alcohol dehydrogenase(s in the cytoplasm for transformation of the aldehyde. In this study, diversity of alcohol dehydrogenases in Lactobacillus species was investigated with a focus on L. reuteri. Nine ADH enzymes were found in L. reuteri DSM20016, out of which 3 (PduQ, ADH6 and ADH7 belong to the group of iron-dependent enzymes that are known to transform aldehydes/ketones to alcohols. L. reuteri mutants were generated in which the three ADHs were deleted individually. The lagging growth phenotype of these deletion mutants revealed that limited NAD+/NADH recycling could be restricting their growth in the absence of ADHs. Notably, it was demonstrated that PduQ is more active in generating NAD+ during glycerol metabolism within the microcompartment by resting cells, while ADH7 functions to balance NAD+/NADH by converting 3-HPA to 1,3-PDO outside the microcompartment in the growing cells. Moreover, evaluation of ADH6 deletion mutant showed strong decrease in ethanol level, supporting the role of this bifuctional alcohol/aldehyde dehydrogenase in ethanol production. To the best of our knowledge, this is the first report revealing both internal and

  9. Myricetin is a novel inhibitor of human inosine 5′-monophosphate dehydrogenase with anti-leukemia activity

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Huiling; Hu, Qian; Wang, Jingyuan; Liu, Zehui; Wu, Dang [Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, 130 Mei Long Road, Shanghai 200237 (China); Lu, Weiqiang, E-mail: wqlu@bio.ecnu.edu.cn [Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, 500 Dongchuan Road, Shanghai 200241 (China); Huang, Jin, E-mail: huangjin@ecust.edu.cn [Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, 130 Mei Long Road, Shanghai 200237 (China)

    2016-09-02

    Human inosine 5′-monophosphate dehydrogenase (hIMPDH) is a rate-limiting enzyme in the de novo biosynthetic pathway of purine nucleotides, playing crucial roles in cellular proliferation, differentiation, and transformation. Dysregulation of hIMPDH expression and activity have been found in a variety of human cancers including leukemia. In this study, we found that myricetin, a naturally occurring phytochemical existed in berries, wine and tea, was a novel inhibitor of human type 1 and type 2 IMPDH (hIMPDH1/2) with IC{sub 50} values of 6.98 ± 0.22 μM and 4.10 ± 0.14 μM, respectively. Enzyme kinetic analysis using Lineweaver-Burk plot revealed that myricetin is a mix-type inhibitor for hIMPDH1/2. Differential scanning fluorimetry and molecular docking simulation data demonstrate that myricetin is capable of binding with hIMPDH1/2. Myricetin treatment exerts potent anti-proliferative and pro-apoptotic effects on K562 human leukemia cells in a dose-dependent manner. Importantly, cytotoxicity of myricetin on K562 cells were markedly attenuated by exogenous addition of guanosine, a salvage pathway of maintaining intracellular pool of guanine nucleotides. Taking together, these results indicate that natural product myricetin exhibits potent anti-leukemia activity by interfering with purine nucleotides biosynthetic pathway through the suppression of hIMPDH1/2 catalytic activity. - Highlights: • Myricetin, a common dietary flavonoid, is a novel inhibitor of hIMPDH1/2. • Myricetin directly binds with hIMPDH1/2 and induces cell cycle arrest and apoptosis of leukemia cells. • The cytotoxicity of myricetin on K562 cells is markedly attenuated by exogenous addition of guanosine.

  10. Hypoxia-induced glucose-6-phosphate dehydrogenase overexpression and -activation in pulmonary artery smooth muscle cells: implication in pulmonary hypertension

    Science.gov (United States)

    Chettimada, Sukrutha; Gupte, Rakhee; Rawat, Dhwajbahadur; Gebb, Sarah A.; McMurtry, Ivan F.

    2014-01-01

    Severe pulmonary hypertension is a debilitating disease with an alarmingly low 5-yr life expectancy. Hypoxia, one of the causes of pulmonary hypertension, elicits constriction and remodeling of the pulmonary arteries. We now know that pulmonary arterial remodeling is a consequence of hyperplasia and hypertrophy of pulmonary artery smooth muscle (PASM), endothelial, myofibroblast, and stem cells. However, our knowledge about the mechanisms that cause these cells to proliferate and hypertrophy in response to hypoxic stimuli is still incomplete, and, hence, the treatment for severe pulmonary arterial hypertension is inadequate. Here we demonstrate that the activity and expression of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the pentose phosphate pathway, are increased in hypoxic PASM cells and in lungs of chronic hypoxic rats. G6PD overexpression and -activation is stimulated by H2O2. Increased G6PD activity contributes to PASM cell proliferation by increasing Sp1 and hypoxia-inducible factor 1α (HIF-1α), which directs the cells to synthesize less contractile (myocardin and SM22α) and more proliferative (cyclin A and phospho-histone H3) proteins. G6PD inhibition with dehydroepiandrosterone increased myocardin expression in remodeled pulmonary arteries of moderate and severe pulmonary hypertensive rats. These observations suggest that altered glucose metabolism and G6PD overactivation play a key role in switching the PASM cells from the contractile to synthetic phenotype by increasing Sp1 and HIF-1α, which suppresses myocardin, a key cofactor that maintains smooth muscle cell in contractile state, and increasing hypoxia-induced PASM cell growth, and hence contribute to pulmonary arterial remodeling and pathogenesis of pulmonary hypertension. PMID:25480333

  11. High fat fed heart failure animals have enhanced mitochondrial function and acyl-coa dehydrogenase activities

    Science.gov (United States)

    We have previously shown that administration of high fat in heart failure (HF) increased mitochondrial respiration and did not alter left ventricular (LV) function. PPARalpha is a nuclear transcription factor that activates expression of genes involved in fatty acid uptake and utilization. We hypoth...

  12. Direct enzyme assay evidence confirms aldehyde reductase function of Ydr541cp and Ygl039wp from Saccharomyces cerevisiae

    Science.gov (United States)

    Aldehyde reductase gene ARI1 is a recently characterized member of intermediate subfamily under SDR (short-chain dehydrogenase/reductase) superfamily that revealed mechanisms of in situ detoxification of furfural and HMF for tolerance of Saccharomyces cerevisiae. Uncharacterized open reading frames ...

  13. Mild reductions in cytosolic NADP-dependent isocitrate dehydrogenase activity result in lower amino acid contents and pigmentation without impacting growth.

    Science.gov (United States)

    Sulpice, Ronan; Sienkiewicz-Porzucek, Agata; Osorio, Sonia; Krahnert, Ina; Stitt, Mark; Fernie, Alisdair R; Nunes-Nesi, Adriano

    2010-10-01

    Transgenic tomato (Solanum lycopersicum) plants were generated targeting the cytosolic NADP-dependent isocitrate dehydrogenase gene (SlICDH1) via the RNA interference approach. The resultant transformants displayed a relatively mild reduction in the expression and activity of the target enzyme in the leaves. However, biochemical analyses revealed that the transgenic lines displayed a considerable shift in metabolism, being characterized by decreases in the levels of the TCA cycle intermediates, total amino acids, photosynthetic pigments, starch and NAD(P)H. The plants showed little change in photosynthesis with the exception of a minor decrease in maximum photosynthetic efficiency (F (v)/F (m)), and a small decrease in growth compared to the wild type. These results reveal that even small changes in cytosolic NADP-dependent isocitrate dehydrogenase activity lead to noticeable alterations in the activities of enzymes involved in primary nitrate assimilation and in the synthesis of 2-oxoglutarate derived amino acids. These data are discussed within the context of current models for the role of the various isoforms of isocitrate dehydrogenase within plant amino acid metabolism.

  14. The effect of ammonium ions on the activity of glutamate dehydrogenase, alanine aminotransferase and aspartate aminotransferase in Cucumis sativus L. seedlings

    Directory of Open Access Journals (Sweden)

    Genowefa Kubiak-Dobosz

    2014-01-01

    Full Text Available Changes in the activity of glutamate dehydrogenase (GDH, alanine aminotransferase (GPT and aspartate aminotransferase (GOT were studied in various organs of Cucumis sativus L. seedlings in relation to the uptake of mineral nitrogen (in form of N03- or NH4+ from the medium. Activity of GDH, GPT, and GOT was higher in young leaves and roots of cucumber seedlings if the plants developed- in an ammonium medium. No similar changes of aminotransferases activity were noted in the cotyledons. Factors affecting varying effect of ammonium ions upon GPT and GOT activity are discussed for particular organs of cucumber seedlings.

  15. Peptide aldehyde inhibitors of bacterial peptide deformylases.

    Science.gov (United States)

    Durand, D J; Gordon Green, B; O'Connell, J F; Grant, S K

    1999-07-15

    Bacterial peptide deformylases (PDF, EC 3.5.1.27) are metalloenzymes that cleave the N-formyl groups from N-blocked methionine polypeptides. Peptide aldehydes containing a methional or norleucinal inhibited recombinant peptide deformylase from gram-negative Escherichia coli and gram-positive Bacillus subtilis. The most potent inhibitor was calpeptin, N-CBZ-Leu-norleucinal, which was a competitive inhibitor of the zinc-containing metalloenzymes, E. coli and B. subtilis PDF with Ki values of 26.0 and 55.6 microM, respectively. Cobalt-substituted E. coli and B. subtilis deformylases were also inhibited by these aldehydes with Ki values for calpeptin of 9.5 and 12.4 microM, respectively. Distinct spectral changes were observed upon binding of calpeptin to the Co(II)-deformylases, consistent with the noncovalent binding of the inhibitor rather than the formation of a covalent complex. In contrast, the chelator 1,10-phenanthroline caused the time-dependent inhibition of B. subtilis Co(II)-PDF activity with the loss of the active site metal. The fact that calpeptin was nearly equipotent against deformylases from both gram-negative and gram-positive bacterial sources lends further support to the idea that a single deformylase inhibitor might have broad-spectrum antibacterial activity. Copyright 1999 Academic Press.

  16. Cloning, expression and characterization of an aryl-alcohol dehydrogenase from the white-rot fungus Phanerochaete chrysosporium strain BKM-F-1767

    Directory of Open Access Journals (Sweden)

    Yang Dong-Dong

    2012-06-01

    Full Text Available Abstract Background The white-rot fungus Phanerochaete chrysosporium is among the small group of fungi that can degrade lignin to carbon dioxide while leaving the crystalline cellulose untouched. The efficient lignin oxidation system of this fungus requires cyclic redox reactions involving the reduction of aryl-aldehydes to the corresponding alcohols by aryl-alcohol dehydrogenase. However, the biochemical properties of this enzyme have not been extensively studied. These are of most interest for the design of metabolic engineering/synthetic biology strategies in the field of biotechnological applications of this enzyme. Results We report here the cloning of an aryl-alcohol dehydrogenase cDNA from the white-rot fungus Phanerochaete chrysosporium, its expression in Escherichia coli and the biochemical characterization of the encoded GST and His6 tagged protein. The purified recombinant enzyme showed optimal activity at 37°C and at pH 6.4 for the reduction of aryl- and linear aldehydes with NADPH as coenzyme. NADH could also be the electron donor, while having a higher Km (220 μM compared to that of NADPH (39 μM. The purified recombinant enzyme was found to be active in the reduction of more than 20 different aryl- and linear aldehydes showing highest specificity for mono- and dimethoxylated Benzaldehyde at positions 3, 4, 3,4 and 3,5. The enzyme was also capable of oxidizing aryl-alcohols with NADP + at 30°C and an optimum pH of 10.3 but with 15 to 100-fold lower catalytic efficiency than for the reduction reaction. Conclusions In this work, we have characterized the biochemical properties of an aryl-alcohol dehydrogenase from the white-rot fungus Phanerochaete chrysosporium. We show that this enzyme functions in the reductive sense under physiological conditions and that it displays relatively large substrate specificity with highest activity towards the natural compound Veratraldehyde.

  17. A novel NADPH-dependent aldehyde reductase gene from Vigna radiata confers resistance to the grapevine fungal toxin eutypine.

    Science.gov (United States)

    Guillén, P; Guis, M; Martínez-Reina, G; Colrat, S; Dalmayrac, S; Deswarte, C; Bouzayen, M; Roustan, J P; Fallot, J; Pech, J C; Latché, A

    1998-11-01

    Eutypine, 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzyl aldehyde, is a toxin produced by Eutypa lata, the causal agent of eutypa dieback of grapevines. It has previously been demonstrated that tolerance of some cultivars to this disease was correlated with their capacity to convert eutypine to the corresponding alcohol, eutypinol, which lacks phytotoxicity. We have thus purified to homogeneity a protein from Vigna radiata that exhibited eutypine-reducing activity and have isolated the corresponding cDNA. This encodes an NADPH-dependent reductase of 36 kDa that we have named Vigna radiata eutypine-reducing enzyme (VR-ERE), based on the capacity of a recombinant form of the protein to reduce eutypine into eutypinol. The strongest homologies (86.8%) of VR-ERE at the amino acid level were found with CPRD14, a drought-inducible gene of unknown function, isolated from Vigna unguiculata and with an aromatic alcohol dehydrogenase (71.7%) from Eucalyptus gunnii. Biochemical characterization of VR-ERE revealed that a variety of compounds containing an aldehyde group can act as substrates. However, the highest affinity was observed with 3-substituted benzaldehydes. Expression of a VR-ERE transgene in Vitis vinifera cells cultured in vitro conferred resistance to the toxin. This discovery opens up new biotechnological approaches for the generation of grapevines resistant to eutypa dieback.

  18. Effect of Follicular Fluid and Platelet-Activating Factor on Lactate Dehydrogenase C Expression in Human Asthenozoospermic Samples

    Directory of Open Access Journals (Sweden)

    Tahereh Esmaeilpour

    2014-01-01

    Full Text Available Background: Application of follicular fluid (FF and platelet-activating factor (PAF in artificial insemination improves sperm motility. Lactate dehydrogenase C (LDH-C is a key enzyme for sperm motility. In this study, the effects of FF and PAF on the sperm motility index and LDH-C expression were investigated. Moreover, LDH-C expression was compared between asthenozoospermic and normozoospermic samples. Methods: The expression of LDH-C was examined by quantitative real-time polymerase chain reaction (q-RT PCR and western blotting after it was treated with optimized concentrations of FF and PAF in twenty asthenozoospermic samples. Also, LDH-C expression was evaluated in five normozoospermic samples. Results: Samples with 75% FF and 100 nM of PAF had an increase in their percentages of progressive and slowly motile sperms and a decrease in their percentages of non-progressive and non-motile sperms. Moreover, LDH-C mRNA transcripts were not changed following PAF and FF treatment, and LDH-C protein was detected in highly progressive motile specimens treated with FF in the asthenozoospermic samples. Furthermore, LDH-C expression was more detectable in the normal sperms. Conclusion: Our results indicated that PAF had more beneficial effects than FF on sperm motility in the asthenozoospermic samples (P=0.0001, although the LDH-C expressions of the sperms were not changed significantly in both groups. We found no association between LDH-C expression and sperm motility after FF and PAF actions. This finding, however, requires further investigation. The fact that LDH-C protein was detected in the normozoospermic, but not asthenozoospermic, samples could be cited as a reason for the infertility in these patients.

  19. Elevated lactate dehydrogenase activity and increased cardiovascular mortality in the arsenic-endemic areas of southwestern Taiwan

    Energy Technology Data Exchange (ETDEWEB)

    Liao, Ya-Tang [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China); Genomics Research Center, Academia Sinica, Taiwan (China); Chen, Chien-Jen [Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China); Genomics Research Center, Academia Sinica, Taiwan (China); Li, Wan-Fen [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Hsu, Ling-I [Genomics Research Center, Academia Sinica, Taiwan (China); Tsai, Li-Yu; Huang, Yeou-Lih [Department of Medical Laboratory Science and Biotechnology, Kaohsiung Medical University, Taiwan (China); Sun, Chien-Wen [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Chen, Wei J., E-mail: wjchen@ntu.edu.tw [Graduate Institute of Epidemiology and Preventive Medicine, College of Public Health, National Taiwan University, Taiwan (China); Genetic Epidemiology Core Laboratory, National Taiwan University Center for Genomic Medicine, Taiwan (China); Wang, Shu-Li, E-mail: slwang@nhri.org.tw [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, Taiwan (China); Department of Public Health, College of Public Health, China Medical University, Taichung, Taiwan (China)

    2012-08-01

    Arsenic ingestion has been linked to increasing global prevalence of and mortality from cardiovascular disease (CVD); arsenic can be removed from drinking water to reduce related health effects. Lactate dehydrogenase (LDH) is used for the evaluation of acute arsenic toxicity in vivo and in vitro, but it is not validated for the evaluation of long-term, chronic arsenic exposure. The present study examined the long-term effect of chronic arsenic exposure on CVD and serum LDH levels, after consideration of arsenic metabolism capacity. A total of 380 subjects from an arseniasis-endemic area and 303 from a non-endemic area of southwestern Taiwan were recruited in 2002. Various urinary arsenic species were analyzed using high-performance liquid chromatography (HPLC) and hydride generation systems. Fasting serum was used for quantitative determination of the total LDH activity. A significant dose–response relationship was observed between arsenic exposure and LDH elevation, independent of urinary arsenic profiles (P < 0.001). Furthermore, abnormal LDH elevation was associated with CVD mortality after adjustment for Framingham risk scores for 10-year CVD and arsenic exposure (hazard ratio, 3.98; 95% confidence interval, 1.07–14.81). LDH was elevated in subjects with arsenic exposure in a dose-dependent manner. LDH is a marker of arsenic toxicity associated with CVD mortality. Results of this study have important implications for use in ascertaining long-term arsenic exposure risk of CVD. -- Highlights: ► We showed that arsenic exposure was correlated with LDH elevation. ► LDH elevation was related to arsenic methylation capacity. ► Abnormal LDH elevation can be a marker of susceptibility to CVD mortality.

  20. Atividade da 6-fosfogliconato desidrogenase em deficientes de glicose-6-fosfato desidrogenase Activity of 6-phosphogluconate dehydrogenase in glucose-6-phosphate dehydrogenase deficiency

    Directory of Open Access Journals (Sweden)

    Daniela B. Nicolielo

    2006-06-01

    to know better the actuation of these enzymes. The goal of this study was to evaluate the 6PGD enzymatic activity on a population with G6PD deficiency, to verify if there is an elevation of the activity of this enzyme, and try to correlate to a possible increase on the number of reticulocytes or the presence of alterations on red series. The research with 2657 male individuals detected 97 deficient for G6PD, which determined a 3.65% prevalence for the Bauru (SP region, with mean enzymatic activity of 1.74 UI.g Hb-1. min-1 at 37ºC, 14,4% of the normal G6PD activity. Mean 6PGD enzymatic activity was 9.5 UI.g Hb-1. min-1 at 37ºC, and was elevated in 47.4% of the G6PD deficient individuals. The result obtained did not confirm the hypothesis that the elevation of the 6PGD enzymatic activity, in G6PD deficient individuals, was due to the presence of an increase of reticulocytes in blood stream, age or erythrocytometric alterations that could denote anemia. The most plausible theory is that the auto-limited hemolysis, imposed by oxidative processes, preserves young erythrocytes that have an elevated enzymatic activity, as naturally these enzymes lose activity with cellular aging.

  1. High yields of active Thermus thermophilus proline dehydrogenase are obtained using maltose-binding protein as a solubility tag.

    NARCIS (Netherlands)

    Huijbers, M.M.E.; Berkel, van W.J.H.

    2015-01-01

    Proline dehydrogenase (ProDH) catalyzes the FAD-dependent oxidation of proline to ¿1-pyrroline-5-carboxylate, the first step of proline catabolism in many organisms. Next to being involved in a number of physiological processes, ProDH is of interest for practical applications because the proline

  2. Regenerative Capacity of Cacti Schlumbergera and Rhipsalidopsis in Relation to Endogenous Phytohormones, Cytokinin Oxidase/Dehydrogenase, and Peroxidase Activities

    Czech Academy of Sciences Publication Activity Database

    Sriskandarajah, S.; Prinsen, E.; Motyka, Václav; Dobrev, Petre; Serek, M.

    2006-01-01

    Roč. 25, č. 1 (2006), s. 79-88 ISSN 0721-7595 R&D Projects: GA ČR GA206/03/0313 Institutional research plan: CEZ:AV0Z50380511 Keywords : Cytokinin dehydrogenase * Cytokinin oxidase * Endogenous phytohormones * In vitro regeneration * Peroxidase * Rhipsalidopsis * Schlumbergera Subject RIV: EF - Botanics Impact factor: 2.107, year: 2006

  3. Effects of organic solvents on the enzyme activity of Trypanosoma cruzi glyceraldehyde-3-phosphate dehydrogenase in calorimetric assays

    DEFF Research Database (Denmark)

    Wiggers, Henrik; Cheleski, J; Zottis, A

    2007-01-01

    OH), in the isothermal titration calorimetry (ITC) kinetic assays for the catalyzed reaction of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from Trypanosoma cruzi. The solvent effects on T. cruzi GAPDH had not yet been studied. This enzyme was shown here to be affected by the organic solvents content up to 5...

  4. Determination of medium chain acyl-CoA dehydrogenase activity in cultured skin fibroblasts using mass spectrometry

    NARCIS (Netherlands)

    Niezen-Koning, K E; Chapman, T E; Mulder, I E; Smit, G P; Reijngoud, D J; Berger, R

    1991-01-01

    Medium chain acyl-CoA dehydrogenase deficiency, a defect of mitochondrial beta-oxidation, is one of the most frequently occurring among inborn errors of metabolism. We describe a rapid and sensitive gas chromatographic/mass spectrometric method allowing reliable assessment of medium chain acyl-CoA

  5. The activity of 11β-hydroxysteroid dehydrogenase type 2 enzyme and cortisol secretion in patients with adrenal incidentalomas.

    Science.gov (United States)

    Morelli, Valentina; Polledri, Elisa; Mercadante, Rosa; Zhukouskaya, Volha; Palmieri, Serena; Beck-Peccoz, Paolo; Spada, Anna; Fustinoni, Silvia; Chiodini, Iacopo

    2016-09-01

    In adrenal incidentaloma (AI) patients, beside the cortisol secretion, a different 11β-hydroxysteroid dehydrogenase type 2 (HSD11B2) activity, measurable by 24-h urinary cortisol/cortisone ratio (R-UFF/UFE) (the higher R-UFF/UFE the lower HSD11B2 activity), could influence the occurrence of the subclinical hypercortisolism (SH)-related complications (hypertension, type 2 diabetes, obesity). We evaluated whether in AI patients, UFF levels are associated to UFE levels, and the HSD11B2 activity to the complications presence. In 156 AI patients (93F, age 65.2 ± 9.5 years), the following were measured: serum cortisol after 1 mg-dexamethasone test (1 mg-DST), ACTH, UFF, UFE levels, and R-UFF/UFE (by liquid chromatography-tandem mass spectrometry), the latter was also evaluated in 63 matched-controls. We diagnosed SH (n = 22) in the presence of ≥2 among ACTH levels, and 1 mg-DST >83 nmol/L. Patients showed higher UFF levels and R-UFF/UFE than controls (75.9 ± 43.1 vs 54.4 ± 22.9 nmol/24 h and 0.26 ± 0.12 vs 0.20 ± 0.07, p levels (291 ± 91.1 vs 268 ± 61.5, p = 0.069). The R-UFF/UFE was higher in patients with high (h-UFF, n = 28, 0.41 ± 0.20) than in those with normal (n-UFF, 0.22 ± 0.10, p levels and in patients with SH than in those without SH (0.30 ± 0.12 vs 0.25 ± 0.12, p = 0.04). UFF levels were associated with R-UFF/UFE (r = 0.849, p levels in n-UFF patients but not in h-UFF patients, and it is not associated with the SH complications.

