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Sample records for succinate dehydrogenase genes

  1. Frequent germ-line succinate dehydrogenase subunit D gene mutations in patients with apparently sporadic parasympathetic paraganglioma

    NARCIS (Netherlands)

    H. Dannenberg (Hilde); W.N.M. Dinjens (Winand); M. Abbou; H. van Urk (Hero); B.K. Pauw; D. Mouwen; W.J. Mooi (Wolter); R.R. de Krijger (Ronald)

    2002-01-01

    textabstractPURPOSE: Recently, familial paraganglioma (PGL) was shown to be caused bymutations in the gene encoding succinate dehydrogenase subunit D (SDHD). However, the prevalence of SDHD mutations in apparently sporadic PGL is unknown. We studied the frequency and spectrum of

  2. Cloning and functional analysis of succinate dehydrogenase gene PsSDHA in Phytophthora sojae.

    Science.gov (United States)

    Pan, Yuemin; Ye, Tao; Gao, Zhimou

    2017-07-01

    Succinate dehydrogenase (SDH) is one of the key enzymes of the tricarboxylic acid cycle (TCA cycle) and a proven target of fungicides for true fungi. To explore the roles of the SDHA gene in Phytophthora sojae, we first cloned PsSDHA to construct the PsSDHA silenced expression vector pHAM34-PsSDHA, and then utilized PEG to mediate the P. sojae protoplast transformation experiment. Through transformation screening, we obtained the silenced mutants A1 and A3, which have significant suppressive effect. Further study showed that the hyphae of the silenced mutant strains were shorter and more bifurcated; the growth of the silenced mutants was clearly inhibited in 10% V8 agar medium containing sodium chloride (NaCl), hydrogen peroxide (H 2 O 2 ) or Congo Red, respectively. The pathogenicity of the silenced mutants was significantly reduced compared with the wild-type strain and the mock. The results could help us better to understand the position and function of SDH in P. sojae and provide a proven target of fungicides for the oomycete. Copyright © 2017. Published by Elsevier Ltd.

  3. No evidence for promoter region methylation of the succinate dehydrogenase and fumarate hydratase tumour suppressor genes in breast cancer

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    Dobrovic Alexander

    2009-09-01

    Full Text Available Abstract Background Succinate dehydrogenase (SDH and fumarate hydratase (FH are tricarboxylic acid (TCA cycle enzymes that are also known to act as tumour suppressor genes. Increased succinate or fumarate levels as a consequence of SDH and FH deficiency inhibit hypoxia inducible factor-1α (HIF-1α prolyl hydroxylases leading to sustained HIF-1α expression in tumours. Since HIF-1α is frequently expressed in breast carcinomas, DNA methylation at the promoter regions of the SDHA, SDHB, SDHC and SDHD and FH genes was evaluated as a possible mechanism in silencing of SDH and FH expression in