  6. Characterization of 17α-hydroxysteroid dehydrogenase activity (17α-HSD and its involvement in the biosynthesis of epitestosterone

    Directory of Open Access Journals (Sweden)

    Breton Rock

    2005-07-01

    Full Text Available Abstract Background Epi-testosterone (epiT is the 17α-epimer of testosterone. It has been found at similar level as testosterone in human biological fluids. This steroid has thus been used as a natural internal standard for assessing testosterone abuse in sports. EpiT has been also shown to accumulate in mammary cyst fluid and in human prostate. It was found to possess antiandrogenic activity as well as neuroprotective effects. So far, the exact pathway leading to the formation of epiT has not been elucidated. Results In this report, we describe the isolation and characterization of the enzyme 17α-hydroxysteroid dehydrogenase. The name is given according to its most potent activity. Using cells stably expressing the enzyme, we show that 17α-HSD catalyzes efficienty the transformation of 4-androstenedione (4-dione, dehydroepiandrosterone (DHEA, 5α-androstane-3,17-dione (5α-dione and androsterone (ADT into their corresponding 17α-hydroxy-steroids : epiT, 5-androstene-3β,17α-diol (epi5diol, 5α-androstane-17α-ol-3-one (epiDHT and 5α-androstane-3α,17α-diol (epi3α-diol, respectively. Similar to other members of the aldo-keto reductase family that possess the ability to reduce the keto-group into hydroxyl-group at different position on the steroid nucleus, 17α-HSD could also catalyze the transformation of DHT, 5α-dione, and 5α-pregnane-3,20-dione (DHP into 3α-diol, ADT and 5α-pregnane-3α-ol-20-one (allopregnanolone through its less potent 3α-HSD activity. We also have over-expressed the 17α-HSD in Escherichia coli and have purified it by affinity chromatography. The purified enzyme exhibits the same catalytic properties that have been observed with cultured HEK-293 stably transfected cells. Using quantitative Realtime-PCR to study tissue distribution of this enzyme in the mouse, we observed that it is expressed at very high levels in the kidney. Conclusion The present study permits to clarify the biosynthesis pathway of epiT. It

  7. Glucose-6-phosphate dehydrogenase and glutathione reductase activity in methemoglobin reduction by methylene blue and cyst amine: study on glucose-6-phosphate dehydrogenase-deficient individuals, on normal subjects and on riboflavin-treated subjects

    Directory of Open Access Journals (Sweden)

    Benedito Barraviera

    1988-10-01

    Full Text Available The authors have standardized methods for evaluation of the activity of the glucose-6-phosphate dehydrogenase and of glutathione reductase. The general principle of the first method was based on methemoglobin formation by sodium nitrite followed by stimulation of the glucose-6-phosphate dehydrogenase with methylene blue. Forty six adults (23 males and 23 females were studied. Subjects were not G6PD deficient and were aged 20 to 30 years. The results showed that methemoglobin reduction by methylene blue was 154.40 and 139.90 mg/min (p<0.05 for males and females, respectively, in whole blood, and 221.10 and 207.85 mg/min (n.s., respectively, in washed red cells. These data showed that using washed red cells and 0.7g% sodium nitrite concentration produced no differences between sexes and also shortened reading time for the residual amount of methemoglobin to 90 minutes. Glutathione reductase activity was evaluated on the basis of the fact that cystamine (a thiol agent binds to the SH groups of hemoglobin, forming complexes. These complexes are reversed by the action of glutathione reductase, with methemoglobin reduction occurring simultaneously with this reaction. Thirty two adults (16 males and 16 females were studied. Subjects were not G6PD deficient and were aged 20 to 30 years. Methemoglobin reduction by cystamine was 81.27 and 91.13 mg/min (p<0.01 for males and females, respectively. These data showed that using washed red cells and 0.1 M cystamine concentration permits a reading of the residual amount of methemoglobin at 180 minutes of incubation. Glutathione reductase activity was evaluated by methemoglobin reduction by cystamine in 14 females before and after treatment with 10 mg riboflavin per day for 8 days. The results were 73.69 and 94.26 jug/min (p<0.01 before and after treatment, showing that riboflavin treatment increase glutathione reductase activity even in normal individuals. Three Black G6PD-deficient individuals (2 males and 1

  8. Glucose-6-phosphate dehydrogenase

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003671.htm Glucose-6-phosphate dehydrogenase test To use the sharing features on this page, please enable JavaScript. Glucose-6-phosphate dehydrogenase (G6PD) is a protein that ...

  9. Xanthine dehydrogenase AtXDH1 from Arabidopsis thaliana is a potent producer of superoxide anions via its NADH oxidase activity.

    Science.gov (United States)

    Zarepour, Maryam; Kaspari, Katrin; Stagge, Stefan; Rethmeier, Ralf; Mendel, Ralf R; Bittner, Florian

    2010-02-01

    Xanthine dehydrogenase AtXDH1 from Arabidopsis thaliana is a key enzyme in purine degradation where it oxidizes hypoxanthine to xanthine and xanthine to uric acid. Electrons released from these substrates are either transferred to NAD(+) or to molecular oxygen, thereby yielding NADH or superoxide, respectively. By an alternative activity, AtXDH1 is capable of oxidizing NADH with concomitant formation of NAD(+) and superoxide. Here we demonstrate that in comparison to the specific activity with xanthine as substrate, the specific activity of recombinant AtXDH1 with NADH as substrate is about 15-times higher accompanied by a doubling in superoxide production. The observation that NAD(+) inhibits NADH oxidase activity of AtXDH1 while NADH suppresses NAD(+)-dependent xanthine oxidation indicates that both NAD(+) and NADH compete for the same binding-site and that both sub-activities are not expressed at the same time. Rather, each sub-activity is determined by specific conditions such as the availability of substrates and co-substrates, which allows regulation of superoxide production by AtXDH1. Since AtXDH1 exhibits the most pronounced NADH oxidase activity among all xanthine dehydrogenase proteins studied thus far, our results imply that in particular by its NADH oxidase activity AtXDH1 is an efficient producer of superoxide also in vivo.

  10. The Effects of Piper Sarmentosum Water Extract on the Expression and Activity of 11β-Hydroxysteroid Dehydrogenase Type 1 in the Bones with Excessive Glucocorticoids

    Directory of Open Access Journals (Sweden)

    Elvy Suhana Mohd Ramli

    2012-03-01

    Full Text Available Background: Long-term glucocorticoid therapy causes secondary osteoporosis leading to pathological fractures. Glucocorticoid action in bone is dependant upon the activity of 11β-hydroxysteroid dehydrogenase type 1 enzyme (11β-HSD1. Piper sarmentosum is a local herb that possesses the ability to inhibit 11-βHSD1 enzyme activity. We aimed to determine the effects of Piper sarmentosum water extract on 11-βHSD1 expressions and activity in the bones of glucocorticoid-treated adrenalectomized rats. Methods: Forty male Sprague–Dawley rats (200-250 g were used. Twenty-four animals were adrenalectomized and received intramuscular injection of dexamethasone (120 μg/kg/day. They were simultaneously administered with either Piper sarmentosum water extract (125 mg/kg/day, GCA (120 mg/kg/day or distilled water as vehicle by oral gavage for two months. Eight animals were sham-operated and given vehicle daily, i.e. intramuscular olive oil and oral distilled water. Results: Following two months treatment, dexamethasone-treated adrenalectomized rats had significantly lower 11β-HSD1 dehydrogenase activity and higher 11β-HSD1 expression in the femoral bones compared to the sham-operated and baseline group. The rats supplemented with Piper sarmentosum water extract had significantly higher 11β-HSD1 dehydrogenase activity and lower 11β-HSD1 expression in the bones. Conclusion: The results showed that Piper sarmentosum water extract had the ability to prevent glucocorcoticoid excess in the bones of glucocorticoid-treated adrenalectomized rats through the local modulation of 11β-HSD1 expression and activity, and may be used as prophylaxis for osteoporosis in patients on long-term glucocorticoid treatment.

  11. Analysis of Isocitrate Dehydrogenase -2 (IDH-2) Activity in Human Serum as a Biomarker in Chemotherapy Patients of Breast Carcinoma: A Case-Control Study.

    Science.gov (United States)

    Gavel, Roshni; Mishra, S P; Khanna, Seema; Khanna, Rahul; Shah, Agni Gautam

    2017-05-01

    Breast cancer represents a major public health problem in women worldwide. For many cancers, serum tumour markers play an important role in patient treatment and monitoring. Isocitrate dehydrogenase enzyme is also used as a biomarker for various types of cancer. The purpose of this study was to determine serum Isocitrate dehydrogenase-2 (IDH-2) enzyme activity in breast cancer patients (pre and post chemotherapy) and also correlate the changes in enzyme activity with stages of cancer and control groups. In this case-control study, histologically confirmed 40 female patients aged 28-80 years who fulfilled the criteria for diagnosis of invasive breast cancer were selected in our study groups from surgery outpatient department of SS Hospital, BHU, Varanasi, India, and 40 healthy age matched females were selected between October 2013 to July 2015. The estimation of serum IDH-2 enzyme activity in before and after two cycles of neoadjuvant chemotherapy patients was performed by spectrophotometry assay. The mean serum IDH-2 activity in cases (Mean±SD) was significantly more than control group (pIDH-2 activity in cases was significantly decrease after neo-adjuvant chemotherapy (p=0.019). In stage II pre chemotherapy patients serum IDH-2 activity was higher than post chemotherapy (pIDH-2 activity was not significant (p-value>0.05). The serum IDH-2 can be a potential biomarker in breast carcinoma and can be used for prognosis and monitoring the chemotherapy response of the patients.

  12. Structural and mechanistic aspects of alcohol dehydrogenase function

    OpenAIRE

    Svensson, Stefan

    1999-01-01

    Vertebrates possess a complex alcohol dehydrogenase (ADH) system composed of multiple molecular forms, which are currently classified into seven classes according to their structural properties. ADHs are dimeric zinc metalloenzymes that catalyze the reversible oxidation of alcohols to aldehydes/ketones using NAD+/NADH as electron acceptor and donor, respectively. The classes have broad but only partially overlapping substrate repertoires. This thesis mainly deals with mechan...

  13. Effect of repeated pesticide applications on soil properties in cotton fields: II. Insecticide residues and impact on dehydrogenase and arginine deaminase activities

    International Nuclear Information System (INIS)

    Vig, K.; Singh, D.K.; Agarwal, H.C.; Dhawan, A.K.; Dureja, P.

    2001-01-01

    Insecticides were applied sequentially at recommended dosages post crop emergence in cotton fields and soil was sampled at regular intervals after each treatment. Soil was analysed for insecticide residues and activity of the enzymes dehydrogenase and arginine deaminase. Insecticide residues detected in the soil were in small quantities and they did not persist for long. Only endosulfan leached below 15 cm. Insecticides had only temporary effects on enzyme activities which disappeared either before the next insecticide treatment or by the end of the experimental period. (author)

  14. Highly stable and reusable immobilized formate dehydrogenases: Promising biocatalysts for in situ regeneration of NADH

    Directory of Open Access Journals (Sweden)

    Barış Binay

    2016-02-01

    Full Text Available This study aimed to prepare robust immobilized formate dehydrogenase (FDH preparations which can be used as effective biocatalysts along with functional oxidoreductases, in which in situ regeneration of NADH is required. For this purpose, Candida methylica FDH was covalently immobilized onto Immobead 150 support (FDHI150, Immobead 150 support modified with ethylenediamine and then activated with glutaraldehyde (FDHIGLU, and Immobead 150 support functionalized with aldehyde groups (FDHIALD. The highest immobilization yield and activity yield were obtained as 90% and 132%, respectively when Immobead 150 functionalized with aldehyde groups was used as support. The half-life times (t1/2 of free FDH, FDHI150, FDHIGLU and FDHIALD were calculated as 10.6, 28.9, 22.4 and 38.5 h, respectively at 35 °C. FDHI150, FDHIGLU and FDHIALD retained 69, 38 and 51% of their initial activities, respectively after 10 reuses. The results show that the FDHI150, FDHIGLU and FDHIALD offer feasible potentials for in situ regeneration of NADH.

  15. Comparison of endogenous cytokinins and cytokinin oxidase/dehydrogenase activity in germinating and thermoinhibited Tagetes minuta achenes

    Czech Academy of Sciences Publication Activity Database

    Stirk, W. A.; Novák, Ondřej; Žižková, Eva; Motyka, Václav; Strnad, Miroslav; van Staden, J.

    2012-01-01

    Roč. 169, č. 7 (2012), s. 696-703 ISSN 0176-1617 R&D Projects: GA MŠk(CZ) LC06034; GA ČR(CZ) GAP506/11/0774 Grant - others:GA MŠk(CZ) ED0007/01/01 Program:ED Institutional research plan: CEZ:AV0Z50380511 Keywords : cytokinin biosynthesis * cytokinin oxidase/dehydrogenase * deactivation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.699, year: 2012

  16. Fluoxetine elevates allopregnanolone in female rat brain but inhibits a steroid microsomal dehydrogenase rather than activating an aldo-keto reductase.

    Science.gov (United States)

    Fry, J P; Li, K Y; Devall, A J; Cockcroft, S; Honour, J W; Lovick, T A

    2014-12-01

    Fluoxetine, a selective serotonin reuptake inhibitor, elevates brain concentrations of the neuroactive progesterone metabolite allopregnanolone, an effect suggested to underlie its use in the treatment of premenstrual dysphoria. One report showed fluoxetine to activate the aldo-keto reductase (AKR) component of 3α-hydroxysteroid dehydrogenase (3α-HSD), which catalyses production of allopregnanolone from 5α-dihydroprogesterone. However, this action was not observed by others. The present study sought to clarify the site of action for fluoxetine in elevating brain allopregnanolone. Adult male rats and female rats in dioestrus were treated with fluoxetine and their brains assayed for allopregnanolone and its precursors, progesterone and 5α-dihydroprogesterone. Subcellular fractions of rat brain were also used to investigate the actions of fluoxetine on 3α-HSD activity in both the reductive direction, producing allopregnanolone from 5α-dihydroprogesterone, and the reverse oxidative direction. Fluoxetine was also tested on these recombinant enzyme activities expressed in HEK cells. Short-term treatment with fluoxetine increased brain allopregnanolone concentrations in female, but not male, rats. Enzyme assays on native rat brain fractions and on activities expressed in HEK cells showed fluoxetine did not affect the AKR producing allopregnanolone from 5α-dihydroprogesterone but did inhibit the microsomal dehydrogenase oxidizing allopregnanolone to 5α-dihydroprogesterone. Fluoxetine elevated allopregnanolone in female rat brain by inhibiting its oxidation to 5α-dihydroprogesterone by a microsomal dehydrogenase. This is a novel site of action for fluoxetine, with implications for the development of new agents and/or dosing regimens to raise brain allopregnanolone. © 2014 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of The British Pharmacological Society.

  17. Chromate reduction by rabbit liver aldehyde oxidase

    Energy Technology Data Exchange (ETDEWEB)

    Banks, R.B.; Cooke, R.T. Jr.

    1986-05-29

    Chromate was reduced during the oxidation of 1-methylnicotinamide chlorine by partially purified rabbit liver aldehyde oxidase. In addition to l-methylnicotinamide, several other electron donor substrates for aldehyde oxidase were able to support the enzymatic chromate reduction. The reduction required the presence of both enzyme and the electron donor substrate. The rate of the chromate reduction was retarded by inhibitors or aldehyde oxidase but was not affected by substrates or inhibitors of xanthine oxidase. These results are consistent with the involvement of aldehyde oxidase in the reduction of chromate by rabbit liver cytosolic enzyme preparations.

  18. Redox Properties of Human Medium-Chain Acyl-CoA Dehydrogenase, Modulation by Charged Active-Site Amino Acid Residues

    OpenAIRE

    Mancini-Samuelson, Gina J.; Kieweg, Volker; Sabaj, Kim Marie; Ghisla, Sandro; Stankovich, Marian T.

    1998-01-01

    The modulation of the electron-transfer properties of human medium-chain acyl-CoA dehydrogenase (hwtMCADH) has been studied using wild-type and site-directed mutants by determining their midpoint potentials at various pH values and estimating the involved pKs. The mutants used were E376D, in which the negative charge is retained; E376Q, in which one negative charge (pKa ≈ 6.0) is removed from the active center; E99G, in which a different negative charge (pKa ≈ 7.3) also is affected; and E376H...

  19. Influence of the isolation procedure on coriander leaf volatiles with some correlation to the enzymatic activity.

    Science.gov (United States)

    To Quynh, Cung Thi; Iijima, Yoko; Kubota, Kikue

    2010-01-27

    Coriander leaves (Coriandrum sativum L.) have become popular worldwide because of their pleasant and delicate aroma. By a hot water extraction method, in which coriander leaves were cut before suspending in boiling water for 2 min, the contents of the main volatile compounds such as alkanals and 2-alkenals from C10 to C14 decreased, while the levels of corresponding alcohols increased in comparison to those obtained by solvent extraction. To investigate the reasons for this variation, an enzyme activity was assayed. By using aliphatic aldehyde as a substrate and NADPH as a coenzyme, strong activity of an aliphatic aldehyde reductase was found for the first time in this herb in the relatively wide pH range of 5.0-9.0, with the maximum activity at pH 8.5. Additionally, the aliphatic aldehyde dehydrogenase, responsible for acid formation, was also found to have a relatively weak activity compared to that of reductase.

  20. Changes in the Activities of Aldolase and of D-Glyceraldehyde-3-Phosphate Dehydrogenase during the Mitotic Cycle in Microspores of Lilium longiflorum

    Science.gov (United States)

    Nasatir, Maimon; Stern, Herbert

    1959-01-01

    Microspores of Lilium longiflorum were isolated at various stages of development surrounding the mitotic interval and were analyzed for changes in the activities of D-glyceraldehyde-3-phosphate dehydrogenase and aldolase. Fructose 1,6 diphosphate was used as substrate. Activities were measured by the increase in optical density due to the reduction of diphosphopyridine nucleotide. It was found that mitosis occurs during the minimal activity of both aldolase and D-glyceraldehyde-3-phosphate, thus indicating that heightened glycolytic capacity is not necessarily related to mitosis. It was also found that soluble-SH levels were highest when the enzymes were least active. It appeared, therefore, that the "—SH enzymes" are not necessarily activated intracellularly by high concentrations of soluble thiol. These results are discussed in connection with the theory that soluble-SH compounds stimulate glycolysis and in this way initiate mitosis. PMID:14426048

  1. Identification and functional evaluation of the reductases and dehydrogenases from Saccharomyces cerevisiae involved in vanillin resistance.

    Science.gov (United States)

    Wang, Xinning; Liang, Zhenzhen; Hou, Jin; Bao, Xiaoming; Shen, Yu

    2016-04-01

    Vanillin, a type of phenolic released during the pre-treatment of lignocellulosic materials, is toxic to microorganisms and therefore its presence inhibits the fermentation. The vanillin can be reduced to vanillyl alcohol, which is much less toxic, by the ethanol producer Saccharomyces cerevisiae. The reducing capacity of S. cerevisiae and its vanillin resistance are strongly correlated. However, the specific enzymes and their contribution to the vanillin reduction are not extensively studied. In our previous work, an evolved vanillin-resistant strain showed an increased vanillin reduction capacity compared with its parent strain. The transcriptome analysis suggested the reductases and dehydrogenases of this vanillin resistant strain were up-regulated. Using this as a starting point, 11 significantly regulated reductases and dehydrogenases were selected in the present work for further study. The roles of these reductases and dehydrogenases in the vanillin tolerance and detoxification abilities of S. cerevisiae are described. Among the candidate genes, the overexpression of the alcohol dehydrogenase gene ADH6, acetaldehyde dehydrogenase gene ALD6, glucose-6-phosphate 1-dehydrogenase gene ZWF1, NADH-dependent aldehyde reductase gene YNL134C, and aldo-keto reductase gene YJR096W increased 177, 25, 6, 15, and 18 % of the strain μmax in the medium containing 1 g L(-1) vanillin. The in vitro detected vanillin reductase activities of strain overexpressing ADH6, YNL134C and YJR096W were notably higher than control. The vanillin specific reduction rate increased by 8 times in ADH6 overexpressed strain but not in YNL134C and YJR096W overexpressed strain. This suggested that the enzymes encoded by YNL134C and YJR096W might prefer other substrate and/or could not show their effects on vanillin on the high background of Adh6p in vivo. Overexpressing ALD6 and ZWF1 mainly increased the [NADPH]/[NADP(+)] and [GSH]/[GSSG] ratios but not the vanillin reductase activities. Their

  2. Kinetic and chemical analyses of the cytokinin dehydrogenase-catalysed reaction : correlations with the crystal structure

    NARCIS (Netherlands)

    Popelková, Hana; Fraaije, Marco W.; Novák, Ondřej; Frébortová, Jitka; Bilyeu, Kristin D.; Frébort, Ivo

    2006-01-01

    CKX (cytokinin dehydrogenase) is a flavoprotein that cleaves cytokinins to adenine and the corresponding side-chain aldehyde using a quinone-type electron acceptor. In the present study, reactions of maize (Zea mays) CKX with five different substrates (N6-isopentenyladenine, trans-zeatin, kinetin,

  3. Emissions of odorous aldehydes from alkyd paint

    Science.gov (United States)

    Chang, John C. S.; Guo, Zhishi

    Aldehyde emissions are widely held responsible for the acrid after-odor of drying alkyd-based paint films. The aldehyde emissions from three different alkyd paints were measured in small environ-mental chambers. It was found that, for each gram of alkyd paint applied, more than 2 mg of aldehydes (mainly hexanal) were emitted during the curing (drying) period. Since no measurable hexanal was found in the original paint, it is suspected that the aldehydes emitted were produced by autoxidation of the unsaturated fatty acid esters in the alkyd resins. The hexanal emission rate was simulated by a model assuming that the autoxidation process was controlled by a consecutive first-order reaction mechanism. Using the emission rate model, indoor air quality simulation indicated that the hexanal emissions can result in prolonged (several days) exposure risk to occupants. The occupant exposure to aldehydes emitted from alkyd paint also could cause sensory irritation and other health concerns.

  4. Mechanism of catalytic action of oxide systems in reactions of aldehyde oxidation to carboxylic acids

    International Nuclear Information System (INIS)

    Andrushkevich, T.V.

    1997-01-01

    Mechanism of selective action of oxide catalysts (on the base of V 2 O 4 , MoO 3 ) of aldehyde oxidation to acids is considered, reaction acrolein oxidation to acrylic acid is taken as an example. Multistage mechanism of the process is established; it involves consequent transformation of coordination-bonded aldehyde into carbonyl-bonded aldehyde and symmetric carboxylate. Principles of active surface construction are formulated, they take into account the activity of stabilization center of concrete intermediate compound and bond energy of oxygen with surface. (author)

  5. INTERACTION OF ALDEHYDES DERIVED FROM LIPID PEROXIDATION AND MEMBRANE PROTEINS.

    Directory of Open Access Journals (Sweden)

    Stefania ePizzimenti

    2013-09-01

    Full Text Available A great variety of compounds are formed during lipid peroxidation of polyunsaturated fatty acids of membrane phospholipids. Among them, bioactive aldehydes, such as 4-hydroxyalkenals, malondialdehyde (MDA and acrolein, have received particular attention since they have been considered as toxic messengers that can propagate and amplify oxidative injury. In the 4-hydroxyalkenal class, 4-hydroxy-2-nonenal (HNE is the most intensively studied aldehyde, in relation not only to its toxic function, but also to its physiological role. Indeed, HNE can be found at low concentrations in human tissues and plasma and participates in the control of biological processes, such as signal transduction, cell proliferation and differentiation. Moreover, at low doses, HNE exerts an anti-cancer effect, by inhibiting cell proliferation, angiogenesis, cell adhesion and by inducing differentiation and/or apoptosis in various tumor cell lines. It is very likely that a substantial fraction of the effects observed in cellular responses, induced by HNE and related aldehydes, be mediated by their interaction with proteins, resulting in the formation of covalent adducts or in the modulation of their expression and/or activity. In this review we focus on membrane proteins affected by lipid peroxidation-derived aldehydes, under physiological and pathological conditions.

  6. The Activity of Escherichia coli Dihydroorotate Dehydrogenase is Dependent on a Conserved Loop Identified by Sequence Homology, Mutagenesis and Limited Proteolysis

    DEFF Research Database (Denmark)

    Björnberg, O.; Grüner, A.-C.; Roepstorff, P.

    1999-01-01

    of dihydroorotate dehydrogenases, but sedimentation in sucrose gradients suggests a dimeric structure also of the E. coli enzyme. Product inhibition showed that the E. coli enzyme, in contrast to the L. lactis enzyme, has separate binding sites for dihydroorotate and the electron acceptor. Trypsin readily cleaved...... the E. coli enzyme into two fragments of 182 and 154 residues, respectively. Cleavage reduced the activity more than 100-fold but left other molecular properties, including the heat stability, intact. The trypsin cleavage site, at R182, is positioned in a conserved region that, in the L. lactis enzyme......, forms a loop where a cysteine residue is very critical for activity. In the corresponding position, the enzyme from E. coli has a serine residue. Mutagenesis of this residue (S175) to alanine or cysteine reduced the activities 10000- and 500-fold, respectively. The S175C mutant was also defective...

  7. A role for glucose-6-phosphate dehydrogenase

    African Journals Online (AJOL)

    STORAGESEVER

    2009-01-19

    -phosphate dehydrogenase activity in male rats. Twelve (12) male rats were divided into two groups of six (6) rats each. Group 1 rats were control rats which received normal saline while group 2 rats were treated with.

  8. Regulation of glutamate dehydrogenase in Bacillus subtilis.

    OpenAIRE

    Kane, J F; Wakim, J; Fischer, R S

    1981-01-01

    The activity of the nicotinamide adenine dinucleotide-dependent glutamate dehydrogenase in Bacillus subtilis was influenced by the carbon source, but not the nitrogen source, in the growth medium. The highest specific activity for this enzyme was found when B. subtilis was grown in a minimal or rich medium that contained glutamate as the carbon source. It is proposed that glutamate dehydrogenase serves a catabolic function in the metabolism of glutamate, is induced by glutamate, and is subjec...

  9. Regulation of glutamate dehydrogenase in Bacillus subtilis.

    Science.gov (United States)

    Kane, J F; Wakim, J; Fischer, R S

    1981-01-01

    The activity of the nicotinamide adenine dinucleotide-dependent glutamate dehydrogenase in Bacillus subtilis was influenced by the carbon source, but not the nitrogen source, in the growth medium. The highest specific activity for this enzyme was found when B. subtilis was grown in a minimal or rich medium that contained glutamate as the carbon source. It is proposed that glutamate dehydrogenase serves a catabolic function in the metabolism of glutamate, is induced by glutamate, and is subject to catabolite repression. PMID:6118356

  10. Activation of Pyruvate Dehydrogenase by Sodium Dichloroacetate Shifts Metabolic Consumption from Amino Acids to Glucose in IPEC-J2 Cells and Intestinal Bacteria in Pigs.

    Science.gov (United States)

    An, Rui; Tang, Zhiru; Li, Yunxia; Li, Tiejun; Xu, Qingqing; Zhen, Jifu; Huang, Feiru; Yang, Jing; Chen, Cheng; Wu, Zhaoliang; Li, Mao; Sun, Jiajing; Zhang, Xiangxin; Chen, Jinchao; Wu, Liuting; Zhao, Shengjun; Qingyan, Jiang; Zhu, Weiyun; Yin, Yulong; Sun, Zhihong

    2018-04-18

    The extensive metabolism of amino acids (AA) as fuel is an important reason for the low use efficiency of protein in pigs. In this study, we investigated whether regulation of the pyruvate dehydrogenase kinase (PDK)/pyruvate dehydrogenase alpha 1 (PDHA1) pathway affected AA consumption by porcine intestinal epithelial (IPEC-J2) cells and intestinal bacteria in pigs. The effects of knockdown of PDHA1 and PDK1 with small interfering RNA (siRNA) on nutrient consumption by IPEC-J2 cells were evaluated. IPEC-J2 cells were then cultured with sodium dichloroacetate (DCA) to quantify AA and glucose consumption and nutrient oxidative metabolism. The results showed that knockdown of PDHA1 using siRNA decreased glucose consumption but increased total AA (TAA) and glutamate (Glu) consumption by IPEC-J2 cells ( P < 0.05). Opposite effects were observed using siRNA targeting PDK1 ( P < 0.05). Additionally, culturing IPEC-J2 cells in the presence of 5 mM DCA markedly increased the phosphorylation of PDHA1 and PDH phosphatase 1, but inhibited PDK1 phosphorylation ( P < 0.05). DCA treatment also reduced TAA and Glu consumption and increased glucose depletion ( P < 0.05). These results indicated that PDH was the regulatory target for shifting from AA metabolism to glucose metabolism and that culturing cells with DCA decreased the consumption of AAs by increasing the depletion of glucose through PDH activation.

  11. Kinetic characterization of recombinant Bacillus coagulans FDP-activated l-lactate dehydrogenase expressed in Escherichia coli and its substrate specificity.

    Science.gov (United States)

    Jiang, Ting; Xu, Yanbing; Sun, Xiucheng; Zheng, Zhaojuan; Ouyang, Jia

    2014-03-01

    Bacillus coagulans is a homofermentative, acid-tolerant and thermophilic sporogenic lactic acid bacterium, which is capable of producing high yields of optically pure lactic acid. The l-(+)-lactate dehydrogenase (l-LDH) from B. coagulans is considered as an ideal biocatalyst for industrial production. In this study, the gene ldhL encoding a thermostable l-LDH was amplified from B. coagulans NL01 genomic DNA and successfully expressed in Escherichia coli BL21 (DE3). The recombinant enzyme was partially purified and its enzymatic properties were characterized. Sequence analysis demonstrated that the l-LDH was a fructose 1,6-diphosphate-activated NAD-dependent lactate dehydrogenase (l-nLDH). Its molecular weight was approximately 34-36kDa. The Km and Vmax values of the purified l-nLDH for pyruvate were 1.91±0.28mM and 2613.57±6.43μmol(minmg)(-1), respectively. The biochemical properties of l-nLDH showed that the specific activity were up to 2323.29U/mg with optimum temperature of 55°C and pH of 6.5 in the pyruvate reduction and 351.01U/mg with temperature of 55°C and pH of 11.5 in the lactate oxidation. The enzyme also showed some activity in the absence of FDP, with a pH optimum of 4.0. Compared to other lactic acid bacterial l-nLDHs, the enzyme was found to be relatively stable at 50°C. Ca(2+), Ba(2+), Mg(2+) and Mn(2+) ions had activated effects on the enzyme activity, and the enzyme was greatly inhibited by Ni(2+) ion. Besides these, l-nLDH showed the higher specificity towards pyruvate esters, such as methyl pyruvate and ethyl pyruvate. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Enhancement of L-3-hydroxybutyryl-CoA dehydrogenase activity and circulating ketone body levels by pantethine. Relevance to dopaminergic injury

    Directory of Open Access Journals (Sweden)

    de Reggi Max

    2010-04-01

    Full Text Available Abstract Background The administration of the ketone bodies hydroxybutyrate and acetoacetate is known to exert a protective effect against metabolic disorders associated with cerebral pathologies. This suggests that the enhancement of their endogenous production might be a rational therapeutic approach. Ketone bodies are generated by fatty acid beta-oxidation, a process involving a mitochondrial oxido-reductase superfamily, with fatty acid-CoA thioesters as substrates. In this report, emphasis is on the penultimate step of the process, i.e. L-3-hydroxybutyryl-CoA dehydrogenase activity. We determined changes in enzyme activity and in circulating ketone body levels in the MPTP mouse model of Parkinson's disease. Since the active moiety of CoA is pantetheine, mice were treated with pantethine, its naturally-occurring form. Pantethine has the advantage of being known as an anti-inflammatory and hypolipidemic agent with very few side effects. Results We found that dehydrogenase activity and circulating ketone body levels were drastically reduced by the neurotoxin MPTP, whereas treatment with pantethine overcame these adverse effects. Pantethine prevented dopaminergic neuron loss and motility disorders. In vivo and in vitro experiments showed that the protection was associated with enhancement of glutathione (GSH production as well as restoration of respiratory chain complex I activity and mitochondrial ATP levels. Remarkably, pantethine treatment boosted the circulating ketone body levels in MPTP-intoxicated mice, but not in normal animals. Conclusions These finding demonstrate the feasibility of the enhancement of endogenous ketone body production and provide a promising therapeutic approach to Parkinson's disease as well as, conceivably, to other neurodegenerative disorders.

  13. Methanol-Water Aqueous-Phase Reforming with the Assistance of Dehydrogenases at Near-Room Temperature.

    Science.gov (United States)

    Shen, Yangbin; Zhan, Yulu; Li, Shuping; Ning, Fandi; Du, Ying; Huang, Yunjie; He, Ting; Zhou, Xiaochun

    2018-01-11

    As an excellent hydrogen-storage medium, methanol has many advantages, such as high hydrogen content (12.6 wt %), low cost, and availability from biomass or photocatalysis. However, conventional methanol-water reforming usually proceeds at high temperatures. In this research, we successfully designed a new effective strategy to generate hydrogen from methanol at near-room temperature. The strategy involved two main processes: CH 3 OH→HCOOH→H 2 and NADH→HCOOH→H 2 . The first process (CH 3 OH→HCOOH→H 2 ) was performed by an alcohol dehydrogenase (ADH), an aldehyde dehydrogenase (ALDH), and an Ir catalyst. The second procedure (NADH→HCOOH→H 2 ) was performed by formate dehydrogenase (FDH) and the Ir catalyst. The Ir catalyst used was a previously reported polymer complex catalyst [Cp*IrCl 2 (ppy); Cp*=pentamethylcyclopentadienyl, ppy=polypyrrole] with high catalytic activity for the decomposition of formic acid at room temperature and is compatible with enzymes, coenzymes, and poisoning chemicals. Our results revealed that the optimum hydrogen generation rate could reach up to 17.8 μmol h -1  g cat -1 under weak basic conditions at 30 °C. This will have high impact on hydrogen storage, production, and applications and should also provide new inspiration for hydrogen generation from methanol. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. The influence of individualizing physical loads on speed, creatine kinase activity and lactate dehydrogenase in football players.

    Directory of Open Access Journals (Sweden)

    2008-06-01

    Full Text Available Introduction: One of the most important training problems in: contemporary football is speed preparation of a player for the season and the ability of keeping it on the same, relatively high level throughout the starting period [1]. The main process used for re-synthesis ATP during single, short-lasting efforts of maximal intensity, is decomposition of phospho-creatine under the influence of creatine kinase enzyme. Physical loads imposed during speed trainings often exceed the possibility of producing energy from phosphogenic reserve through oxygen - lactate free processes, because the supply of phospho-creatine is used very quickly. In such circumstances the lacking energy is refilled through processes called oxygen free glicolise with the help of lactate dehydrogenase enzyme. The aim of the work was to answer the question:

  15. Aryl-aldehyde formation in fungal polyketides: discovery and characterization of a distinct biosynthetic mechanism.

    Science.gov (United States)

    Wang, Meng; Beissner, Mirko; Zhao, Huimin

    2014-02-20

    Aryl-aldehydes are a common feature in fungal polyketides, which are considered to be exclusively generated by the R domain of nonreducing polyketide synthases (NR-PKSs). However, by cloning and heterologous expression of both cryptic NR-PKS and nonribosomal peptide synthase (NRPS)-like genes from Aspergillus terreus in Saccharomyces cerevisiae, we identified a distinct mechanism for aryl-aldehyde formation in which a NRPS-like protein activates and reduces an aryl-acid produced by the accompanying NR-PKS to an aryl-aldehyde. Bioinformatics study indicates that such a mechanism may be widely used throughout the fungi kingdom. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Functional dissection of a trigger enzyme: mutations of the bacillus subtilis glutamate dehydrogenase RocG that affect differentially its catalytic activity and regulatory properties.

    Science.gov (United States)

    Gunka, Katrin; Newman, Joseph A; Commichau, Fabian M; Herzberg, Christina; Rodrigues, Cecilia; Hewitt, Lorraine; Lewis, Richard J; Stülke, Jörg

    2010-07-23

    Any signal transduction requires communication between a sensory component and an effector. Some enzymes engage in signal perception and transduction, as well as in catalysis, and these proteins are known as "trigger" enzymes. In this report, we detail the trigger properties of RocG, the glutamate dehydrogenase of Bacillus subtilis. RocG not only deaminates the key metabolite glutamate to form alpha-ketoglutarate but also interacts directly with GltC, a LysR-type transcription factor that regulates glutamate biosynthesis from alpha-ketoglutarate, thus linking the two metabolic pathways. We have isolated mutants of RocG that separate the two functions. Several mutations resulted in permanent inactivation of GltC as long as a source of glutamate was present. These RocG proteins have lost their ability to catabolize glutamate due to a strongly reduced affinity for glutamate. The second class of mutants is exemplified by the replacement of aspartate residue 122 by asparagine. This mutant protein has retained enzymatic activity but has lost the ability to control the activity of GltC. Crystal structures of glutamate dehydrogenases that permit a molecular explanation of the properties of the various mutants are presented. Specifically, we may propose that D122N replacement affects the surface of RocG. Our data provide evidence for a correlation between the enzymatic activity of RocG and its ability to inactivate GltC, and thus give insights into the mechanism that couples the enzymatic activity of a trigger enzyme to its regulatory function. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  17. S-Nitrosomycothiol Reductase and Mycothiol Are Required for Survival Under Aldehyde Stress and Biofilm Formation in Mycobacterium smegmatis

    Science.gov (United States)

    Vargas, Derek; Hageman, Samantha; Gulati, Megha; Nobile, Clarissa J.; Rawat, Mamta

    2017-01-01

    We show that Mycobacterium smegmatis mutants disrupted in mscR, coding for a dual function S-nitrosomycothiol reductase and formaldehyde dehydrogenase, and mshC, coding for a mycothiol ligase and lacking mycothiol (MSH), are more susceptible to S-nitrosoglutathione (GSNO) and aldehydes than wild type. MSH is a cofactor for MscR, and both mshC and mscR are induced by GSNO and aldehydes. We also show that a mutant disrupted in egtA, coding for a γ-glutamyl cysteine synthetase and lacking in ergothioneine, is sensitive to nitrosative stress but not to aldehydes. In addition, we find that MSH and S-nitrosomycothiol reductase are required for normal biofilm formation in M. smegmatis, suggesting potential new therapeutic pathways to target to inhibit or disrupt biofilm formation. PMID:27321674

  18. Nitrate reductase, arginine deaminase, urease and dehydrogenase activities in natural soil (ridges with forest) and in cotton soil after acetamiprid treatments.

    Science.gov (United States)

    Singh, Dileep K; Kumar, Sunil

    2008-03-01

    Soil enzymes are indicators of microbial activities in soil and are often considered as an indicator of soil health and fertility. They are very sensitive to the agricultural practices, pH of the soil, nutrients, inhibitors and weather conditions. To understand the effect of an insecticide, acetamiprid (IUPAC Name: (E)-N1-[(6-chloro-3-pyridyl) methyl]-N2-cyano-N1-methyl acetamidine) on different soil enzyme activities, the experiments were conducted for three consecutive years (2003--2005) at control and cotton experimental fields of Indian Agricultural Research Institute (IARI) and natural area (ridges with forest) in Delhi. The combined results for all three years were presented here to understand the impact of acetamiprid on soil enzyme activities. Acetamiprid was applied three times in one crop season after 41, 48 and 73 days of sowing, to control the pest. Soil of treated fields was analyzed for insecticide residues immediately after first insecticide treatment and thereafter at definite period. The residues of acetamiprid in experimental soil was varied from 0.30+/-0.13 to 22.67+/-0.2 microg g(-1)d.wt. soil, during the crop period of 2003. The insecticide residues for 2004 ranged between 0.59+/-0.38 and 13.42+/-0.71 microg g(-1)d.wt. soil and for 2005 it ranged between 0.48+/-0.22 and 19.81+/-0.33 microg g(-1)d.wt. soil. An average half life of acetamiprid in our treated field was 11.2+/-1.7 days for all three years. Similarly, the soil from natural area and control were also tested for insecticide residues. No detectable insecticide residues had been found. Soil from three localities i.e. natural, control and experimental fields were tested for different enzyme activities. Nitrate reductase, arginine deaminase, urease and dehydrogenase activities were high in natural soil in comparison to control soil and insecticide treated soil in all three experimental years. At the same time, nitrate reductase activity was all time low in acetamiprid treated soil

  19. Myoglobin-Catalyzed Olefination of Aldehydes.

    Science.gov (United States)

    Tyagi, Vikas; Fasan, Rudi

    2016-02-12

    The olefination of aldehydes constitutes a most valuable and widely adopted strategy for constructing carbon-carbon double bonds in organic chemistry. While various synthetic methods have been made available for this purpose, no biocatalysts are known to mediate this transformation. Reported herein is that engineered myoglobin variants can catalyze the olefination of aldehydes in the presence of α-diazoesters with high catalytic efficiency (up to 4,900 turnovers) and excellent E diastereoselectivity (92-99.9 % de). This transformation could be applied to the olefination of a variety of substituted benzaldehydes and heteroaromatic aldehydes, also in combination with different alkyl α-diazoacetate reagents. This work provides a first example of biocatalytic aldehyde olefination and extends the spectrum of synthetically valuable chemical transformations accessible using metalloprotein-based catalysts. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Substrate specificity of an aflatoxin-metabolizing aldehyde reductase.

    OpenAIRE

    Ellis, E M; Hayes, J D

    1995-01-01

    The enzyme from rat liver that reduces aflatoxin B1-dialdehyde exhibits a unique catalytic specificity distinct from that of other aldo-keto reductases. This enzyme, designated AFAR, displays high activity towards dicarbonyl-containing compounds with ketone groups on adjacent carbon atoms; 9,10-phenanthrenequinone, acenaphthenequinone and camphorquinone were found to be good substrates. Although AFAR can also reduce aromatic and aliphatic aldehydes such as succinic semialdehyde, it is inactiv...

  1. Aldehyde-sequestering drugs: tools for studying protein damage by lipid peroxidation products.

    Science.gov (United States)

    Burcham, Philip C; Kaminskas, Lisa M; Fontaine, Frank R; Petersen, Dennis R; Pyke, Simon M

    2002-12-27

    Elevated levels of reactive alpha,beta-unsaturated aldehydes (e.g. malondialdehyde, 4-hydroxynonenal and acrolein) in the affected tissues of various degenerative conditions suggest these substances are active propagators of the disease process. One experimental approach to attenuating damage by these intermediates employs 'aldehyde-sequestering drugs' as sacrificial nucleophiles, thereby sparing cell macromolecules and perhaps slowing disease progression. Drugs with demonstrated trapping activity toward lipid-derived aldehydes include various amine compounds such as aminoguanidine, carnosine and pyridoxamine. We have focused on identifying scavengers of acrolein, perhaps the most toxic aldehyde formed during lipid peroxidation cascades. Various phthalazine compounds (hydralazine and dihydralazine) were found to trap acrolein readily, forming hydrazone derivatives in a rapid Schiff-type reaction. These compounds strongly protect against acrolein-mediated toxicity in isolated hepatocytes.

  2. Chapter 18 (Part 2): Aldehydes & Ketones

    OpenAIRE

    Christiansen, Mike A

    2012-01-01

    In this video I'll teach you about what happens when we add acetylide, cyanide, and Grignard reagents to aldehydes and ketones. I also provide in-depth coverage on the reaction of aldehydes, ketones, carboxylic acids, esters, amides, and acyl (acid) chlorides with sodium borohydride (NaBH4), lithium aluminum hydride (LiAlH4), and DIBAL-H (or "diisobutyl aluminum hydride). --Dr. Mike Christiansen from Utah State University

  3. Efficient and Highly Aldehyde Selective Wacker Oxidation

    KAUST Repository

    Teo, Peili

    2012-07-06

    A method for efficient and aldehyde-selective Wacker oxidation of aryl-substituted olefins using PdCl 2(MeCN) 2, 1,4-benzoquinone, and t-BuOH in air is described. Up to a 96% yield of aldehyde can be obtained, and up to 99% selectivity can be achieved with styrene-related substrates. © 2012 American Chemical Society.

  4. Stilbene Glucoside, a Putative Sleep Promoting Constituent from Polygonum multiflorum Affects Sleep Homeostasis by Affecting the Activities of Lactate Dehydrogenase and Salivary Alpha Amylase.

    Science.gov (United States)

    Wei, Qian; Ta, Guang; He, Wenjing; Wang, Wei; Wu, Qiucheng

    2017-01-01

    Chinese herbal medicine (CHM) has been used for treating insomnia for centuries. The most used CHM for insomnia was Polygonum multiflorum. However, the molecular mechanism for CHM preventing insomnia is unknown. Stilbene glucoside (THSG), an important active component of P. multiflorum, may play an important role for treating insomnia. To test the hypothesis, Kunming mice were treated with different dosages of THSG. To examine the sleep duration, a computer-controlled sleep-wake detection system was implemented. Electroencephalogram (EEG) and electromyogram (EMG) electrodes were implanted to determine sleep-wake state. RT-PCR and Western blot was used to measure the levels of lactate dehydrogenase (LDH) and saliva alpha amylase. Spearman's rank correlation coefficient was used to identify the strength of correlation between the variables. The results showed that THSG significantly prolonged the sleep time of the mice (palpha amylase (palpha amylase (pamylase were negatively associated with sleep duration (palpha amylase.

  5. Structure of Cryptosporidium IMP dehydrogenase bound to an inhibitor with in vivo antiparasitic activity

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Youngchang; Makowska-Grzyska, Magdalena; Gorla, Suresh Kumar; Gollapalli, Deviprasad R.; Cuny, Gregory D.; Joachimiak, Andrzej; Hedstrom, Lizbeth

    2015-04-21

    Inosine 5'-monophosphate dehydrogenase (IMPDH) is a promising target for the treatment ofCryptosporidiuminfections. Here, the structure ofC. parvumIMPDH (CpIMPDH) in complex with inosine 5'-monophosphate (IMP) and P131, an inhibitor within vivoanticryptosporidial activity, is reported. P131 contains two aromatic groups, one of which interacts with the hypoxanthine ring of IMP, while the second interacts with the aromatic ring of a tyrosine in the adjacent subunit. In addition, the amine and NO2moieties bind in hydrated cavities, forming water-mediated hydrogen bonds to the protein. The design of compounds to replace these water molecules is a new strategy for the further optimization ofC. parvuminhibitors for both antiparasitic and antibacterial applications.

  6. In vitro effects of radioactive properties of 99mTc and 99mTc-MDP on human glucose 6-phosphate dehydrogenase activity

    Science.gov (United States)

    Sahin, Ali; Senturk, Murat

    2017-04-01

    The inhibitory effects of Na99mTcO4 (Sodium pertechnetate) and Na99mTcO4-metilendifosfonat (MDP) on human erythrocyte glucose 6-phosphate dehydrogenase (hG6PD) activity were investigated. For this purpose, hG6PD was initially purified 557-fold at a yield of 51.43% using 2',5'-adenosine diphosphate (ADP) sepharose 4B affinity gel chromatography. The in vitro effects of these compounds on hG6PD enzyme were studied. It was detected in in vitro studies that the hG6PD enzyme is inhibited due to Na99mTcO4 and Na99mTcO4-metilendifosfonat (MDP).

  7. Role of aldehydes in the toxic and mutagenic effects of nitrosamines

    OpenAIRE

    Peterson, Lisa A.; Urban, Anna M.; Vu, Choua C.; Cummings, Meredith E.; Brown, Lee C.; Warmka, Janel K.; Li, Li; Wattenberg, Elizabeth V.; Patel, Yesha; Stram, Daniel O.; Pegg, Anthony E.

    2013-01-01

    α-Hydroxynitrosamine metabolites of nitrosamines decompose to a reactive diazohydroxide and an aldehyde. To test the hypothesis that the aldehydes contribute to the harmful effects of nitrosamines, the toxic and mutagenic activity of three model methylating agents were compared in Chinese hamster ovary cells expressing human O6-alkylguanine DNA alkyltransferase (AGT) or not. N-Nitrosomethylurethane (NMUr), acetoxymethylmethylnitrosamine (AMMN) and 4-(methylnitrosamino)-4-acetoxy-1-(3-pyridyl)...

  8. Mutagenesis Study of the Cytochrome c Subunit Responsible for the Direct Electron Transfer-Type Catalytic Activity of FAD-Dependent Glucose Dehydrogenase

    Directory of Open Access Journals (Sweden)

    Yuki Yamashita

    2018-03-01

    Full Text Available The FAD-dependent glucose dehydrogenase from Burkholderia cepacia (FADGDH is a hetero-oligomeric enzyme that is capable of direct electron transfer (DET with an electrode. The cytochrome c (cyt c subunit, which possesses three hemes (heme 1, heme 2, and heme 3, from the N-terminal sequence, is known to enable DET; however, details of the electron transfer pathway remain unknown. A mutagenesis investigation of the heme axial ligands was carried out to elucidate the electron transfer pathway to the electron mediators and/or the electrode. The sixth axial ligand for each of the three heme irons, Met109, Met263, and Met386 were substituted with His. The catalytic activities of the wild-type (WT and mutant enzymes were compared by investigating their dye-mediated dehydrogenase activities and their DET abilities toward the electrode. The results suggested that (1 heme 1 with Met109 as an axial ligand is mainly responsible for the electron transfer with electron acceptors in the solution, but not for the DET with the electrode; (2 heme 2 with Met263 is responsible for the DET-type reaction with the electrode; and (3 heme 3 with Met386 seemed to be the electron acceptor from the catalytic subunit. From these results, two electron transfer pathways were proposed depending on the electron acceptors. Electrons are transferred from the catalytic subunit to heme 3, then to heme 2, to heme 1 and, finally, to electron acceptors in solution. However, if the enzyme complex is immobilized on the electrode and is used as electron acceptors, electrons are passed to the electrode from heme 2.

  9. Attenuated mitochondrial NADP+-dependent isocitrate dehydrogenase activity induces apoptosis and hypertrophy of H9c2 cardiomyocytes.

    Science.gov (United States)

    Lee, Jun Ho; Park, Jeen-Woo

    2014-04-01

    Oxidative stress, characterized by the accumulation of reactive oxygen species (ROS), is known to have numerous detrimental effects on the myocardium such as the induction of apoptotic cell death, hypertrophy, fibrosis, dysfunction, and dilatation. Over the past several years, we have shown that mitochondrial NADP(+)-dependent isocitrate dehydrogenase (IDPm) functions as an antioxidant and anti-apoptotic protein by supplying NADPH to antioxidant systems. Here, we showed that transfection of H9c2 clonal myoblastic cells with small interfering RNA (siRNA) specific for IDPm markedly attenuated IDPm expression and substantially induced apoptosis, senescence, and hypertrophy as indicated by increased atrial natriuretic peptide (ANP) gene expression, a marker of cardiomyocyte hypertrophy, and a larger cell size. Knockdown of IDPm expression resulted in the modulation of cellular and mitochondrial redox status, mitochondrial function, and cellular oxidative damage. Taken together, our results suggest that the suppression of IDPm expression by siRNA induces apoptosis and hypertrophy of cultured cardiomyocytes through the disruption of cellular redox balance. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  10. Plasmodium pyruvate dehydrogenase activity is only essential for the parasite's progression from liver infection to blood infection.

    Science.gov (United States)

    Pei, Ying; Tarun, Alice S; Vaughan, Ashley M; Herman, Rob W; Soliman, Joanne M B; Erickson-Wayman, Alyssa; Kappe, Stefan H I

    2010-02-01

    Plasmodium parasites possess a single pyruvate dehydrogenase (PDH) enzyme complex that is localized to the plastid-like organelle known as the apicoplast. Unlike most eukaryotes, Plasmodium parasites lack a mitochondrial PDH. The PDH complex catalyses the conversion of pyruvate to acetyl-CoA, an important precursor for the tricarboxylic acid cycle and type II fatty acid synthesis (FAS II). In this study, using a rodent malaria model, we show that the PDH E1 alpha and E3 subunits colocalize with the FAS II enzyme FabI in the apicoplast of liver stages but are not significantly expressed in blood stages. Deletion of the E1 alpha or E3 subunit genes of Plasmodium yoelii PDH caused no defect in blood stage development, mosquito stage development or early liver stage development. However, the gene deletions completely blocked the ability of the e1 alpha(-) and e3(-) parasites to form exo-erythrocytic merozoites during late liver stage development, thus preventing the initiation of a blood stage infection. This phenotype is similar to that observed for deletions of genes involved in FAS II elongation. The data strongly support the hypothesis that the sole role of PDH is to provide acetyl-CoA for FAS II.

  11. Role of Lipid Peroxidation-Derived α, β-Unsaturated Aldehydes in Vascular Dysfunction

    Directory of Open Access Journals (Sweden)

    Seung Eun Lee

    2013-01-01

    Full Text Available Vascular diseases are the most prominent cause of death, and inflammation and vascular dysfunction are key initiators of the pathophysiology of vascular disease. Lipid peroxidation products, such as acrolein and other α, β-unsaturated aldehydes, have been implicated as mediators of inflammation and vascular dysfunction. α, β-Unsaturated aldehydes are toxic because of their high reactivity with nucleophiles and their ability to form protein and DNA adducts without prior metabolic activation. This strong reactivity leads to electrophilic stress that disrupts normal cellular function. Furthermore, α, β-unsaturated aldehydes are reported to cause endothelial dysfunction by induction of oxidative stress, redox-sensitive mechanisms, and inflammatory changes such as induction of cyclooxygenase-2 and cytokines. This review provides an overview of the effects of lipid peroxidation products, α, β-unsaturated aldehydes, on inflammation and vascular dysfunction.

  12. Monoterpenic aldehydes as potential anti-Leishmania agents: activity of Cymbopogon citratus and citral on L. infantum, L. tropica and L. major.

    Science.gov (United States)

    Machado, M; Pires, P; Dinis, A M; Santos-Rosa, M; Alves, V; Salgueiro, L; Cavaleiro, C; Sousa, M C

    2012-03-01

    In order to contribute for the search of new drugs for leishmaniasis, we study the susceptibility of Leishmania infantum, Leishmania tropica and Leishmania major to Cymbopogon citratus essential oil and major compounds, mrycene and citral. C. citratus and citral were the most active inhibiting L. infantum, L. tropica and L. major growth at IC(50) concentrations ranging from 25 to 52 μg/ml and from 34 to 42 μg/ml, respectively. L. infantum promastigotes exposed to essential oil and citral underwent considerable ultrastructural alterations, namely mitochondrial and kinetoplast swelling, autophagosomal structures, disruption of nuclear membrane and nuclear chromatin condensation. C. citratus essential oil and citral promoted the leishmanicidal effect by triggering a programmed cell death. In fact, the leishmanicidal activity was mediated via apoptosis as evidenced by externalization of phosphatidylserine, loss of mitochondrial membrane potential, and cell-cycle arrest at the G(0)/G(1) phase. Taken together, ours findings lead us to propose that citral was responsible for anti-Leishmania activity of the C. citratus and both may represent a valuable source for therapeutic control of leishmaniasis. Copyright © 2011 Elsevier Inc. All rights reserved.

  13. Expression levels of chaperones influence biotransformation activity of recombinant Escherichia coli expressing Micrococcus luteus alcohol dehydrogenase and Pseudomonas putida Baeyer-Villiger monooxygenase.

    Science.gov (United States)

    Baek, A-Hyong; Jeon, Eun-Yeong; Lee, Sun-Mee; Park, Jin-Byung

    2015-05-01

    We demonstrated for the first time that the archaeal chaperones (i.e., γ-prefoldin and thermosome) can stabilize enzyme activity in vivo. Ricinoleic acid biotransformation activity of recombinant Escherichia coli expressing Micrococcus luteus alcohol dehydrogenase and the Pseudomonas putida KT2440 Baeyer-Villiger monooxygenase improved significantly with co-expression of γ-prefoldin or recombinant themosome originating from the deep-sea hyperthermophile archaea Methanocaldococcus jannaschii. Furthermore, the degree of enhanced activity was dependent on the expression levels of the chaperones. For example, whole-cell biotransformation activity was highest at 12 µmol/g dry cells/min when γ-prefoldin expression level was approximately 46% of the theoretical maximum. This value was approximately two-fold greater than that in E. coli, where the γ-prefoldin expression level was zero or set to the theoretical maximum. Therefore, it was assumed that the expression levels of chaperones must be optimized to achieve maximum biotransformation activity in whole-cell biocatalysts. © 2014 Wiley Periodicals, Inc.

  14. [Studies on antipeptic ulcer agents: the synthesis and structure-activity relationship analysis of heterocycle aldehyde (thio) semicarbazones and acyl hydrazones].

    Science.gov (United States)

    Guo, Z R; Yang, G Z; Chu, F M; Xu, G S; Zhang, J J; Zhang, S R; Yu, Y W

    1989-01-01

    Twenty-eight condensation products of heterocycle-alpha-carboaldehydes with N-aminooxazolidones, semicarbazides, thiosemicarbazides, aminoguanidines, aromatic hydrazides and benzoxycarbonyl hydrazide were synthesized so as to deduce the antiulcer pharmacophore or fragment of furazolidone(I), the prototype, which shows therapeutic efficacy for patients with gastric ulcer. Analysis of the SAR of the compounds indicate that the substitution of furan, thiophene, pyrrole and N-methyl pyrrole rings for the 5-nitrofuran and the cleavage of the oxazolidone ring obtain the activity to some extent. The necessary electronic density of carbonyl group of compounds is of importance. A lead structure, therefore, is derived for further optimization.

  15. Maximal-intensity exercise does not fully restore muscle pyruvate dehydrogenase complex activation after 3 days of high-fat dietary intake.

    Science.gov (United States)

    Constantin-Teodosiu, D; Cederblad, G; Bergström, M; Greenhaff, P L

    2018-02-15

    Exercise activates muscle pyruvate dehydrogenase complex (PDC), but moderate intensity exercise fails to fully activate muscle PDC after high-fat diet [1]. We investigated whether maximal intensity exercise overcomes this inhibition. Quadriceps femoris muscle biopsy samples were obtained from healthy males at rest, and after 46 and 92 electrically-evoked maximal intermittent isometric contractions, which were preceded by 3 days of either low- (18%) or high- (69%) isocaloric dietary fat intake (LFD and HFD, respectively). The ratio of PDCa (active form) to total PDCt (fully activated) at rest was 50% less after HFD (0.32 ± 0.01 vs 0.15 ± 0.01; P < 0.05). This ratio increased to 0.77 ± 0.06 after 46 contractions (P < 0.001) and to 0.98 ± 0.07 after 92 contractions (P < 0.001) in LFD. The corresponding values after HFD were less (0.54 ± 0.06; P < 0.01 and 0.70 ± 0.07; P < 0.01, respectively). Resting muscle acetyl-CoA and acetylcarnitine content was greater after HFD than LFD (both P < 0.05), but their rate of accumulation in the former was reduced during contraction. Muscle lactate content after 92 contractions was 30% greater after HFD (P < 0.05). Muscle force generation during contraction was no different between interventions, but HFD lengthened muscle relaxation time (P < 0.05). Daily urinary total carnitine excretion after HFD was 2.5-fold greater than after LFD (P < 0.01). A bout of maximal intense exercise did not overcome dietary fat-mediated inhibition of muscle pyruvate dehydrogenase complex activation, and was associated with greater muscle lactate accumulation, as a result of lower PDC flux, and increased muscle relaxation time. Copyright © 2018. Published by Elsevier Ltd.

  16. Effect of bioactive aldehydes on gelatin properties

    Directory of Open Access Journals (Sweden)

    I. P. Krysyuk

    2015-04-01

    Full Text Available Bioactive aldehydes are among main factors of proteins postsynthetic modifications, which are the cause and consequence of many diseases. Comparative study of some aldehydes modifying action on gelatin was carried out in vitro. Gelatin samples (20 mM were incubated with: ribose, deoxyribose, glyoxal, methylglyoxal, formaldehyde, acrolein (20 mM each and their combinations in 0.1 M Na-phosphate buffer (pH 7.4 containing 0.02% sodium azide at 37 °C in the dark for 30 days. We investigated the fluorescent properties of these samples and their molecular weight distribution by electrophoresis. It has been revealed that formed adducts had different fluorescence spectra. According to fluorescence intensity these aldehydes were put in order: formaldehyde < methylglyoxal < acrolein < ribose < deoxy­ribose < glyoxal. The electrophoresis results showed fragments of gelatin molecular weight redistribution. By this index, the aldehydes rating was as follows: ribose < deoxyribose < acrolein < glyoxal < formaldehyde < methylglyoxal. Comparison of these two ratings indicates that aldehydes with a lower ability to form fluorescent adducts have higher abili­ty to form intermolecular crosslinks. Therefore, the traditional clinical fluorescent test of a patients’ skin surface for collagen crosslinks determination has to be verified by other tests for proteins postsynthetic modifications.

  17. Structural characterization of a β-hydroxyacid dehydrogenase from Geobacter sulfurreducens and Geobacter metallireducens with succinic semialdehyde reductase activity

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Yanfeng; Zheng, Yi; Qin, Ling; Wang, Shihua; Buchko, Garry W.; Garavito, Michael R.

    2014-07-30

    Beta-hydroxyacid dehydrogenase (β-HAD) genes have been identified in all sequenced genomes of eukaryotes and prokaryotes. Their gene products catalyze the NAD+- or NADP+-dependent oxidation of various β-hydroxy acid substrates into their corresponding semialdehyde. In many fungal and bacterial genomes, multiple β-HAD genes are observed leading to the hypothesis that these gene products may have unique, uncharacterized metabolic roles specific to their species. The genomes of Geobacter sulfurreducens and Geobacter metallireducens each contain two potential β-HAD genes. The protein sequences of one pair of these genes, Gs-βHAD (Q74DE4) and Gm-βHAD (Q39R98), have 65% sequence identity and 77% sequence similarity with each other. Both proteins reduce succinic semialdehyde, a metabolite of the GABA shunt. To further explore the structural and functional characteristics of these two β-HADs with a potentially unique substrate specificity, crystal structures for Gs-βHAD and Gm-βHAD in complex with NADP+ were determined to a resolution of 1.89 Å and 2.07 Å, respectively. The structure of both proteins are similar, composed of 14 α-helices and nine β-strands organized into two domains. Domain One (1-165) adopts a typical Rossmann fold composed of two α/β units: a six-strand parallel β-sheet surrounded by six α-helices (α1 – α6) followed by a mixed three-strand β-sheet surrounded by two α-helices (α7 and α8). Domain Two (166-287) is composed of a bundle of seven α-helices (α9 – α14). Four functional regions conserved in all β-HADs are spatially located near each other at the interdomain cleft in both Gs-βHAD and Gm-βHAD with a buried molecule of NADP+. The structural features of Gs-βHAD and Gm-βHAD are described in relation to the four conserved consensus sequences characteristic of β-HADs and the potential biochemical importance of these enzymes as an alternative pathway for the degradation of succinic semialdehyde.

  18. Leucine-induced activation of translational initiation is partly regulated by the branched-chain α-keto acid dehydrogenase complex in C2C12 cells

    International Nuclear Information System (INIS)

    Nakai, Naoya; Shimomura, Yoshiharu; Tamura, Tomohiro; Tamura, Noriko; Hamada, Koichiro; Kawano, Fuminori; Ohira, Yoshinobu

    2006-01-01

    Branched-chain amino acid leucine has been shown to activate the translational regulators through the mammalian target of rapamycin. However, the leucine's effects are self-limiting because leucine promotes its own disposal by an oxidative pathway. The irreversible and rate-limiting step in the leucine oxidation pathway is catalyzed by the branched-chain α-keto acid dehydrogenase (BCKDH) complex. The complex contains E1 (α2β2), E2, and E3 subunits, and its activity is abolished by phosphorylation of the E1α subunit by BCKDH kinase. The relationship between the activity of BCKDH complex and leucine-mediated activation of the protein translation was investigated using the technique of RNA interference. The activity of BCKDH complex in C2C12 cell was modulated by transfection of small interfering RNA (siRNA) for BCKDH E2 subunit or BCKDH kinase. Transfection of siRNAs decreased the mRNA expression and protein amount of corresponding gene. Suppression of either E2 subunit or kinase produced opposite effects on the cell proliferation and the activation of translational regulators by leucine. Suppression of BCKDH kinase for 48 h resulted in decreasing cell proliferation. In contrast, E2 suppression led to increased amount of total cellular protein. The phosphorylation of p70 S6 kinase by leucine was increased in E2-siRNA transfected C2C12 cells, whereas the leucine's effect was diminished in kinase-siRNA transfected cells. These results suggest that the activation of the translational regulators by leucine was partly regulated by the activity of BCKDH complex

  19. Inhibitory action of marine algae extracts on the Trypanosoma cruzi dihydroorotate dehydrogenase activity and on the protozoan growth in mammalian cells.

    Science.gov (United States)

    Nara, Takeshi; Kamei, Yuto; Tsubouchi, Akiko; Annoura, Takeshi; Hirota, Kenichiro; Iizumi, Kyoichi; Dohmoto, Yuki; Ono, Takeaki; Aoki, Takashi

    2005-03-01

    Trypanosoma cruzi, the causative agent of Chagas' disease, replicates in mammalian cells and relies on the de novo pyrimidine biosynthetic pathway that supplies essential precursors for nucleic acid synthesis. The protozoan dihydroorotate dehydrogenase (DHOD), the fourth enzyme of the pathway catalyzing production of orotate from dihydroorotate, markedly differs from the human enzyme. This study was thus aimed to search for potent inhibitors against T. cruzi DHOD activity, and a number of methanol extracts prepared from green, brown, and red algae were assayed. The extracts from two brown algae, Fucus evanescens and Pelvetia babingtonii, yielded 59 and 58% decrease in the recombinant DHOD activity, respectively, at the concentration of 50 microg/ml. Inhibition by these extracts was noncompetitive with respect to dihydroorotate, with apparent Ki values of 35.3+/-5.9 and 10.3+/-4.4 microg/ml, respectively. Further, in an in vitro T. cruzi-HeLa cell infection system, ethanol-reconstituted F. evanescens and P. babingtonii extracts at the concentration of 1 microg/ml, respectively, decreased significantly the infection rate of host cells and the average parasite number per infected cell. These results imply that F. evanescens and P. babingtonii contain inhibitor(s) against the T. cruzi DHOD activity and against the protozoan infection and proliferation in mammalian cells. Identification of inhibitor(s) in these two brown algae and further screening of other marine algae may facilitate the discovery of new, anti-trypanosomal lead compounds.

  20. Effect of in vivo ozone exposure to Dorset sheep, an animal model with low levels of erythrocyte glucose-6-phosphate dehydrogenase activity

    Energy Technology Data Exchange (ETDEWEB)

    Moore, G.S.; Calabrese, E.J.; Schulz, E.

    1981-02-01

    Considerable interest has recently been directed to the possible extrapulmonary effects caused by exposure to ambient ozone. As a result of ozone induced in vivo alteration of red cell function within human subjects, it has been hypothesized that individuals with an erythrocyte glucose-6-phosphate dehydrogenase (G-6-PD) deficiency would be at increased hemolytic risk to elevated ambient ozone exposure. In order to evaluate such an hypothesis in an experimental setting it would be of great value to have an appropriate animal model with erythrocyte G-6-PD activity similar to the absolute activity range found in the human population. While no such unique animal model is presently known, the literature has revealed that Dorset sheep have an erythrocyte G-6-PD activity comparable in absolute units to a human G-6-PD deficient. Based on this information, we evaluated the mechanisms by which sheep and human G-6-PD deficient red cells handle oxidant stress. We evaluated the effects of in vivo ozone exposure in Dorset sheep over a broad range of concentrations.

  1. Effects of long-term oxygen treatment on α-ketoglutarate dehydrogenase activity and oxidative modifications in mitochondria of the guinea pig heart

    Directory of Open Access Journals (Sweden)

    Kaplan P

    2009-12-01

    Full Text Available Abstract Objective Oxygen therapy is used for the treatment of various diseases, but prolonged exposure to high concentrations of O2 is also associated with formation of free radicals and oxidative damage. Methods In the present study we compared α-ketoglutarate dehydrogenase (KGDH activity and mitochondrial oxidative damage in the hearts of guinea pigs after long-term (17 and 60 h oxygenation with 100% normobaric O2 and with partially negatively (O2 neg or positively (O2 posit ionized oxygen. Results Inhalation of O2 led to significant loss in KGDH activity and thiol group content and accumulation of bityrosines. Inhalation of O2 neg was accompanied by more pronounced KGDH inhibition, possibly due to additional formation of protein-lipid conjugates. In contrast, O2 posit prevented loss in KGDH activity and diminished mitochondrial oxidative damage. Conclusions These findings suggest that oxygen treatment is associated with impairment of heart energy metabolism and support the view that inhalation of O2 posit optimizes the beneficial effects of oxygen therapy.

  2. Effects of temperature, pH-values and sodium chloride concentrations on the glucose-6-phosphate dehydrogenase activity by thermotolerant Bacillus strains

    Directory of Open Access Journals (Sweden)

    HAZEM AQEL

    2012-01-01

    Full Text Available Thirteen new isolated thermotolerant Bacillus strains and four known Bacillus species were used to evaluate the effect of growth temperature, pH-values and NaCl concentrations on the intracellular glucose-6-phosphate dehydrogenase (G6PDH activity. Results had shown a significant difference in G6PDH production among all species at all used temperatures (p<0.05. The response of individual new isolates and controls for production of G6PDH under growth conditions was variable. The optimal growth conditions did not correspond to the optimal cultivation conditions for maximum G6PDH production. The growth temperature showed the most significant effect on G6PDH activity. The combined effect of temperature and NaCl on the G6PDH activity was strongly pronounced in comparison with the combined effect of temperature and pH or pH and NaCl. Thermal stability at 53ºC and electrophoretic mobility were also investigated. G6PDH from HUTB41 was the most thermostable G6PDH enzyme with T50% of more than 360 minutes. Electrophoretic study demonstrated that G6PDH was composed of two isoenzymes for all strains except B. marinus and B. schlegelii that had three isoenzymes.

  3. Association of glucose-6-phosphate dehydrogenase activity with oocyte cytoplasmic lipid content, developmental competence, and expression of candidate genes in a sheep model.

    Science.gov (United States)

    Mohammadi-Sangcheshmeh, Abdollah; Veshkini, Arash; Hajarizadeh, Athena; Jamshidi-Adegani, Fatemeh; Zhandi, Mahdi; Abazari-Kia, Amir Hossein; Cinar, Mehmet Ulas; Soleimani, Masoud; Gastal, Eduardo L

    2014-08-01

    To evaluate associations of glucose-6-phosphate dehydrogenase (G6PDH) activity in sheep oocytes with cytoplasmic lipid content, maturational competence, developmental competence to the blastocyst stage, and gene expression of certain molecular markers. Before brilliant cresyl blue (BCB) staining test, oocytes were classified as high, middle, and low cytoplasmic lipid content (HCLC, MCLC, and LCLC) and after the test as having low or high G6PDH-activity (BCB(+) and BCB(-), respectively). After maturation in vitro, a group of oocytes were subjected to IVF followed by in vitro embryo culture and another group was used for evaluation of expression of candidate genes. The cleavage and blastosyst rates were lowest (P BCB(+), and higher (P BCB(+) oocytes than the BCB(-) oocytes. Our gene expression data indicated that mRNA transcript abundance of ITGB2, pZP3, BMP15, and GDF9 genes was similar between BCB oocytes groups. However, the expression of ATP1A1 was higher (P BCB(+) oocytes compared to BCB(-) oocytes. In addition, BAX transcript abundance was similar (P > 0.05) among BCB(+), BCB(-), and control groups, before and after maturation in vitro. Activity of G6PDH in sheep oocytes is highly associated with lipid content, and compared with the morphological parameters might be a more precise and objective predictor for subsequent developmental competence in vitro.

  4. ALD5, PAD1, ATF1 and ATF2 facilitate the catabolism of coniferyl aldehyde, ferulic acid and p-coumaric acid in Saccharomyces cerevisiae

    Science.gov (United States)

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

    2017-01-01

    The ability of Saccharomyces cerevisiae to catabolize phenolic compounds remains to be fully elucidated. Conversion of coniferyl aldehyde, ferulic acid and p-coumaric acid by S. cerevisiae under aerobic conditions was previously reported. A conversion pathway was also proposed. In the present study, possible enzymes involved in the reported conversion were investigated. Aldehyde dehydrogenase Ald5, phenylacrylic acid decarboxylase Pad1, and alcohol acetyltransferases Atf1 and Atf2, were hypothesised to be involved. Corresponding genes for the four enzymes were overexpressed in a S. cerevisiae strain named APT_1. The ability of APT_1 to tolerate and convert the three phenolic compounds was tested. APT_1 was also compared to strains B_CALD heterologously expressing coniferyl aldehyde dehydrogenase from Pseudomonas, and an ald5Δ strain, all previously reported. APT_1 exhibited the fastest conversion of coniferyl aldehyde, ferulic acid and p-coumaric acid. Using the intermediates and conversion products of each compound, the catabolic route of coniferyl aldehyde, ferulic acid and p-coumaric acid in S. cerevisiae was studied in greater detail. PMID:28205618

  5. [EFFECT OF BIOACTIVE ALDEHYDES ON GELATIN PROPERTIES].

    Science.gov (United States)

    Krysyuk, I P; Dzvonkevych, N D; Volodina, T T; Popova, N N; Shandrenko, S G

    2015-01-01

    Bioactive aldehydes are among main factors of proteins postsynthetic modifications, which are the cause and consequence of many diseases. Comparative study of some aldehydes modifying action on gelatin was carried out in vitro. Gelatin samples (20 mM) were incubated with: ribose, deoxyribose, glyoxal, methylglyoxal, formaldehyde, acrolein (20 mM each) and their combinations in 0.1 M Naphosphate buffer (pH 7.4) containing 0.02% sodium azide at 37 °C in the dark for 30 days. We investigated the fluorescent properties of these samples and their molecular weight distribution by electrophoresis. It has been revealed that formed adducts had different fluorescence spectra. According to fluorescence intensity these aldehydes were put in order: formaldehyde acrolein acrolein test of a patients' skin surface for collagen crosslinks determination has to be verified by other tests for proteins postsynthetic modifications.

  6. Colorimetric Recognition of Aldehydes and Ketones.

    Science.gov (United States)

    Li, Zheng; Fang, Ming; LaGasse, Maria K; Askim, Jon R; Suslick, Kenneth S

    2017-08-07

    A colorimetric sensor array has been designed for the identification of and discrimination among aldehydes and ketones in vapor phase. Due to rapid chemical reactions between the solid-state sensor elements and gaseous analytes, distinct color difference patterns were produced and digitally imaged for chemometric analysis. The sensor array was developed from classical spot tests using aniline and phenylhydrazine dyes that enable molecular recognition of a wide variety of aliphatic or aromatic aldehydes and ketones, as demonstrated by hierarchical cluster, principal component, and support vector machine analyses. The aldehyde/ketone-specific sensors were further employed for differentiation among and identification of ten liquor samples (whiskies, brandy, vodka) and ethanol controls, showing its potential applications in the beverage industry. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. An Ultrasensitive Plasmonic Nanosensor for Aldehydes.

    Science.gov (United States)

    Li, Meng; Shi, Lei; Xie, Tao; Jing, Chao; Xiu, Guangli; Long, Yi-Tao

    2017-02-24

    Glucose is the most common but important aldehyde, and it is necessary to create biosensors with high sensitivity and anti-interference to detect it. Under the existence of silver ions and aldehyde compounds, single gold nanoparticles and freshly formed silver atoms could respectively act as core and shell, which finally form a core-shell structure. By observing the reaction between glucose and Tollens' reagent, metallic silver was found to be reduced on the surface of gold nanoparticles and formed Au@Ag nanoparticles that lead to a direct wavelength shift. Based on this principle and combined with in situ plasmon resonance scattering spectra, a plasmonic nanosensor was successfully applied in identifying aldehyde compounds with excellent sensitivity and specificity. This ultrasensitive sensor was successfully further utilized to detect blood glucose in mice serum samples, exhibiting good anti-interference ability and great promise for future clinical application.

  8. Redox properties of human medium-chain acyl-CoA dehydrogenase, modulation by charged active-site amino acid residues.

    Science.gov (United States)

    Mancini-Samuelson, G J; Kieweg, V; Sabaj, K M; Ghisla, S; Stankovich, M T

    1998-10-13

    The modulation of the electron-transfer properties of human medium-chain acyl-CoA dehydrogenase (hwtMCADH) has been studied using wild-type and site-directed mutants by determining their midpoint potentials at various pH values and estimating the involved pKs. The mutants used were E376D, in which the negative charge is retained; E376Q, in which one negative charge (pKa approximately 6. 0) is removed from the active center; E99G, in which a different negative charge (pKa approximately 7.3) also is affected; and E376H (pKa approximately 9.3) in which a positive charge is present. Em for hwtMCADH at pH 7.6 is -0.114 V. Results for the site-directed mutants indicate that loss of a negative charge in the active site causes a +0.033 V potential shift. This is consistent with the assumption that electrostatic interactions (as in the case of flavodoxins) and specific charges are important in the modulation of the electron-transfer properties of this class of dehydrogenases. Specifically, these charge interactions appear to correlate with the positive Em shift observed upon binding of substrate/product couple to MCADH [Lenn, N. D., Stankovich, M. T., and Liu, H. (1990) Biochemistry 29, 3709-3715], which coincides with a pK increase of Glu376-COOH from approximately 6 to 8-9 [Rudik, I., Ghisla, S., and Thorpe, C. (1998) Biochemistry 37, 8437-8445]. From the pH dependence of the midpoint potentials of hwtMCADH two mechanistically important ionizations are estimated. The pKa value of approximately 6.0 is assigned to the catalytic base, Glu376-COOH, in the oxidized enzyme based on comparison with the pH behavior of the E376H mutant, it thus coincides with the pK value recently estimated [Vock, P., Engst, S., Eder, M., and Ghisla, S. (1998) Biochemistry 37, 1848-1860]. The pKa of approximately 7.1 is assigned to Glu376-COOH in reduced hwtMCADH. Comparable values for these pKas for Glu376-COOH in pig kidney MCADH are pKox = 6.5 and pKred = 7.9. The Em measured for K304E-MCADH (a

  9. Contribution of ALDH1A1 isozyme to detoxification of aldehydes present in food products.

    Science.gov (United States)

    Sołobodowska, Sylwia; Giebułtowicz, Joanna; Wolinowska, Renata; Wroczyński, Piotr

    2012-01-01

    Even though food awareness is so developed and more and more people pay attention to what their diet is composed of, it is not possible to exclude all potentially dangerous substances present in our diet. One group of such compounds may be aldehydes as several studies indicate that they can be mutagenic, carcinogenic, genotoxic and cytotoxic. These relatively reactive organic molecules are natural constituents of food. They are also extensively used by food industry as additives giving aroma and taste. Fortunately many enzyme systems were developed to protect us against these toxic compounds, one of which is aldehyde dehydrogenase enzyme superfamily. As mouth is the first part of digestive system it seems crucial for detoxifying toxic substances introduced with our diet. The only ALDH isozyme present in saliva is ALDH3A1, which has very high affinity towards aromatic aldehydes commonly found in food. However, because of hyposalivation, which is not uncommon nowadays, the effectiveness of this barrier can be drastically diminished. As another member of this enzyme family, isozyme ALDH1A1 is also present in digestive system its possible contribution to detoxification of "food" aldehydes was addressed. Kinetic parameters (Km, Vmax) of recombinant ALDH1A1 towards several aliphatic and aromatic aldehydes occurring in food products (vanillin, citral, furfural, cinnamaldehyde, anisaldehyde, benzaldehyde and trans-hexenal) were determined by measuring the increase of NADH fluorescence after adding various concentrations of aldehyde substrates. Rates were used to construct the Lineweaver-Burk plot from which Km and Vmax (measured relative to that of benzaldehyde which was assigned the value of 100) values were calculated. The following results were obtained: 0.04 +/- 0.06 microM and 277 +/- 81 for anisaldehyde, 0.86 +/- 0.03 mciroM and 50 +/- 3 for vanillin, 0.18 +/- 0.05 mciroM and 93 +/- 9 for trans-2-hexenal, 0.17 +/- 0.03 microM and 201 +/- 32 for cinnamaldehyde, 5

  10. Effect of bioactive aldehydes on gelatin properties

    OpenAIRE

    I. P. Krysyuk; N. D. Dzvonkevych; T. T. Volodina; N. N. Popova; S. G. Shandrenko

    2015-01-01

    Bioactive aldehydes are among main factors of proteins postsynthetic modifications, which are the cause and consequence of many diseases. Comparative study of some aldehydes modifying action on gelatin was carried out in vitro. Gelatin samples (20 mM) were incubated with: ribose, deoxyribose, glyoxal, methylglyoxal, formaldehyde, acrolein (20 mM each) and their combinations in 0.1 M Na-phosphate buffer (pH 7.4) containing 0.02% sodium azide at 37 °C in the dark for 30 days. We investigated t...

  11. Disruption of seven hypothetical aryl alcohol dehydrogenase genes from Saccharomyces cerevisiae and construction of a multiple knock-out strain.

    Science.gov (United States)

    Delneri, D; Gardner, D C; Bruschi, C V; Oliver, S G

    1999-11-01

    By in silicio analysis, we have discovered that there are seven open reading frames (ORFs) in Saccharomyces cerevisiae whose protein products show a high degree of amino acid sequence similarity to the aryl alcohol dehydrogenase (AAD) of the lignin-degrading fungus Phanerochaete chrysosporium. Yeast cultures grown to stationary phase display a significant aryl alcohol dehydrogenase activity by degrading aromatic aldehydes to the corresponding alcohols. To study the biochemical and the biological role of each of the AAD genes, a series of mutant strains carrying deletion of one or more of the AAD-coding sequences was constructed by PCR-mediated gene replacement, using the readily selectable marker kanMX. The correct targeting of the PCR-generated disruption cassette into the genomic locus was verified by analytical PCR and by pulse-field gel electrophoresis (PFGE) followed by Southern blot analysis. Double, triple and quadruple mutant strains were obtained by classical genetic methods, while the construction of the quintuple, sextuple and septuple mutants was achieved by using the marker URA3 from Kluyveromyces lactis, HIS3 from Schizosaccharomyces pombe and TRP1 from S. cerevisiae. None of the knock-out strains revealed any mutant phenotype when tested for the degradation of aromatic aldehydes using both spectrophotometry and high performance liquid chromatography (HPLC). Specific tests for changes in the ergosterol and phospholipids profiles did not reveal any mutant phenotype and mating and sporulation efficiencies were not affected in the septuple deletant. Compared to the wild-type strain, the septuple deletant showed an increased resistance to the anisaldehyde, but there is a possibility that the nutritional markers used for gene replacement are causing this effect. Copyright 1999 John Wiley & Sons, Ltd.

  12. Evaluation of Milk Trace Elements, Lactate Dehydrogenase, Alkaline Phosphatase and Aspartate Aminotransferase Activity of Subclinical Mastitis as and Indicator of Subclinical Mastitis in Riverine Buffalo (Bubalus bubalis)

    Science.gov (United States)

    Guha, Anirban; Gera, Sandeep; Sharma, Anshu

    2012-01-01

    Mastitis is a highly morbid disease that requires detection at the subclinical stage. Tropical countries like India mainly depend on milch buffaloes for milk. The present study was conducted to investigate whether the trace minerals viz. copper (Cu), iron (Fe), zinc (Zn), cobalt (Co) and manganese (Mn) and enzyme activity of lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and aspartate aminotransferase (AST) in riverine buffalo milk can be used as an indicator of subclinical mastitis (SCM) with the aim of developing suitable diagnostic kit for SCM. Trace elements and enzyme activity in milk were estimated with Atomic absorption Spectrophotometer, GBC 932 plus and biochemical methods, respectively. Somatic cell count (SCC) was done microscopically. The cultural examination revealed Gram positive bacteria as the most prevalent etiological agent. A statistically significant (p<0.01) increase in SCC, Fe, Zn, Co and LDH occurred in SCM milk containing gram positive bacterial agents only. ALP was found to be elevated in milk infected by both gram positive and negative bacteria. The percent sensitivity, specificity and accuracy, predictive values and likelihood ratios were calculated taking bacterial culture examination and SCC≥2×105 cells/ml of milk as the benchmark. Only ALP and Zn, the former being superior, were found to be suitable for diagnosis of SCM irrespective of etiological agents. LDH, Co and Fe can be introduced in the screening programs where Gram positive bacteria are omnipresent. It is recommended that both ALP and Zn be measured together in milk to diagnose buffalo SCM, irrespective of etiology. PMID:25049573

  13. Neuronal Glud1 (glutamate dehydrogenase 1) over-expressing mice: increased glutamate formation and synaptic release, loss of synaptic activity, and adaptive changes in genomic expression.

    Science.gov (United States)

    Michaelis, E K; Wang, X; Pal, R; Bao, X; Hascup, K N; Wang, Y; Wang, W-T; Hui, D; Agbas, A; Choi, I-Y; Belousov, A; Gerhardt, G A

    2011-09-01

    Glutamate dehydrogenase 1 (GLUD1) is a mitochondrial enzyme expressed in all tissues, including brain. Although this enzyme is expressed in glutamatergic pathways, its function as a regulator of glutamate neurotransmitter levels is still not well defined. In order to gain an understanding of the role of GLUD1 in the control of glutamate levels and synaptic release in mammalian brain, we generated transgenic (Tg) mice that over-express this enzyme in neurons of the central nervous system. The Tg mice have increased activity of GLUD, as well as elevated levels and increased synaptic and depolarization-induced release of glutamate. These mice suffer age-associated losses of dendritic spines, nerve terminals, and neurons. The neuronal losses and dendrite structural changes occur in select regions of the brain. At the transcriptional level in the hippocampus, cells respond by increasing the expression of genes related to neurite growth and synapse formation, indications of adaptive or compensatory responses to the effects of increases in the release and action of glutamate at synapses. Because these Tg mice live to a relatively old age they are a good model of the effects of a "hyperglutamatergic" state on the aging process in the nervous system. The mice are also useful in defining the molecular pathways affected by the over-activation of GLUD in glutamatergic neurons of the brain and spinal cord. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. NADH:ubiquinone reductase and succinate dehydrogenase activity in the liver of rats with acetaminophen-induced toxic hepatitis on the background of alimentary protein deficiency

    Directory of Open Access Journals (Sweden)

    G. P. Kopylchuk

    2015-02-01

    Full Text Available The ratio between the redox forms of the nicotinamide coenzymes and key enzymatic activity of the I and II respiratory chain complexes in the liver cells mitochondria of rats with acetaminophen-induced hepatitis under the conditions of alimentary deprivation of protein was studied. It was estimated, that under the conditions of acute acetaminophen-induced hepatitis of rats kept on a low-protein diet during 4 weeks a significant decrease of the NADH:ubiquinone reductase and succinate dehydrogenase activity with simultaneous increase of the ratio between redox forms of the nicotinamide coenzymes (NAD+/NADН is observed compared to the same indices in the liver cells of animals with experimental hepatitis kept on the ration balanced by all nutrients. Results of research may become basic ones for the biochemical rationale for the approaches directed to the correction and elimination of the consequences­ of energy exchange in the toxic hepatitis, induced on the background of protein deficiency.

  15. L-arginine supplementation attenuates capillary regression without increasing integrated succinate dehydrogenase activity and VEGF expression in skeletal muscle during hindlimb unloading.

    Science.gov (United States)

    Uchida, Kensaku; Tanaka, Minoru; Kondo, Hiroyo; Ishihara, Akihiko; Fujino, Hidemi

    2016-10-01

    Decreased capillary number is observed in atrophied muscle. The change in capillary number is regulated by angiogenic factors. L-arginine enhances expression of endothelial nitric oxide synthase (eNOS), angiogenic factor, in skeletal muscle. Therefore, the aim of this study was to evaluate the effects of L-arginine supplementation on capillary regression during hindlimb unloading. Twenty-four male Wistar rats were divided into four treatment groups: (1) control, (2) L-arginine supplementation, (3) hindlimb unloading, and (4) hindlimb unloading with L-arginine supplementation. Hindlimb unloading resulted in a decrease of capillary-to-muscle fibre (C/F) ratio, eNOS expression, and integrated succinate dehydrogenase (SDH) activity. L-arginine supplementation attenuated the decrease in both eNOS expression and C/F ratio, although it did not increase integrated SDH activity in skeletal muscle. These results indicate that L-arginine supplementation is effective for maintaining capillary number in atrophied muscle, and that elevation of eNOS expression may be one mechanism associated with these responses.

  16. α-(Substituted-phenoxyacetoxy)-α-heterocyclylmethylphosphonates: synthesis, herbicidal activity, inhibition on pyruvate dehydrogenase complex (PDHc), and application as postemergent herbicide against broadleaf weeds.

    Science.gov (United States)

    He, Hong-Wu; Peng, Hao; Wang, Tao; Wang, Chubei; Yuan, Jun-Lin; Chen, Ting; He, Junbo; Tan, Xiaosong

    2013-03-13

    Pyruvate dehydrogenase complex (PDHc) is the site of action of a new class of herbicides. On the basis of the previous work for O,O'-dimethyl α-(substituted-phenoxyacetoxy)alkylphosphonates (I), further synthetic modifications were made by introducing a fural and a thienyl group to structure I. A series of α-(substituted-phenoxyacetoxy)-α-heterocyclylmethylphosphonate derivatives (II) were synthesized as potential inhibitors of PDHc. The postemergent activity of the title compounds II was evaluated in greenhouse experiments. The in vitro efficacy of II against PDHc was also examined. Compounds II with fural as R(3) and 2,4-dichloro as X and Y showed significant herbicidal activity and effective inhibition against PDHc from plants. O,O'-Dimethyl α-(2,4-dichlorophenoxyacetoxy)-α-(furan-2-yl)methylphosphonate II-17 had higher inhibitory potency against PDHc from Pisum sativum than against PDHc from Oryza sativa in vitro and was most effective against broadleaf weeds at 50 and 300 ai g/ha. II-17 was safe for maize and rice even at the dose of 900-1200 ai g/ha. Field trials at different regions in China showed that II-17 (HWS) could control a broad spectrum of broad-leaved and sedge weeds at the rate of 225-375 ai g/ha for postemergent applications in maize fields. II-17 (HWS) displayed potential utility as a selective herbicide.

  17. Isoniazid acetylating phenotype in patients with paracoccidioidomycosis and its relationship with serum sulfadoxin levels, glucose-6-phosphate dehydrogenase and glutathione reductase activities

    Directory of Open Access Journals (Sweden)

    Benedito Barraviera

    1991-06-01

    Full Text Available The authors evaluated the isoniazid acetylating phenotype and measured hematocrit, hemoglobin, glucose-6-phosphate dehydrogenase and glutathione reductase activities plus serum sulfadoxin levels in 39 patients with paracoccidioidomycosis (33 males and 6 females aged 17 to 58 years. Twenty one (53.84% of the patients presented a slow acetylatingphenotype and 18(46.16% a fast acetylating phenotype. Glucose-6-phosphate- dehydrogenase (G6PD acti vity was decreased in 5(23.80% slow acetylators and in 4(22.22% fast acetylators. Glutathione reductase activity was decreased in 14 (66.66% slow acetylators and in 12 (66.66% fast acetylators. Serum levels of free and total sulfadoxin Were higher in slow acetylator (p Os autores avaliaram o fenótipo acetilador da isoniazida, hematócrito, hemoglobina, atividade da glicose-6- fosfato desidrogenase, glutationa redutase e os níveis séricos de sulfadoxina de 39 doentes com paracoccidíoidomicose, senão 33 do sexo masculino e 6 do feminino, com idades compreendidas entre 17 e 58 anos. Vinte e um (53,84% doentes apresentaram fenótipo acetilador lento e 18 (46,16% rápido. A atividade da glicose-6-fosfato desidrogenase (G6PD esteve diminuída em 5 (23,80% acetiladores lentos e 4 (22,22% rápidos. A atividade da glutationa redutase esteve diminuída em 14 (66,66% acetiladores lentos e 12 (66,66% rápidos. Os níveis séricos de sulfadoxina livre e total foram maiores nos acetiladores lentos (p < 0,02. A análise dos resultados permite concluir que os níveis séricos de sulfadoxina relaciona-se com o fenótipo acetilador. Além disso, os níveis estiveram sempre acima de 50 µg/ml, níveis estes considerados terapêuticos. Por outro lado, a deficiência de glutationa redutase pode estar relacionada com a má absorção intestinal de nutrientes, entre eles riboflavina, vitamina precursora de FAD.

  18. Decreased 11β-Hydroxysteroid Dehydrogenase 1 Level and Activity in Murine Pancreatic Islets Caused by Insulin-Like Growth Factor I Overexpression.

    Directory of Open Access Journals (Sweden)

    Subrata Chowdhury

    Full Text Available We have reported a high expression of IGF-I in pancreatic islet β-cells of transgenic mice under the metallothionein promoter. cDNA microarray analysis of the islets revealed that the expression of 82 genes was significantly altered compared to wild-type mice. Of these, 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1, which is responsible for the conversion of inert cortisone (11-dehydrocorticosterone, DHC in rodents to active cortisol (corticosterone in the liver and adipose tissues, has not been identified previously as an IGF-I target in pancreatic islets. We characterized the changes in its protein level, enzyme activity and glucose-stimulated insulin secretion. In freshly isolated islets, the level of 11β-HSD1 protein was significantly lower in MT-IGF mice. Using dual-labeled immunofluorescence, 11β-HSD1 was observed exclusively in glucagon-producing, islet α-cells but at a lower level in transgenic vs. wild-type animals. MT-IGF islets also exhibited reduced enzymatic activities. Dexamethasone (DEX and DHC inhibited glucose-stimulated insulin secretion from freshly isolated islets of wild-type mice. In the islets of MT-IGF mice, 48-h pre-incubation of DEX caused a significant decrease in insulin release, while the effect of DHC was largely blunted consistent with diminished 11β-HSD1 activity. In order to establish the function of intracrine glucocorticoids, we overexpressed 11β-HSD1 cDNA in MIN6 insulinoma cells, which together with DHC caused apoptosis and a significant decrease in proliferation. Both effects were abolished with the treatment of an 11β-HSD1 inhibitor. Our results demonstrate an inhibitory effect of IGF-I on 11β-HSD1 expression and activity within the pancreatic islets, which may mediate part of the IGF-I effects on cell proliferation, survival and insulin secretion.

  19. A membrane-associated adenylate cyclase modulates lactate dehydrogenase and creatine kinase activities required for bull sperm capacitation induced by hyaluronic acid.

    Science.gov (United States)

    Fernández, Silvina; Córdoba, Mariana

    2017-04-01

    Hyaluronic acid, as well as heparin, is a glycosaminoglycan present in the female genital tract of cattle. The aim of this study was to evaluate oxidative metabolism and intracellular signals mediated by a membrane-associated adenylate cyclase (mAC), in sperm capacitation with hyaluronic acid and heparin, in cryopreserved bull sperm. The mAC inhibitor, 2',5'-dideoxyadenosine, was used in the present study. Lactate dehydrogenase (LDH) and creatine kinase (CK) activities and lactate concentration were determined spectrophotometrically in the incubation medium. Capacitation and acrosome reaction were evaluated by chlortetracycline technique, while plasma membrane and acrosome integrity were determined by trypan blue stain/differential interference contrast microscopy. Heparin capacitated samples had a significant decrease in LDH and CK activities, while in hyaluronic acid capacitated samples LDH and CK activities both increased compared to control samples, in heparin and hyaluronic acid capacitation conditions, respectively. A significant increase in lactate concentration in the incubation medium occurred in hyaluronic acid-treated sperm samples compared to heparin treatment, indicating this energetic metabolite is produced during capacitation. The LDH and CK enzyme activities and lactate concentrations in the incubation medium were decreased with 2',5'-dideoxyadenosine treatment in hyaluronic acid samples. The mAC inhibitor significantly inhibited heparin-induced capacitation of sperm cells, but did not completely inhibit hyaluronic acid capacitation. Therefore, hyaluronic acid and heparin are physiological glycosaminoglycans capable of inducing in vitro capacitation in cryopreserved bull sperm, stimulating different enzymatic pathways and intracellular signals modulated by a mAC. Hyaluronic acid induces sperm capacitation involving LDH and CK activities, thereby reducing oxidative metabolism, and this process is mediated by mAC. Copyright © 2017 Elsevier B.V. All

  20. Elevation of 11β-hydroxysteroid dehydrogenase type 2 activity in Holocaust survivor offspring: evidence for an intergenerational effect of maternal trauma exposure.

    Science.gov (United States)

    Bierer, Linda M; Bader, Heather N; Daskalakis, Nikolaos P; Lehrner, Amy L; Makotkine, Iouri; Seckl, Jonathan R; Yehuda, Rachel

    2014-10-01

    Adult offspring of Holocaust survivors comprise an informative cohort in which to study intergenerational transmission of the effects of trauma exposure. Lower cortisol and enhanced glucocorticoid sensitivity have been previously demonstrated in Holocaust survivors with PTSD, and in offspring of Holocaust survivors in association with maternal PTSD. In other work, reduction in the activity of the enzyme 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD-2), which inactivates cortisol, was identified in Holocaust survivors in comparison to age-matched, unexposed Jewish controls. Therefore, we investigated glucocorticoid metabolism in offspring of Holocaust survivors to evaluate if similar enzymatic decrements would be observed that might help to explain glucocorticoid alterations previously shown for Holocaust offspring. Holocaust offspring (n=85) and comparison subjects (n=27) were evaluated with clinical diagnostic interview and self-rating scales, and asked to collect a 24-h urine sample from which concentrations of cortisol and glucocorticoid metabolites were assayed by GCMS. 11β-HSD-2 activity was determined as the ratio of urinary cortisone to cortisol. Significantly reduced cortisol excretion was observed in Holocaust offspring compared to controls (p=.046), as had been shown for Holocaust survivors. However, 11β-HSD-2 activity was elevated for offspring compared to controls (p=.008), particularly among those whose mothers had been children, rather than adolescents or adults, during World War II (p=.032). The effect of paternal Holocaust exposure could not be reliably investigated in the current sample. The inverse association of offspring 11β-HSD-2 activity with maternal age at Holocaust exposure is consistent with the influence of glucocorticoid programming. Whereas a long standing reduction in 11β-HSD-2 activity among survivors is readily interpreted in the context of Holocaust related deprivation, understanding the directional effect on offspring will

  1. Major differences exist in the function and tissue-specific expression of human aflatoxin B1 aldehyde reductase and the principal human aldo-keto reductase AKR1 family members.

    Science.gov (United States)

    O'connor, T; Ireland, L S; Harrison, D J; Hayes, J D

    1999-01-01

    Complementary DNA clones encoding human aflatoxin B(1) aldehyde reductase (AKR7A2), aldehyde reductase (AKR1A1), aldose reductase (AKR1B1), dihydrodiol dehydrogenase 1 (AKR1C1) and chlordecone reductase (AKR1C4) have been expressed in Escherichia coli. These members of the aldo-keto reductase (AKR) superfamily have been purified from E. coli as recombinant proteins. The recently identified AKR7A2 was shown to differ from the AKR1 isoenzymes in being able to catalyse the reduction of 2-carboxybenzaldehyde. Also, AKR7A2 was found to exhibit a narrow substrate specificity, with activity being restricted to succinic semialdehyde (SSA), 2-nitrobenzaldehyde, pyridine-2-aldehyde, isatin, 1,2-naphthoquinone (1,2-NQ) and 9,10-phenanthrenequinone. In contrast, AKR1A1 reduces a broad spectrum of carbonyl-containing compounds, displaying highest specific activity for SSA, 4-carboxybenzaldehyde, 4-nitrobenzaldehyde, pyridine-3-aldehyde, pyridine-4-aldehyde, 4-hydroxynonenal, phenylglyoxal, methylglyoxal, 2,3-hexanedione, 1, 2-NQ, 16-ketoestrone and d-glucuronic acid. Comparison between the kinetic properties of AKR7A2 and AKR1A1 showed that both recombinant enzymes exhibited roughly similar k(cat)/K(m) values for SSA, 1,2-NQ and 16-ketoestrone. Many of the compounds which are substrates for AKR1A1 also serve as substrates for AKR1B1, though the latter enzyme was shown to display a specific activity significantly less than that of AKR1A1 for most of the aromatic and aliphatic aldehydes studied. Neither AKR1C1 nor AKR1C4 was found to possess high reductase activity towards aliphatic aldehydes, aromatic aldehydes, aldoses or dicarbonyls. However, unlike AKR1A1 and AKR1B1, both AKR1C1 and AKR1C4 were able to catalyse the oxidation of 1-acenaphthenol and, in addition, AKR1C4 could oxidize di- and tri-hydroxylated bile acids. Specific antibodies raised against AKR7A2, AKR1A1, AKR1B1, AKR1C1 and AKR1C4 have been used to show the presence of all of the reductases in human hepatic

  2. Gastric alcohol dehydrogenase activity in man: influence of gender, age, alcohol consumption and smoking in a caucasian population

    DEFF Research Database (Denmark)

    Parlesak, Alexandr; Billinger, M. H.; Bode, C.

    2002-01-01

    at ethanol concentrations between 150 and 500 mM. Mean ADH activity was higher in antral specimens than in those from the gastric corpus of the same subjects. ADH activity decreased with increasing age in males, while the values in females aged 41-60 years were higher than those in women aged 20-40 or 61...

  3. Antitrypanosomal compounds from the essential oil and extracts of Keetia leucantha leaves with inhibitor activity on Trypanosoma brucei glyceraldehyde-3-phosphate dehydrogenase.

    Science.gov (United States)

    Bero, J; Beaufay, C; Hannaert, V; Hérent, M-F; Michels, P A; Quetin-Leclercq, J

    2013-02-15

    Keetia leucantha is a West African tree used in traditional medicine to treat several diseases among which parasitic infections. The dichloromethane extract of leaves was previously shown to possess growth-inhibitory activities on Plasmodium falciparum, Trypanosoma brucei brucei and Leishmania mexicana mexicana with low or no cytotoxicity (>100 μg/ml on human normal fibroblasts) (Bero et al. 2009, 2011). In continuation of our investigations on the antitrypanosomal compounds from this dichloromethane extract, we analyzed by GC-FID and GC-MS the essential oil of its leaves obtained by hydrodistillation and the major triterpenic acids in this extract by LC-MS. Twenty-seven compounds were identified in the oil whose percentages were calculated using the normalization method. The essential oil, seven of its constituents and the three triterpenic acids were evaluated for their antitrypanosomal activity on Trypanosoma brucei brucei bloodstream forms (Tbb BSF) and procyclic forms (Tbb PF) to identify an activity on the glycolytic process of trypanosomes. The oil showed an IC(50) of 20.9 μg/ml on Tbb BSF and no activity was observed on Tbb PF. The best antitrypanosomal activity was observed for ursolic acid with IC(50) of 2.5 and 6.5 μg/ml respectively on Tbb BSF and Tbb PF. The inhibitory activity on a glycolytic enzyme of T. brucei, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), was also evaluated for betulinic acid, olenaolic acid, ursolic acid, phytol, α-ionone and β-ionone. The three triterpenic acids and β-ionone showed inhibitory activities on GAPDH with oleanolic acid being the most active with an inhibition of 72.63% at 20 μg/ml. This paper reports for the first time the composition and antitrypanosomal activity of the essential oil of Keetia leucantha. Several of its constituents and three triterpenic acids present in the dichloromethane leaves extract showed a higher antitrypanosomal activity on bloodstream forms of Tbb as compared to procyclic forms

  4. Palladium complexes of pyrrole-2-aldehyde thiosemicarbazone ...

    Indian Academy of Sciences (India)

    Vol. 126, No. 5, September 2014, pp. 1547–1555. c Indian Academy of Sciences. Palladium complexes of pyrrole-2-aldehyde thiosemicarbazone: Synthesis, structure and spectral properties. PIYALI PAUL and SAMARESH BHATTACHARYA. ∗. Department of Chemistry, Inorganic Chemistry Section, Jadavpur University, ...

  5. Aspirin inhibits glucose‑6‑phosphate dehydrogenase activity in HCT 116 cells through acetylation: Identification of aspirin-acetylated sites.

    Science.gov (United States)

    Ai, Guoqiang; Dachineni, Rakesh; Kumar, D Ramesh; Alfonso, Lloyd F; Marimuthu, Srinivasan; Bhat, G Jayarama

    2016-08-01

    Glucose-6-phosphate dehydrogenase (G6PD) catalyzes the first reaction in the pentose phosphate pathway, and generates ribose sugars, which are required for nucleic acid synthesis, and nicotinamide adenine dinucleotide phosphate (NADPH), which is important for neutralization of oxidative stress. The expression of G6PD is elevated in several types of tumor, including colon, breast and lung cancer, and has been implicated in cancer cell growth. Our previous study demonstrated that exposure of HCT 116 human colorectal cancer cells to aspirin caused acetylation of G6PD, and this was associated with a decrease in its enzyme activity. In the present study, this observation was expanded to HT‑29 colorectal cancer cells, in order to compare aspirin‑mediated acetylation of G6PD and its activity between HCT 116 and HT‑29 cells. In addition, the present study aimed to determine the acetylation targets of aspirin on recombinant G6PD to provide an insight into the mechanisms of inhibition. The results demonstrated that the extent of G6PD acetylation was significantly higher in HCT 116 cells compared with in HT‑29 cells; accordingly, a greater reduction in G6PD enzyme activity was observed in the HCT 116 cells. Mass spectrometry analysis of aspirin‑acetylated G6PD (isoform a) revealed that aspirin acetylated a total of 14 lysine residues, which were dispersed throughout the length of the G6PD protein. One of the important amino acid targets of aspirin included lysine 235 (K235, in isoform a) and this corresponds to K205 in isoform b, which has previously been identified as being important for catalysis. Acetylation of G6PD at several sites, including K235 (K205 in isoform b), may mediate inhibition of G6PD activity, which may contribute to the ability of aspirin to exert anticancer effects through decreased synthesis of ribose sugars and NADPH.

  6. Glutamates 78 and 122 in the active site of saccharopine dehydrogenase contribute to reactant binding and modulate the basicity of the acid-base catalysts.

    Science.gov (United States)

    Ekanayake, Devi K; Andi, Babak; Bobyk, Kostyantyn D; West, Ann H; Cook, Paul F

    2010-07-02

    Saccharopine dehydrogenase catalyzes the NAD-dependent oxidative deamination of saccharopine to give l-lysine and alpha-ketoglutarate. There are a number of conserved hydrophilic, ionizable residues in the active site, all of which must be important to the overall reaction. In an attempt to determine the contribution to binding and rate enhancement of each of the residues in the active site, mutations at each residue are being made, and double mutants are being made to estimate the interrelationship between residues. Here, we report the effects of mutations of active site glutamate residues, Glu(78) and Glu(122), on reactant binding and catalysis. Site-directed mutagenesis was used to generate E78Q, E122Q, E78Q/E122Q, E78A, E122A, and E78A/E122A mutant enzymes. Mutation of these residues increases the positive charge of the active site and is expected to affect the pK(a) values of the catalytic groups. Each mutant enzyme was completely characterized with respect to its kinetic and chemical mechanism. The kinetic mechanism remains the same as that of wild type enzymes for all of the mutant enzymes, with the exception of E78A, which exhibits binding of alpha-ketoglutarate to E and E.NADH. Large changes in V/K(Lys), but not V, suggest that Glu(78) and Glu(122) contribute binding energy for lysine. Shifts of more than a pH unit to higher and lower pH of the pK(a) values observed in the V/K(Lys) pH-rate profile of the mutant enzymes suggests that the presence of Glu(78) and Glu(122) modulates the basicity of the catalytic groups.

  7. Effects of L-carnitine and Pentoxifylline on the Activity of Lactate Dehydrogenase C4 isozyme and Motility of Testicular Spermatozoa in Mice.

    Science.gov (United States)

    Aliabadi, Elham; Karimi, Fatemeh; Rasti, Mozhgan; Akmali, Masoumeh; Esmaeilpour, Tahereh

    2013-04-01

    Extracted sperm from the testis have poor motility. Moreover, their motility changes during their journey through epidydimis. Meanwhile, they face high concentration of L-carnitin. In addition, lactate dehydrogenase C4 (LDH-C4) gene disorders has been shown to cause impaired sperm motility, leading to infertility in male mice. The aim of this study was to evaluate sperm motility and LDH-C4 enzyme activity upon L-carnitine (LC) and Pentoxifylline (PTX) administrations in mice. We extracted testicular sperm of 48 mice and divided them into three equal parts. One part was incubated with Ham's F10 medium (control), the other parts were treated with Ham's F10 containing LC and PTX with a final concentration of 1.76 mM, for 30 min at room temperature. Sperm motility was assessed according to the World Health Organization (WHO) criteria. Sperm LDH-C4 enzyme activity was measured by spectrophotometery method. Statistical analyses were performed using ANOVA and Fisher's LSD test, and a p-value less than 0.05 was considered as a statistically significant difference. Sperm motility increased after 30 min of incubation in LC- and PTX-treated group (p<0.001). LC and PTX administrations showed a significant increase in the LDHC4 enzyme activity of sperm compared to that of the controls after 30 min (P=0.04 and 0.01, respectively). The effects of LC and PTX on motility of sperm can be explained by an increase in LDH-C4 enzyme activity that may influence male fertility status. We suggest that LC as a non-toxic antioxidant is more suitable for use in assisted reproductive technique protocols than PTX.

  8. Prediction of oocyte developmental competence in ovine using glucose-6-phosphate dehydrogenase (G6PDH) activity determined at retrieval time.

    Science.gov (United States)

    Mohammadi-Sangcheshmeh, Abdollah; Soleimani, Masoud; Deldar, Hamid; Salehi, Mohammad; Soudi, Sara; Hashemi, Seyed Mahmoud; Schellander, Karl; Hoelker, Michael

    2012-02-01

    To determine whether G6PDH-activity measured by Brilliant Cresyl Blue known as BCB dye, predicts developmental competence within cohorts of ovine oocytes. Ovine oocytes were exposed to BCB staining and categorized into two groups: BCB+ (blue cytoplasm, low G6PDH-activity) and BCB- (colorless cytoplasm, high G6PDH-activity). After maturation in vitro, oocytes were subjected to fertilization followed by in vitro embryo culture. We observed a significant difference in oocyte diameter considering BCB+ and BCB- oocytes. BCB+ and Control groups showed significantly higher maturation rates compared to BCB- group. There were significantly more cleaved embryos in BCB+ and control groups than in BCB- group. Blastocyst rate was significantly higher for BCB+ group compared to control and BCB- groups with control group being significantly higher than BCB- group. G6PDH-activity is a strong predictive marker of oocyte competence and may be useful in identifying oocytes with a good prognosis for further develop.

  9. Crassiflorone derivatives that inhibit Trypanosoma brucei glyceraldehyde-3-phosphate dehydrogenase (TbGAPDH) and Trypanosoma cruzi trypanothione reductase (TcTR) and display trypanocidal activity.

    Science.gov (United States)

    Uliassi, Elisa; Fiorani, Giulia; Krauth-Siegel, R Luise; Bergamini, Christian; Fato, Romana; Bianchini, Giulia; Carlos Menéndez, J; Molina, Maria Teresa; López-Montero, Eulogio; Falchi, Federico; Cavalli, Andrea; Gul, Sheraz; Kuzikov, Maria; Ellinger, Bernhard; Witt, Gesa; Moraes, Carolina B; Freitas-Junior, Lucio H; Borsari, Chiara; Costi, Maria Paola; Bolognesi, Maria Laura

    2017-12-01

    Crassiflorone is a natural product with anti-mycobacterial and anti-gonorrhoeal properties, isolated from the stem bark of the African ebony tree Diospyros crassiflora. We noticed that its pentacyclic core possesses structural resemblance to the quinone-coumarin hybrid 3, which we reported to exhibit a dual-targeted inhibitory profile towards Trypanosoma brucei glyceraldehyde-3-phosphate dehydrogenase (TbGAPDH) and Trypanosoma cruzi trypanothione reductase (TcTR). Following this basic idea, we synthesized a small library of crassiflorone derivatives 15-23 and investigated their potential as anti-trypanosomatid agents. 19 is the only compound of the series showing a balanced dual profile at 10 μM (% inhibition TbGAPDH  = 64% and % inhibition TcTR  = 65%). In phenotypic assay, the most active compounds were 18 and 21, which at 5 μM inhibited Tb bloodstream-form growth by 29% and 38%, respectively. Notably, all the newly synthesized compounds at 10 μM did not affect viability and the status of mitochondria in human A549 and 786-O cell lines, respectively. However, further optimization that addresses metabolic liabilities including solubility, as well as cytochromes P450 (CYP1A2, CYP2C9, CYP2C19, and CYP2D6) inhibition, is required before this class of natural product-derived compounds can be further progressed. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  10. Site-directed mutagenesis under the direction of in silico protein docking modeling reveals the active site residues of 3-ketosteroid-Δ1-dehydrogenase from Mycobacterium neoaurum.

    Science.gov (United States)

    Qin, Ning; Shen, Yanbing; Yang, Xu; Su, Liqiu; Tang, Rui; Li, Wei; Wang, Min

    2017-07-01

    3-Ketosteroid-Δ 1 -dehydrogenases (KsdD) from Mycobacterium neoaurum could transform androst-4-ene-3,17-dione (AD) to androst-1,4-diene-3,17-dione. This reaction has a significant effect on the product of pharmaceutical steroid. The crystal structure and active site residues information of KsdD from Mycobacterium is not yet available, which result in the engineering of KsdD is tedious. In this study, by the way of protein modeling and site-directed mutagenesis, we find that, Y122, Y125, S138, E140 and Y541 from the FAD-binding domain and Y365 from the catalytic domain play a key role in this transformation. Compared with the wild type, the decline in AD conversion for mutants illustrated that Y125, Y365, and Y541 were essential to the function of KsdD. Y122, S138 and E140 contributed to the catalysis of KsdD. The following analysis revealed the catalysis mechanism of these mutations in KsdD of Mycobacterium. These information presented here facilitate the manipulation of the catalytic properties of the enzyme to improve its application in the pharmaceutical steroid industry.

  11. Identification and characterization of an antennae-specific aldehyde oxidase from the navel orangeworm.

    Directory of Open Access Journals (Sweden)

    Young-Moo Choo

    Full Text Available Antennae-specific odorant-degrading enzymes (ODEs are postulated to inactivate odorant molecules after they convey their signal. Different classes of insect ODEs are specific to esters, alcohols, and aldehydes--the major functional groups of female-produced, hydrophobic sex pheromones from moth species. Esterases that rapidly inactive acetate and other esters have been well-studied, but less is known about aldehyde oxidases (AOXs. Here we report cloning of an aldehyde oxidase, AtraAOX2, from the antennae of the navel orangeworm (NOW, Amyelois transitella, and the first activity characterization of a recombinant insect AOX. AtraAOX2 gene spans 3,813 bp and encodes a protein with 1,270 amino acid residues. AtraAOX2 cDNA was expressed in baculovirus-infected insect Sf21 cells as a ≈280 kDa homodimer with 140 kDa subunits. Recombinant AtraAOX2 degraded Z11Z13-16Ald and plant volatile aldehydes as substrates. However, as expected for aldehyde oxidases, recombinant AtraAOX2 did not show specificity for Z11Z13-16Ald, the main constituent of the sex pheromone, but showed high activity for plant volatile aldehydes. Our data suggest AtraAOX2 might be involved in degradation of a diversity of aldehydes including sex pheromones, plant-derived semiochemicals, and chemical cues for oviposition sites. Additionally, AtraAOX2 could protect the insect's olfactory system from xenobiotics, including pesticides that might reach the sensillar lymph surrounding the olfactory receptor neurons.

  12. Cloning, functional expression and characterization of a bifunctional 3-hydroxybutanal dehydrogenase /reductase involved in acetone metabolism by Desulfococcus biacutus.

    Science.gov (United States)

    Frey, Jasmin; Rusche, Hendrik; Schink, Bernhard; Schleheck, David

    2016-11-25

    The strictly anaerobic, sulfate-reducing bacterium Desulfococcus biacutus can utilize acetone as sole carbon and energy source for growth. Whereas in aerobic and nitrate-reducing bacteria acetone is activated by carboxylation with CO 2 to acetoacetate, D. biacutus involves CO as a cosubstrate for acetone activation through a different, so far unknown pathway. Proteomic studies indicated that, among others, a predicted medium-chain dehydrogenase/reductase (MDR) superfamily, zinc-dependent alcohol dehydrogenase (locus tag DebiaDRAFT_04514) is specifically and highly produced during growth with acetone. The MDR gene DebiaDRAFT_04514 was cloned and overexpressed in E. coli. The purified recombinant protein required zinc as cofactor, and accepted NADH/NAD + but not NADPH/NADP + as electron donor/acceptor. The pH optimum was at pH 8, and the temperature optimum at 45 °C. Highest specific activities were observed for reduction of C 3 - C 5 -aldehydes with NADH, such as propanal to propanol (380 ± 15 mU mg -1 protein), butanal to butanol (300 ± 24 mU mg -1 ), and 3-hydroxybutanal to 1,3-butanediol (248 ± 60 mU mg -1 ), however, the enzyme also oxidized 3-hydroxybutanal with NAD + to acetoacetaldehyde (83 ± 18 mU mg -1 ). The enzyme might play a key role in acetone degradation by D. biacutus, for example as a bifunctional 3-hydroxybutanal dehydrogenase/reductase. Its recombinant production may represent an important step in the elucidation of the complete degradation pathway.

  13. Initial steps of the peroxidase-catalyzed polymerization of coniferyl alcohol and/or sinapyl aldehyde: capillary zone electrophoresis study of pH effect.

    Science.gov (United States)

    Fournand, David; Cathala, Bernard; Lapierre, Catherine

    2003-01-01

    Capillary zone electrophoresis has been used to monitor the first steps of the dehydrogenative polymerization of coniferyl alcohol, sinapyl aldehyde, or a mixture of both, catalyzed by the horseradish peroxidase (HRP)-H(2)O(2) system. When coniferyl alcohol was the unique HRP substrate, three major dimers were observed (beta-5, beta-beta, and beta-O-4 interunit linkages) and their initial formation velocity as well as their relative abundance varied with pH. The beta-O-4 interunit linkage was thus slightly favored at lower pH values. In contrast, sinapyl aldehyde turned out to be a very poor substrate for HRP except in basic conditions (pH 8). The major dimer observed was the beta,beta'-di-sinapyl aldehyde, a red-brown exhibiting compound which might partly participate in the red coloration usually observed in cinnamyl alcohol dehydrogenase-deficient angiosperms. Finally, when a mixture of coniferyl alcohol and sinapyl aldehyde was used, it looked as if sinapyl aldehyde became a very good substrate for HRP. Indeed, coniferyl alcohol turned out to serve as a redox mediator (i.e. "shuttle oxidant") for the sinapyl aldehyde incorporation in the lignin-like polymer. This means that in particular conditions the specificity of oxidative enzymes might not hinder the incorporation of poor substrates into the growing lignin polymer.

  14. 40 CFR 721.5762 - Aromatic aldehyde phenolic resin (generic).

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Aromatic aldehyde phenolic resin... Specific Chemical Substances § 721.5762 Aromatic aldehyde phenolic resin (generic). (a) Chemical substance... aromatic aldehyde phenolic resin (PMN P-01-573) is subject to reporting under this section for the...

  15. Studies on lipoamide dehydrogenase

    NARCIS (Netherlands)

    Benen, J.A.E.

    1992-01-01

    At the onset of the investigations described in this thesis progress was being made on the elucidation of the crystal structure of the Azotobactervinelandii lipoamide dehydrogenase. Also the gene encoding this enzyme was cloned in our laboratory. By this, a

  16. Studies on lipoamide dehydrogenase

    NARCIS (Netherlands)

    Visser, J.

    1969-01-01

    Gel-filtration, ultracentrifugation and sucrose density gradient centrifugation demonstrated differences in physico-chemical properties of holoenzyme and apoenzyme of lipoamide dehydrogenase. The native apoenzyme has a mol.wt. of approx. 52,000 which is half that of the native holoenzyme. The

  17. Characterization of a novel PQQ-dependent quinohemoprotein pyranose dehydrogenase from Coprinopsis cinerea classified into auxiliary activities family 12 in carbohydrate-active enzymes.

    Directory of Open Access Journals (Sweden)

    Kouta Takeda

    Full Text Available The basidiomycete Coprinopsis cinerea contains a quinohemoprotein (CcPDH named as CcSDH in our previous paper, which is a new type of pyrroloquinoline-quinone (PQQ-dependent pyranose dehydrogenase and is the first found among all eukaryotes. This enzyme has a three-domain structure consisting of an N-terminal heme b containing a cytochrome domain that is homologous to the cytochrome domain of cellobiose dehydrogenase (CDH; EC 1.1.99.18 from the wood-rotting basidiomycete Phanerochaete chrysosporium, a C-terminal family 1-type carbohydrate-binding module, and a novel central catalytic domain containing PQQ as a cofactor. Here, we describe the biochemical and electrochemical characterization of recombinant CcPDH. UV-vis and resonance Raman spectroscopic studies clearly reveal characteristics of a 6-coordinated low-spin heme b in both the ferric and ferrous states, as well as intramolecular electron transfer from the PQQ to heme b. Moreover, the formal potential of the heme was evaluated to be 130 mV vs. NHE by cyclic voltammetry. These results indicate that the cytochrome domain of CcPDH possesses similar biophysical properties to that in CDH. A comparison of the conformations of monosaccharides as substrates and the associated catalytic efficiency (kcat/Km of CcPDH indicates that the enzyme prefers monosaccharides with equatorial C-2, C-3 hydroxyl groups and an axial C-4 hydroxyl group in the 1C4 chair conformation. Furthermore, a binding study shows a high binding affinity of CcPDH for cellulose, suggesting that CcPDH function is related to the enzymatic degradation of plant cell wall.

  18. Hypoxanthine-guanine phosphoribosyltransferase and inosine 5’-monophosphate dehydrogenase activities in three mammalian species: aquatic (Mirounga angustirostris, semiaquatic (Lontra longicaudis annectens and terrestrial (Sus scrofa

    Directory of Open Access Journals (Sweden)

    Myrna eBarjau Perez-Milicua

    2015-07-01

    Full Text Available Aquatic and semiaquatic mammals have the capacity of breath hold (apnea diving. Northern elephant seals (Mirounga angustirostris have the ability to perform deep and long duration dives; during a routine dive, adults can hold their breath for 25 min. Neotropical river otters (Lontra longicaudis annectens can hold their breath for about 30 sec. Such periods of apnea may result in reduced oxygen concentration (hypoxia and reduced blood supply (ischemia to tissues. Production of adenosine 5’-triphosphate (ATP requires oxygen, and most mammalian species, like the domestic pig (Sus scrofa, are not adapted to tolerate hypoxia and ischemia, conditions that result in ATP degradation. The objective of this study was to explore the differences in purine synthesis and recycling in erythrocytes and plasma of three mammalian species adapted to different environments: aquatic (northern elephant seal (n=11, semiaquatic (neotropical river otter (n=4 and terrestrial (domestic pig (n=11. Enzymatic activity of hypoxanthine-guanine phosphoribosyltransferase (HGPRT was determined by spectrophotometry, and activity of inosine 5’-monophosphate dehydrogenase (IMPDH and the concentration of hypoxanthine (HX, inosine 5’-monophosphate (IMP, adenosine 5’-monophosphate (AMP, adenosine 5’-diphosphate (ADP, ATP, guanosine 5’-diphosphate (GDP, guanosine 5’-triphosphate (GTP, and xanthosine 5’-monophosphate (XMP were determined by high-performance liquid chromatography (HPLC. The activities of HGPRT and IMPDH and the concentration of HX, IMP, AMP, ADP, ATP, GTP and XMP in erythrocytes of domestic pigs were higher than in erythrocytes of northern elephant seals and river otters. These results suggest that under basal conditions (no diving, sleep apnea or exercise, aquatic and semiaquatic mammals have less purine mobilization than their terrestrial counterparts.

  19. A trade off between catalytic activity and protein stability determines the clinical manifestations of glucose-6-phosphate dehydrogenase (G6PD) deficiency.

    Science.gov (United States)

    Boonyuen, Usa; Chamchoy, Kamonwan; Swangsri, Thitiluck; Junkree, Thanyaphorn; Day, Nicholas P J; White, Nicholas J; Imwong, Mallika

    2017-11-01

    Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common polymorphism and enzymopathy in humans, affecting approximately 400 million people worldwide. It is responsible for various clinical manifestations, including favism, hemolytic anemia, chronic non-spherocytic hemolytic anemia, spontaneous abortion, and neonatal hyperbilirubinemia. Understanding the molecular mechanisms underlying the severity of G6PD deficiency is of great importance but that of many G6PD variants are still unknown. In this study, we report the construction, expression, purification, and biochemical characterization in terms of kinetic properties and stability of five clinical G6PD variants-G6PD Bangkok, G6PD Bangkok noi, G6PD Songklanagarind, G6PD Canton+Bangkok noi, and G6PD Union+Viangchan. G6PD Bangkok and G6PD Canton+Bangkok noi showed a complete loss of catalytic activity and moderate reduction in thermal stability when compared with the native G6PD. G6PD Bangkok noi and G6PD Union+Viangchan showed a significant reduction in catalytic efficiency, whereas G6PD Songklanagarind showed a catalytic activity comparable to the wild-type enzyme. The Union+Viangchan mutation showed a remarkable effect on the global stability of the enzyme. In addition, our results indicate that the location of mutations in G6PD variants affects their catalytic activity, stability, and structure. Hence, our results provide a molecular explanation for clinical manifestations observed in individuals with G6PD deficiency. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  20. Cloning and expression analysis of alcohol dehydrogenase ( Adh ...

    African Journals Online (AJOL)

    Hybrid promoters are created by shuffling of DNA fragments while keeping intact regulatory regions crucial of promoter activity. Two fragments of alcohol dehydrogenase (Adh) promoter from Zea mays were selected to generate hybrid promoter. Sequence analysis of both alcohol dehydrogenase promoter fragments through ...

  1. Some Properties of Glutamate Dehydrogenase from the Marine Red ...

    African Journals Online (AJOL)

    Keywords: ammonia assimilation, glutamate dehydrogenase, GDH, Gracilaria sordida, red alga, enzyme activity. Glutamate dehydrogenases (GDH, EC ... Anabolic functions could be assimilation of ammonia released during photorespiration and synthesis of N-rich transport compounds. Western Indian Ocean Journal of ...

  2. Succinic semialdehyde dehydrogenase deficiency, a disorder of GABA metabolism: an update on pharmacological and enzyme-replacement therapeutic strategies.

    Science.gov (United States)

    Vogel, Kara R; Ainslie, Garrett R; Walters, Dana C; McConnell, Alice; Dhamne, Sameer C; Rotenberg, Alexander; Roullet, Jean-Baptiste; Gibson, K Michael

    2018-02-19

    We present an update to the status of research on succinic semialdehyde dehydrogenase (SSADH) deficiency (SSADHD), a rare disorder of GABA metabolism. This is an unusual disorder featuring the accumulation of both GABA and its neuromodulatory analog, gamma-hydroxybutyric acid (GHB), and recent studies have advanced the potential clinical application of NCS-382, a putative GHB receptor antagonist. Animal studies have provided proof-of-concept that enzyme replacement therapy could represent a long-term therapeutic option. The characterization of neuronal stem cells (NSCs) derived from aldehyde dehydrogenase 5a1 -/- (aldh5a1 -/- ) mice, the murine model of SSADHD, has highlighted NSC utility as an in vitro system in which to study therapeutics and associated toxicological properties. Gene expression analyses have revealed that transcripts encoding GABA A receptors are down-regulated and may remain largely immature in aldh5a1 -/- brain, characterized by excitatory as opposed to inhibitory outputs, the latter being the expected action in the mature central nervous system. This indicates that agents altering chloride channel activity may be therapeutically relevant in SSADHD. The most recent therapeutic prospects include mTOR (mechanistic target of rapamycin) inhibitors, drugs that have received attention with the elucidation of the effects of elevated GABA on autophagy. The outlook for novel therapeutic trials in SSADHD continues to improve.

  3. Co-production of hydrogen and ethanol from glucose inEscherichia coliby activation of pentose-phosphate pathway through deletion of phosphoglucose isomerase (pgi) and overexpression of glucose-6-phosphate dehydrogenase (zwf) and 6-phosphogluconate dehydrogenase (gnd).

    Science.gov (United States)

    Sundara Sekar, Balaji; Seol, Eunhee; Park, Sunghoon

    2017-01-01

    Biologically, hydrogen (H 2 ) can be produced through dark fermentation and photofermentation. Dark fermentation is fast in rate and simple in reactor design, but H 2 production yield is unsatisfactorily low as glucose. To address this challenge, simultaneous production of H 2 and ethanol has been suggested. Co-production of ethanol and H 2 requires enhanced formation of NAD(P)H during catabolism of glucose, which can be accomplished by diversion of glycolytic flux from the Embden-Meyerhof-Parnas (EMP) pathway to the pentose-phosphate (PP) pathway in Escherichia coli . However, the disruption of pgi ( p hospho g lucose i somerase) for complete diversion of carbon flux to the PP pathway made E. coli unable to grow on glucose under anaerobic condition. Here, we demonstrate that, when glucose-6-phosphate dehydrogenase (Zwf) and 6-phosphogluconate dehydrogenase (Gnd), two major enzymes of the PP pathway, are homologously overexpressed, E. coli Δ pgi can recover its anaerobic growth capability on glucose. Further, with additional deletions of Δ hycA , Δ hyaAB , Δ hybBC , Δ ldhA , and Δ frdAB , the recombinant Δ pgi mutant could produce 1.69 mol H 2 and 1.50 mol ethanol from 1 mol glucose. However, acetate was produced at 0.18 mol mol -1 glucose, indicating that some carbon is metabolized through the Entner-Doudoroff (ED) pathway. To further improve the flux via the PP pathway, heterologous zwf and gnd from Leuconostoc mesenteroides and Gluconobacter oxydans , respectively, which are less inhibited by NADPH, were overexpressed. The new recombinant produced more ethanol at 1.62 mol mol -1 glucose along with 1.74 mol H 2  mol -1 glucose, which are close to the theoretically maximal yields, 1.67 mol mol -1 each for ethanol and H 2 . However, the attempt to delete the ED pathway in the Δ pgi mutant to operate the PP pathway as the sole glycolytic route, was unsuccessful. By deletion of pgi and overexpression of heterologous zwf and gnd in E. coli Δ hyc

  4. Succinate dehydrogenase assembly factor 2 is needed for assembly and activity of mitochondrial complex II and for normal root elongation in Arabidopsis.

    Science.gov (United States)

    Huang, Shaobai; Taylor, Nicolas L; Ströher, Elke; Fenske, Ricarda; Millar, A Harvey

    2013-02-01

    Mitochondria complex II (succinate dehydrogenase, SDH) plays a central role in respiratory metabolism as a component of both the electron transport chain and the tricarboxylic acid cycle. We report the identification of an SDH assembly factor by analysis of T-DNA insertions in At5g51040, a protein with unknown function that was identified by mass spectrometry analysis as a low abundance mitochondrial protein. This gene is co-expressed with a number of genes encoding mitochondrial proteins, including SDH1-1, and has low partial sequence similarity to human SDHAF2, a protein required for flavin-adenine dinucleotide (FAD) insertion into SDH. In contrast to observations of other SDH deficient lines in Arabidopsis, the sdhaf2 line did not affect photosynthetic rate or stomatal conductance, but instead showed inhibition of primary root elongation with early lateral root emergence, presumably due to the low SDH activity caused by the reduced abundance of SDHAF2. Both roots and leaves showed succinate accumulation but different responses in the abundance of other organic acids and amino acids assayed. Isolated mitochondria showed lowered SDH1 protein abundance, lowered maximal SDH activity and less protein-bound flavin-adenine dinucleotide (FAD) at the molecular mass of SDH1 in the gel separation. The short root phenotype and SDH function of sdhaf2 was fully complemented by transformation with SDHAF2. Application of the SDH inhibitor, malonate, phenocopied the sdhaf2 root architecture in WT. Whole root respiratory assays showed no difference between WT and sdhaf2, but micro-respirometry of the tips of roots clearly showed low oxygen consumption in sdhaf2 which could explain a metabolic deficit responsible for root tip growth. © 2012 The Authors The Plant Journal © 2012 Blackwell Publishing Ltd.

  5. Evaluation of Milk Trace Elements, Lactate Dehydrogenase, Alkaline Phosphatase and Aspartate Aminotransferase Activity of Subclinical Mastitis as and Indicator of Subclinical Mastitis in Riverine Buffalo (

    Directory of Open Access Journals (Sweden)

    Anirban Guha

    2012-03-01

    Full Text Available Mastitis is a highly morbid disease that requires detection at the subclinical stage. Tropical countries like India mainly depend on milch buffaloes for milk. The present study was conducted to investigate whether the trace minerals viz. copper (Cu, iron (Fe, zinc (Zn, cobalt (Co and manganese (Mn and enzyme activity of lactate dehydrogenase (LDH, alkaline phosphatase (ALP and aspartate aminotransferase (AST in riverine buffalo milk can be used as an indicator of subclinical mastitis (SCM with the aim of developing suitable diagnostic kit for SCM. Trace elements and enzyme activity in milk were estimated with Atomic absorption Spectrophotometer, GBC 932 plus and biochemical methods, respectively. Somatic cell count (SCC was done microscopically. The cultural examination revealed Gram positive bacteria as the most prevalent etiological agent. A statistically significant (p<0.01 increase in SCC, Fe, Zn, Co and LDH occurred in SCM milk containing gram positive bacterial agents only. ALP was found to be elevated in milk infected by both gram positive and negative bacteria. The percent sensitivity, specificity and accuracy, predictive values and likelihood ratios were calculated taking bacterial culture examination and SCC≥2×105 cells/ml of milk as the benchmark. Only ALP and Zn, the former being superior, were found to be suitable for diagnosis of SCM irrespective of etiological agents. LDH, Co and Fe can be introduced in the screening programs where Gram positive bacteria are omnipresent. It is recommended that both ALP and Zn be measured together in milk to diagnose buffalo SCM, irrespective of etiology.

  6. Synthesis and SAR Study of Novel Peptide Aldehydes as Inhibitors of 20S Proteasome

    Directory of Open Access Journals (Sweden)

    Lihe Zhang

    2011-09-01

    Full Text Available Based on the analysis of the crystal structure of MG101 (1 and 20S proteasomes, a new series of peptide aldehyde derivatives were designed and synthesized. Their ability to inhibit 20S proteasome was assayed. Among them, Cbz-Glu(OtBu-Phe-Leucinal (3c, Cbz-Glu(OtBu-Leu-Leucinal (3d, and Boc-Ser(OBzl-Leu-Leucinal (3o exhibited the most activity, which represented an order of magnitude enhancement compared with MG132 (2. The covalent docking protocol was used to explore the binding mode. The structure-activity relationship of the peptide aldehyde inhibitors is discussed.

  7. Kinetics and specificity of guinea pig liver aldehyde oxidase and bovine milk xanthine oxidase towards substituted benzaldehydes.

    Science.gov (United States)

    Panoutsopoulos, Georgios I; Beedham, Christine

    2004-01-01

    Molybdenum-containing enzymes, aldehyde oxidase and xanthine oxidase, are important in the oxidation of N-heterocyclic xenobiotics. However, the role of these enzymes in the oxidation of drug-derived aldehydes has not been established. The present investigation describes the interaction of eleven structurally related benzaldehydes with guinea pig liver aldehyde oxidase and bovine milk xanthine oxidase, since they have similar substrate specificity to human molybdenum hydroxylases. The compounds under test included mono-hydroxy and mono-methoxy benzaldehydes as well as 3,4-dihydroxy-, 3-hydroxy-4-methoxy-, 4-hydroxy-3-methoxy-, and 3,4-dimethoxy-benzaldehydes. In addition, various amines and catechols were tested with the molybdenum hydroxylases as inhibitors of benzaldehyde oxidation. The kinetic constants have shown that hydroxy-, and methoxy-benzaldehydes are excellent substrates for aldehyde oxidase (Km values 5x10(-6) M to 1x10(-5) M) with lower affinities for xanthine oxidase (Km values around 10(-4) M). Therefore, aldehyde oxidase activity may be a significant factor in the oxidation of the aromatic aldehydes generated from amines and alkyl benzenes during drug metabolism. Compounds with a 3-methoxy group showed relatively high Vmax values with aldehyde oxidase, whereas the presence of a 3-hydroxy group resulted in minimal Vmax values or no reaction. In addition, amines acted as weak inhibitors, whereas catechols had a more pronounced inhibitory effect on the aldehyde oxidase activity. It is therefore possible that aldehyde oxidase may be critical in the oxidation of the analogous phenylacetaldehydes derived from dopamine and noradrenaline.

  8. [Influence of Hovenia dulcis on alcohol concentration in blood and activity of alcohol dehydrogenase (ADH) of animals after drinking].

    Science.gov (United States)

    Chen, Shao-hong; Zhong, Gan-sheng; Li, Ai-li; Li, Shao-hua; Wu, Li-kun

    2006-07-01

    To observe the effects of H. dulcis on relieving alcohol toxicity by animal experiments. Male kunming mice were ovraiectomized and randomly divided into 5 groups: control group, model group, and aqueous extracts of H. dulcis group at 0.5, 0.25, 0.125 g x mL(-1). The acute alcohlism animals induced by gastral administration with "Er Guo Tou" and the alchol concentrations in serum were detected after treated with the extracts within 0.5-3 h by biochemical enzymes. The alcohol concentration in blood was up to the maximum in 0.5-1.5 h. However, the alcohol concentrations in blood of aqueous extract from H. dulcis group were decreased in 0.5-3 h. The activity of ADH in the liver in aqueous extract of H. dulcis group was increased in 2-3 h, while it was significantly increased in 1-1.5 h (P <0.05). The aqueous extract of H. dulcis could reduce the alchol concentration in blood of animals and inrease the activity of ADH after given alcohol. It means the extract has the effect of relieving alcohol toxicity and preventing drunkenness through restraining the absorption of alcohol in the gastrointestinal tract and promoting the metabolism of alcohol in the liver.

  9. Functional screening of aldehyde decarbonylases for long-chain alkane production by Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Kang, Min-Kyoung; Zhou, Yongjin J.; Buijs, Nicolaas A.

    2017-01-01

    performed functional screening to identify efficient ADs that can improve alkane production by S. cerevisiae. Results: A comparative study of ADs originated from a plant, insects, and cyanobacteria were conducted in S. cerevisiae. As a result, expression of aldehyde deformylating oxygenases (ADOs), which......Background: Low catalytic activities of pathway enzymes are often a limitation when using microbial based chemical production. Recent studies indicated that the enzyme activity of aldehyde decarbonylase (AD) is a critical bottleneck for alkane biosynthesis in Saccharomyces cerevisiae. We therefore...... are cyanobacterial ADs, from Synechococcus elongatus and Crocosphaera watsonii converted fatty aldehydes to corresponding Cn-1 alkanes and alkenes. The CwADO showed the highest alkane titer (0.13 mg/L/OD600) and the lowest fatty alcohol production (0.55 mg/L/OD600). However, no measurable alkanes and alkenes were...

  10. Volatile aldehydes are promising broad-spectrum postharvest insecticides.

    Science.gov (United States)

    Hammond, D G; Rangel, S; Kubo, I

    2000-09-01

    A variety of naturally occurring aldehydes common in plants have been evaluated for their insecticidal activity and for phytotoxicity to postharvest fruits, vegetables, and grains. Twenty-nine compounds were initially screened for their activity against aphids on fava bean leaf disks. Application under reduced pressure (partial vacuum) for the first quarter of fumigation increased insecticidal activity severalfold. The 11 best aldehydes were assayed against aphids placed under the third leaf of whole heads of iceberg lettuce using the same two-tier reduced-pressure regime, which caused no additional detriment to the commodity over fumigation at atmospheric pressure. Phytotoxicity to naked and wrapped iceburg lettuce, green and red table grapes, lemon, grapefruit, orange, broccoli, avocado, cabbage, pinto bean, and rice at doses that killed 100% of aphids was recorded for three promising fumigants: propanal, (E)-2-pentenal, and 2-methyl-(E)-2-butenal. These three compounds have excellent potential as affordable postharvest insect control agents, killing 100% of the aphids with little or no detectable harm to a majority of the commodities tested. Preliminary assays indicate that similar doses are also effective against mealybugs, thrips, and whitefly.

  11. Biossensor enzimático para detecção de fungicidas ditiocarbamatos: estudo cinético da enzima aldeído desidrogenase e otimização do biossensor Enzymatic biosensor for the detection of dithiocarbamate fungicides: kinetic study of aldehyde dehydrogenase enzyme and biosensor optimization

    Directory of Open Access Journals (Sweden)

    Roberval Soares Lima

    2007-02-01

    Full Text Available Initially, all major factors that affect the rate of the AldH-catalyzed reaction (enzyme concentration, substrate concentration, temperature and pH were investigated. Optimal activity was observed between pH values of 7.5 and 9.5 in the temperature range of 25 to 50 ºC. Kinetic parameters, such as Km (2.92 µmol L-1 and Vmax (1.33 10-2 µmol min-1 demonstrate a strong enzyme-substrate affinity. The sensors were based on screen-printed electrodes modified with the Meldola Blue-Reinecke salt (MBRS combination. Operational conditions (NAD+ and substrate contents, enzyme loading and response time were optimized. Also, two enzyme immobilization procedures were tested: entrapment in poly(vinyl alcohol bearing styrylpyridinium groups (PVA-SbQ and crosslinking with glutaraldehyde. Chronoamperometry was employed to observe the biosensor responses during enzymatic hydrolysis of propionaldehyde and also to construct inhibition curves with maneb and zineb fungicides. Best results were found with the following conditions: [NAD+] = 0.25 mmol L-1; [propionaldehyde] = 80 µmol L-1; enzyme loading = 0.8 U per electrode; response time = 10 min, and inhibition time = 10 min. Current intensities around 103 ± 13 nA with the sensors and good stability was obtained for both immobilization procedures. Detection limits, calculated using 10% inhibition were 31.5 µg L-1 and 35 µg L-1 for maneb and zineb, respectively. Results obtained with other MBRS-modified electrodes consisting of mono and bi-enzymic sensors were compared. The ability to catalyze NADH oxidation by MB was also highlighted.

  12. Reduced activity of 11beta-hydroxysteroid dehydrogenase type 2 is not responsible for sodium retention in nephrotic rats

    DEFF Research Database (Denmark)

    Bistrup, C; Thiesson, H C; Jensen, B L

    2005-01-01

    has been suggested to allow glucocorticoids to stimulate MR, thereby contributing to sodium retention. We tested this hypothesis in the puromycin aminonucleoside model of nephrotic syndrome in rats. METHODS: Complete sodium and potassium intakes and excretions (faeces and urine) were measured in rats......)] to suppress endogenous glucocorticoids in the proteinuric stage during active sodium retention. RESULTS: Nephrotic rats developed proteinuria, positive sodium balance, decreased plasma aldosterone concentration, and decreased urinary Na(+)/K(+) ratio. 11betaHSD2 mRNA expression was down-regulated but protein...... expression was unchanged. SGK mRNA and phosphorylated SGK protein were up-regulated while total SGK protein expression was unchanged. Dexamethasone treatment, which suppressed plasma corticosterone concentration, did not correct sodium balance or fluid retention in nephrotic rats. CONCLUSION: Our results do...

  13. Hibernation impact on the catalytic activities of the mitochondrial D-3-hydroxybutyrate dehydrogenase in liver and brain tissues of jerboa (Jaculus orientalis

    Directory of Open Access Journals (Sweden)

    Hafiani Assia

    2003-09-01

    Full Text Available Abstract Background Jerboa (Jaculus orientalis is a deep hibernating rodent native to subdesert highlands. During hibernation, a high level of ketone bodies i.e. acetoacetate (AcAc and D-3-hydroxybutyrate (BOH are produced in liver, which are used in brain as energetic fuel. These compounds are bioconverted by mitochondrial D-3-hydroxybutyrate dehydrogenase (BDH E.C. 1.1.1.30. Here we report, the function and the expression of BDH in terms of catalytic activities, kinetic parameters, levels of protein and mRNA in both tissues i.e brain and liver, in relation to the hibernating process. Results We found that: 1/ In euthemic jerboa the specific activity in liver is 2.4- and 6.4- fold higher than in brain, respectively for AcAc reduction and for BOH oxidation. The same differences were found in the hibernation state. 2/ In euthermic jerboa, the Michaelis constants, KM BOH and KM NAD+ are different in liver and in brain while KM AcAc, KM NADH and the dissociation constants, KD NAD+and KD NADH are similar. 3/ During prehibernating state, as compared to euthermic state, the liver BDH activity is reduced by half, while kinetic constants are strongly increased except KD NAD+. 4/ During hibernating state, BDH activity is significantly enhanced, moreover, kinetic constants (KM and KD are strongly modified as compared to the euthermic state; i.e. KD NAD+ in liver and KM AcAc in brain decrease 5 and 3 times respectively, while KD NADH in brain strongly increases up to 5.6 fold. 5/ Both protein content and mRNA level of BDH remain unchanged during the cold adaptation process. Conclusions These results cumulatively explained and are consistent with the existence of two BDH enzymatic forms in the liver and the brain. The apoenzyme would be subjected to differential conformational folding depending on the hibernation state. This regulation could be a result of either post-translational modifications and/or a modification of the mitochondrial membrane state

  14. Aldehyde decarbonylation catalysis under mild conditions

    Energy Technology Data Exchange (ETDEWEB)

    Beck, C.M.; Rathmill, S.E.; Park, Y.J.; Chen, J.; Crabtree, R.H.; Liable-Sands, L.M.; Rheingold, A.L.

    1999-12-06

    Reaction of [RhCl(NBD)]{sub 2} with 2.0 equiv of triphos (triphos = bis(2-diphenylphosphinoethyl)phenylphosphine; NBD = bicyclo[2.2.1]hepta-2,5-diene) in THF solution at room temperature affords [Rh(NBD)(triphos)][Cl] (4a), which was isolated as [Rh(NBD)(triphos)][SbF{sub 6}] (4b) in 67% yield. Treatment of 4b with aqueous formaldehyde in THF solution at 80 C forms [Rh(CO)(triphos)][SbF{sub 6}] (2a), which reversibly binds a second equivalent of CO{sub (g)} to give [Rh(CO){sub 2}(triphos)][SbF{sub 6}] (2b). The complex [Rh(CO)(triphos)][SbF{sub 6}] has been found to be an effective aldehyde decarbonylation catalyst for primary and aryl aldehydes at temperatures as low as that of refluxing dioxane, with little or no undesirable side products resulting from {beta} elimination or radical rearrangement.

  15. Toxic Diatom Aldehydes Affect Defence Gene Networks in Sea Urchins.

    Science.gov (United States)

    Varrella, Stefano; Romano, Giovanna; Costantini, Susan; Ruocco, Nadia; Ianora, Adrianna; Bentley, Matt G; Costantini, Maria

    2016-01-01

    Marine organisms possess a series of cellular strategies to counteract the negative effects of toxic compounds, including the massive reorganization of gene expression networks. Here we report the modulated dose-dependent response of activated genes by diatom polyunsaturated aldehydes (PUAs) in the sea urchin Paracentrotus lividus. PUAs are secondary metabolites deriving from the oxidation of fatty acids, inducing deleterious effects on the reproduction and development of planktonic and benthic organisms that feed on these unicellular algae and with anti-cancer activity. Our previous results showed that PUAs target several genes, implicated in different functional processes in this sea urchin. Using interactomic Ingenuity Pathway Analysis we now show that the genes targeted by PUAs are correlated with four HUB genes, NF-κB, p53, δ-2-catenin and HIF1A, which have not been previously reported for P. lividus. We propose a working model describing hypothetical pathways potentially involved in toxic aldehyde stress response in sea urchins. This represents the first report on gene networks affected by PUAs, opening new perspectives in understanding the cellular mechanisms underlying the response of benthic organisms to diatom exposure.

  16. Toxic Diatom Aldehydes Affect Defence Gene Networks in Sea Urchins.

    Directory of Open Access Journals (Sweden)

    Stefano Varrella

    Full Text Available Marine organisms possess a series of cellular strategies to counteract the negative effects of toxic compounds, including the massive reorganization of gene expression networks. Here we report the modulated dose-dependent response of activated genes by diatom polyunsaturated aldehydes (PUAs in the sea urchin Paracentrotus lividus. PUAs are secondary metabolites deriving from the oxidation of fatty acids, inducing deleterious effects on the reproduction and development of planktonic and benthic organisms that feed on these unicellular algae and with anti-cancer activity. Our previous results showed that PUAs target several genes, implicated in different functional processes in this sea urchin. Using interactomic Ingenuity Pathway Analysis we now show that the genes targeted by PUAs are correlated with four HUB genes, NF-κB, p53, δ-2-catenin and HIF1A, which have not been previously reported for P. lividus. We propose a working model describing hypothetical pathways potentially involved in toxic aldehyde stress response in sea urchins. This represents the first report on gene networks affected by PUAs, opening new perspectives in understanding the cellular mechanisms underlying the response of benthic organisms to diatom exposure.

  17. Seasonal and diel activity of Ixodes ricinus (Acari: Ixodidae) subpopulations in Denmark. Aspects of size, physiological age, and malate dehydrogenase genotype in a forest site without any undergrowth.

    Science.gov (United States)

    Jensen, P M; Kaufmann, U

    2003-01-01

    The underlying population dynamics and the behavioural patterns of the vectors are key issues in understanding the transmission of vector borne pathogens. For the tick Ixodes ricinus both seasonal and diel activity have been described as bimodal patterns, which in seasonal aspect has been interpreted as representing two cohorts. However, recent studies have shown that this interpretation may be incorrect. The aim of this study was to obtain more detailed information on nymph host seeking by studying subpopulations of ticks during the day and season. The study was designed to allow for comparisons of the diel variation and seasonal variation in their dependency in a number of tick characteristics. The study took place in a forest with planted beech trees without any undergrowth. Ticks were collected by flagging the dead leaves on the forest floor. For each nymph, a number of visual observations were made. The size and physiological age was observed and the nymphs were genotyped in the malate dehydrogenase locus (MDH, E.C. 1.1.1.37). Briefly the main results can be given as: (i) There were significant differences in the composition of size classes during the season, but only limited trends in time. (ii) The proportion of the small nymphs was highly variable, with a variation from 3% to 24% in October and September, respectively. (iii) The diel variation in MDH genotypes was significant in May and August. (iv) Nymph size classes and physiological age appeared to interact. The non-random interaction was caused by a relatively even distribution of small nymphs in all four age classes, while large nymphs tended to fall into age class 2 and 3. The length by age interaction for the individual months was noted to be significant in May, July, August and September, but not in June. Similarly the interaction was significant in the morning and afternoon, but not at midday. The overall results describe the seasonal and diel activity patterns as changing systematically for

  18. Host cell and expression engineering for development of an E. coli ketoreductase catalyst: Enhancement of formate dehydrogenase activity for regeneration of NADH

    Directory of Open Access Journals (Sweden)

    Mädje Katharina

    2012-01-01

    Full Text Available Abstract Background Enzymatic NADH or NADPH-dependent reduction is a widely applied approach for the synthesis of optically active organic compounds. The overall biocatalytic conversion usually involves in situ regeneration of the expensive NAD(PH. Oxidation of formate to carbon dioxide, catalyzed by formate dehydrogenase (EC 1.2.1.2; FDH, presents an almost ideal process solution for coenzyme regeneration that has been well established for NADH. Because isolated FDH is relatively unstable under a range of process conditions, whole cells often constitute the preferred form of the biocatalyst, combining the advantage of enzyme protection in the cellular environment with ease of enzyme production. However, the most prominent FDH used in biotransformations, the enzyme from the yeast Candida boidinii, is usually expressed in limiting amounts of activity in the prime host for whole cell biocatalysis, Escherichia coli. We therefore performed expression engineering with the aim of enhancing FDH activity in an E. coli ketoreductase catalyst. The benefit resulting from improved NADH regeneration capacity is demonstrated in two transformations of technological relevance: xylose conversion into xylitol, and synthesis of (S-1-(2-chlorophenylethanol from o-chloroacetophenone. Results As compared to individual expression of C. boidinii FDH in E. coli BL21 (DE3 that gave an intracellular enzyme activity of 400 units/gCDW, co-expression of the FDH with the ketoreductase (Candida tenuis xylose reductase; XR resulted in a substantial decline in FDH activity. The remaining FDH activity of only 85 U/gCDW was strongly limiting the overall catalytic activity of the whole cell system. Combined effects from increase in FDH gene copy number, supply of rare tRNAs in a Rosetta strain of E. coli, dampened expression of the ketoreductase, and induction at low temperature (18°C brought up the FDH activity threefold to a level of 250 U/gCDW while reducing the XR activity by

  19. Characterization and Targeting of the Aldehyde Dehydrogenase Subpopulation in Ovarian Cancer

    Science.gov (United States)

    2013-07-01

    pancreatic cancer. Pancreas. 2008 Oct;37(3):275-81. 28. Hu D, Wang X, Mao Y, Zhou L. Identification of CD105 (endoglin)-positive stem-like cells in...Women who have breaks in ovulation due to pregnancy and breast- feeding have lower risk of disease.19,20 Moreover, women who take oral con

  20. Characterization and Targeting of the Aldehyde Dehydrogenase Subpopulation in Ovarian Cancer

    Science.gov (United States)

    2014-07-01

    modification; however, for publication of PCNA figures, contrast was enhanced to an entire image by using the "Auto Contrast" tool in Photoshop to avoid bias...Page Introduction…………………………………………………………….………..….. 1 Body ...provide the opportunity to more fully characterize which cells are mediating survival of primary therapy. BODY : Task 1: Determine

  1. Characterization and Targeting of the Aldehyde Dehydrogenase Subpopulation in Ovarian Cancer

    Science.gov (United States)

    2015-07-01

    Sharp BA, Underwood PB. Cancer patients’ satisfaction with physicians: PMH-SPQ-MD questionnaire results. Am J Obstet Gynecol 188(5):1177-1179, 2003...outcome in uterine cancer : Molecular explanations. Proceedings of the 39th Annual Society of Gynecologic Oncologists Meeting, 2008. Page 16 Revised 8/20...basis for the impact of EphA2 overexpression on clinical outcome in uterine cancer Proceedings of the American Association of Cancer Research, 2008

  2. Peroxisome proliferator-activated receptor-gamma stimulates 11beta-hydroxysteroid dehydrogenase type 1 in rat vascular smooth muscle cells

    Czech Academy of Sciences Publication Activity Database

    Vagnerová, Karla; Loukotová, Jana; Ergang, Peter; Musílková, Jana; Mikšík, Ivan; Pácha, Jiří

    2011-01-01

    Roč. 76, č. 6 (2011), s. 577-581 ISSN 0039-128X R&D Projects: GA ČR(CZ) GAP303/10/0969 Institutional research plan: CEZ:AV0Z50110509 Keywords : 11beta-hydroxysteroid dehydrogenase * thiazolidinediones Subject RIV: ED - Physiology Impact factor: 2.829, year: 2011

  3. Catalytic production of methyl acrylates by gold-mediated cross coupling of unsaturated aldehydes with methanol

    Science.gov (United States)

    Karakalos, Stavros; Zugic, Branko; Stowers, Kara J.; Biener, Monika M.; Biener, Juergen; Friend, Cynthia M.; Madix, Robert J.

    2016-10-01

    Modern methods of esterification, one of the most important reactions in organic synthesis, are reaching their limits, as far as waste and expense are concerned. Novel chemical approaches to ester formation are therefore of importance. Here we report a simple procedure free of caustic reagents or byproducts for the facile direct oxidative methyl esterification of aldehydes over nanoporous Au catalysts. Complementary model studies on single crystal gold surfaces establish the fundamental reactions involved. We find that methanol more readily reacts with adsorbed active oxygen than do the aldehydes, but that once the aldehydes do react, they form strongly-bound acrylates that block reactive sites and decrease the yields of acrylic esters under steady flow conditions at 420 K. Significant improvements in yield can be achieved by operating at higher temperatures, which render the site-blocking acrylates unstable.

  4. Red Xylem and Higher Lignin Extractability by Down-Regulating a Cinnamyl Alcohol Dehydrogenase in Poplar.

    Science.gov (United States)

    Baucher, M.; Chabbert, B.; Pilate, G.; Van Doorsselaere, J.; Tollier, M. T.; Petit-Conil, M.; Cornu, D.; Monties, B.; Van Montagu, M.; Inze, D.; Jouanin, L.; Boerjan, W.

    1996-12-01

    Cinnamyl alcohol dehydrogenase (CAD) catalyzes the last step in the biosynthesis of the lignin precursors, the monolignols. We have down-regulated CAD in transgenic poplar (Populus tremula X Populus alba) by both antisense and co-suppression strategies. Several antisense and sense CAD transgenic poplars had an approximately 70% reduced CAD activity that was associated with a red coloration of the xylem tissue. Neither the lignin amount nor the lignin monomeric composition (syringyl/guaiacyl) were significantly modified. However, phloroglucinol-HCl staining was different in the down-regulated CAD plants, suggesting changes in the number of aldehyde units in the lignin. Furthermore, the reactivity of the cell wall toward alkali treatment was altered: a lower amount of lignin was found in the insoluble, saponified residue and more lignin could be precipitated from the soluble alkali fraction. Moreover, large amounts of phenolic compounds, vanillin and especially syringaldehyde, were detected in the soluble alkali fraction of the CAD down-regulated poplars. Alkaline pulping experiments on 3-month-old trees showed a reduction of the kappa number without affecting the degree of cellulose degradation. These results indicate that reducing the CAD activity in trees might be a valuable strategy to optimize certain processes of the wood industry, especially those of the pulp and paper industry.

  5. Temperature-sensitive glutamate dehydrogenase mutants of Salmonella typhimurium.

    OpenAIRE

    Dendinger, S M; Brenchley, J E

    1980-01-01

    Mutants of Salmonella typhimurium defective in glutamate dehydrogenase activity were isolated in parent strains lacking glutamate synthase activity by localizcd mutagenesis or by a general mutagenesis combined with a cycloserine enrichment for glutamate auxotrophs. Two mutants with temperature-sensitive phenotypes had glutamate dehydrogenase activities that were more thermolabile than that of an isogenic control strain. Eight other mutants had less than 10% of the wild-type glutamate dehydrog...

  6. Structural Determinants of Oligomerization of !1-Pyrroline-5-Carboxylate Dehydrogenase: Identification of a Hexamerization Hot Spot

    Science.gov (United States)

    Luo, Min; Singh, Ranjan K.; Tanner, John J.

    2013-01-01

    The aldehyde dehydrogenase (ALDH) superfamily member !1-pyrroline-5-carboxylate dehydrogenase (P5CDH) catalyzes the NAD+-dependent oxidation of glutamate semialdehyde to glutamate, which is the final step of proline catabolism. Defects in P5CDH activity lead to the metabolic disorder type II hyperprolinemia, P5CDH is essential for virulence of the fungal pathogen Cryptococcus neoformans, and bacterial P5CDHs have been targeted for vaccine development. Although the enzyme oligomeric state is known to be important for ALDH function, the oligomerization of P5CDH has remained relatively unstudied. Here we determine the oligomeric states and quaternary structures of four bacterial P5CDHs using a combination of small-angle X-ray scattering, X-ray crystallography, and dynamic light scattering. The P5CDHs from Thermus thermophilus and Deinococcus radiodurans form trimer-of-dimers hexamers in solution, which is the first observation of a hexameric ALDH in solution. In contrast, two Bacillus P5CDHs form dimers in solution but do not assemble into a higher order oligomer. Site-directed mutagenesis was used to identify a hexamerization hot spot that is centered on an arginine residue in the NAD+-binding domain. Mutation of this critical Arg residue to Ala in either of the hexameric enzymes prevents hexamer formation in solution. Paradoxically, the dimeric Arg-to-Ala T. thermophilus mutant enzyme packs as a hexamer in the crystal state, which illustrates the challenges associated with predicting the biological assembly in solution from crystal structures. The observation of different oligomeric states among P5CDHs suggests potential differences in cooperativity and protein-protein interactions. PMID:23747974

  7. Cloning and sequencing of the gene encoding the 72-kilodalton dehydrogenase subunit of alcohol dehydrogenase from Acetobacter aceti.

    Science.gov (United States)

    Inoue, T; Sunagawa, M; Mori, A; Imai, C; Fukuda, M; Takagi, M; Yano, K

    1989-06-01

    A genomic library of Acetobacter aceti DNA was constructed by using a broad-host-range cosmid vector. Complementation of a spontaneous alcohol dehydrogenase-deficient mutant resulted in the isolation of a plasmid designated pAA701. Subcloning and deletion analysis of pAA701 limited the region that complemented the deficiency in alcohol dehydrogenase activity of the mutant. The nucleotide sequence of this region was determined and showed that this region contained the full structural gene for the 72-kilodalton dehydrogenase subunit of the alcohol dehydrogenase enzyme complex. The predicted amino acid sequence of the gene showed homology with sequences of methanol dehydrogenase structural genes of Paracoccus denitrificans and Methylobacterium organophilum.

  8. MOLECULAR MODELLING OF HUMAN ALDEHYDE OXIDASE AND IDENTIFICATION OF THE KEY INTERACTIONS IN THE ENZYME-SUBSTRATE COMPLEX

    Directory of Open Access Journals (Sweden)

    Siavoush Dastmalchi

    2005-05-01

    Full Text Available Aldehyde oxidase (EC 1.2.3.1, a cytosolic enzyme containing FAD, molybdenum and iron-sulphur cluster, is a member of non-cytochrome P-450 enzymes called molybdenum hydroxylases which is involved in the metabolism of a wide range of endogenous compounds and many drug substances. Drug metabolism is one of the important characteristics which influences many aspects of a therapeutic agent such as routes of administration, drug interaction and toxicity and therefore, characterisation of the key interactions between enzymes and substrates is very important from drug development point of view. The aim of this study was to generate a three-dimensional model of human aldehyde oxidase (AO in order to assist us to identify the mode of interaction between enzyme and a set of phethalazine/quinazoline derivatives. Both sequence-based (BLAST and inverse protein fold recognition methods (THREADER were used to identify the crystal structure of bovine xanthine dehydrogenase (pdb code of 1FO4 as the suitable template for comparative modelling of human AO. Model structure was generated by aligning and then threading the sequence of human AO onto the template structure, incorporating the associated cofactors, and molecular dynamics simulations and energy minimization using GROMACS program. Different criteria which were measured by the PROCHECK, QPACK, VERIFY-3D were indicative of a proper fold for the predicted structural model of human AO. For example, 97.9 percentages of phi and psi angles were in the favoured and most favoured regions in the ramachandran plot, and all residues in the model are assigned environmentally positive compatibility scores. Further evaluation on the model quality was performed by investigation of AO-mediated oxidation of a set of phthalazine/quinazoline derivatives to develop QSAR model capable of describing the extent of the oxidation. Substrates were aligned by docking onto the active site of the enzyme using GOLD technology and then

  9. Aldehyde stress-mediated novel modification of proteins: epimerization of the N-terminal amino acid.

    Science.gov (United States)

    Kajita, Ryo; Goto, Takaaki; Lee, Seon Hwa; Oe, Tomoyuki

    2013-12-16

    Various kinds of aldehyde-mediated chemical modifications of proteins have been identified as being exclusively covalent. We report a unique noncovalent modification: the aldehyde-mediated epimerization of the N-terminal amino acid. Epimerization of amino acids is thought to cause conformational changes that alter their biological activity. However, few mechanistic studies have been performed, because epimerization of an amino acid is a miniscule change in a whole protein. Furthermore, it does not produce a mass shift, making mass spectrometric analysis difficult. Here, we have demonstrated epimerization mediated by endogenous aldehydes. A model peptide, with an N-terminal l- or d-FMRFamide, was incubated with an endogenous or synthetic aldehyde [acetaldehyde, methylglyoxal, pyridoxal 5'-phosphate (PLP), 4-oxo-2(E)-nonenal, 4-hydroxy-2(E)-nonenal, d-glucose (Glc), 4- or 2-pyridinecarboxaldehyde] under physiological conditions. Each reaction mixture was analyzed by liquid chromatography with ultraviolet detection and/or electrospray ionization mass spectrometry. Considerable epimerization occurred after incubation with some endogenous aldehydes (PLP, 40.6% after 1 day; Glc with copper ions, 6.5% after 7 days). Moreover, the epimerization also occurred in whole proteins (human serum albumin and PLP, 26.3% after 1 day). Tandem mass spectrometric studies, including deuterium labeling and sodium borohydride reduction, suggested that the epimerization results from initial Schiff base formation followed by tautomerization to ketimine that causes the chirality to be lost. This suggests that the epimerization of the N-terminal amino acid can also occur in vivo as a post-translational modification under a high level of aldehyde stress.

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

    International Nuclear Information System (INIS)

    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. 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 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

  11. Cortisol metabolism in healthy young adults: sexual dimorphism in activities of A-ring reductases, but not 11beta-hydroxysteroid dehydrogenases.

    Science.gov (United States)

    Finken, M J; Andrews, R C; Andrew, R; Walker, B R

    1999-09-01

    Cortisol is metabolized irreversibly by A-ring reductases (5alpha- and 5beta-reductases) and reversibly (to cortisone) by 11beta-hydroxysteroid dehydrogenases (11betaHSDs). In rats, estradiol down-regulates 11betaHSD1 expression. In humans, ratios of urinary cortisol/cortisone metabolites differ in men and women. In this study, urinary cortisol metabolites and hepatic 11betaHSD1 activity were measured in healthy young men and women at different phases of the menstrual cycle. Ten men and 10 women with regular menstrual cycles collected a 24-h urine sample, took 250 microg oral dexamethasone at 2300 h, took 25 mg oral cortisone at 0900 h (after fasting), and had blood sampled for plasma cortisol estimation over the subsequent 150 min. Women repeated the tests in random order in menstrual, follicular, and luteal phases. Women excreted disproportionately less A-ring-reduced metabolites of cortisol [median 5alpha-tetrahydrocortisol, 1811 (interquartile range, 1391-2300) microg/day in menstrual phase vs. 2723 (interquartile range, 2454-3154) in men (P = 0.01); 5beta-tetrahydrocortisol, 1600 (interquartile range, 1419-1968) vs. 2197 (interquartile range, 1748-2995; P = 0.03)] but similar amounts of cortisol, cortisone, and tetrahydrocortisone. Analogous differences were observed in urinary excretion of androgen metabolites. Conversion of cortisone to cortisol on hepatic first pass metabolism was not different (peak plasma cortisol, 733 +/- 60 nmol/L in women vs. 684 +/- 53 nmol/L in men; mean +/- SEM; P = 0.55). There were no differences in cortisol or androgen metabolism between phases of the menstrual cycle. We conclude that sexual dimorphism in cortisol metabolite excretion is attributable to less A-ring reduction of cortisol in women, rather than less reactivation of cortisone to cortisol by 11betaHSD1. This difference is not influenced acutely by gonadal steroids. 11BetaHSD1 has been suggested to modulate insulin sensitivity and body fat distribution, but caution

  12. Substrate specificity of an aflatoxin-metabolizing aldehyde reductase.

    Science.gov (United States)

    Ellis, E M; Hayes, J D

    1995-01-01

    The enzyme from rat liver that reduces aflatoxin B1-dialdehyde exhibits a unique catalytic specificity distinct from that of other aldo-keto reductases. This enzyme, designated AFAR, displays high activity towards dicarbonyl-containing compounds with ketone groups on adjacent carbon atoms; 9,10-phenanthrenequinone, acenaphthenequinone and camphorquinone were found to be good substrates. Although AFAR can also reduce aromatic and aliphatic aldehydes such as succinic semialdehyde, it is inactive with glucose, galactose and xylose. The enzyme also exhibits low activity towards alpha,beta-unsaturated carbonyl-containing compounds. Determination of the apparent Km reveals that AFAR has highest affinity for 9,10-phenanthrenequinone and succinic semialdehyde, and low affinity for glyoxal and DL-glyceraldehyde. PMID:8526867

  13. Probing Lactate Dehydrogenase Activity in Tumors by Measuring Hydrogen/Deuterium Exchange in Hyperpolarized l-[1-13C,U-2H]Lactate

    OpenAIRE

    Kennedy, Brett W. C.; Kettunen, Mikko I.; Hu, De-En; Brindle, Kevin M.

    2012-01-01

    13C magnetic resonance spectroscopy and spectroscopic imaging measurements of hyperpolarized 13C label exchange between exogenously administered [1-13C]pyruvate and endogenous lactate, catalyzed by lactate dehydrogenase (LDH), has proved to be a powerful approach for probing tissue metabolism in vivo. This experiment has clinical potential, particularly in oncology, where it could be used to assess tumor grade and response to treatment. A limitation of the method is that pyruvate must be admi...

  14. Effect of exogenous cytokinins, auxins and adenine on cytokinin N-glucosylation and cytokinin oxidase/dehydrogenase activity in de-rooted radish seedlings

    Czech Academy of Sciences Publication Activity Database

    Blagoeva, Elitsa; Dobrev, Petre; Malbeck, Jiří; Motyka, Václav; Gaudinová, Alena; Vaňková, Radomíra

    2004-01-01

    Roč. 44, č. 1 (2004), s. 15-23 ISSN 0167-6903 R&D Projects: GA ČR GA522/99/1130; GA AV ČR IAA6038002; GA MŠk LN00A081; GA MŠk ME 505 Institutional research plan: CEZ:AV0Z5038910 Keywords : Auxin * Cytokinin * Cytokinin oxidase/dehydrogenase Subject RIV: GE - Plant Breeding Impact factor: 0.693, year: 2004

  15. A trade off between catalytic activity and protein stability determines the clinical manifestations of glucose-6-phosphate dehydrogenase (G6PD) deficiency

    OpenAIRE

    Boonyuen, Usa; Chamchoy, Kamonwan; Swangsri, Thitiluck; Junkree, Thanyaphorn; Day, Nicholas P.J.; White, Nicholas J.; Imwong, Mallika

    2017-01-01

    Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common polymorphism and enzymopathy in humans, affecting approximately 400 million people worldwide. It is responsible for various clinical manifestations, including favism, hemolytic anemia, chronic non-spherocytic hemolytic anemia, spontaneous abortion, and neonatal hyperbilirubinemia. Understanding the molecular mechanisms underlying the severity of G6PD deficiency is of great importance but that of many G6PD variants are stil...

  16. Genetics Home Reference: lactate dehydrogenase deficiency

    Science.gov (United States)

    ... this condition: lactate dehydrogenase-A deficiency (sometimes called glycogen storage disease XI) and lactate dehydrogenase-B deficiency. People with ... Resources Genetic Testing (2 links) Genetic Testing Registry: Glycogen storage disease XI Genetic Testing Registry: Lactate dehydrogenase B deficiency ...

  17. External NAD(P)H dehydrogenases in Acanthamoeba castellanii mitochondria.

    Science.gov (United States)

    Antos-Krzeminska, Nina; Jarmuszkiewicz, Wieslawa

    2014-09-01

    The mitochondrial respiratory chain of plants and some fungi contains multiple rotenone-insensitive NAD(P)H dehydrogenases, of which at least two are located on the outer surface of the inner membrane (i.e., external NADH and external NADPH dehydrogenases). Annotated sequences of the putative alternative NAD(P)H dehydrogenases of the protozoan Acanthamoeba castellanii demonstrated similarity to plant and fungal sequences. We also studied activity of these dehydrogenases in isolated A. castellanii mitochondria. External NADPH oxidation was observed for the first time in protist mitochondria. The coupling parameters were similar for external NADH oxidation and external NADPH oxidation, indicating similar efficiencies of ATP synthesis. Both external NADH oxidation and external NADPH oxidation had an optimal pH of 6.8 independent of relevant ubiquinol-oxidiz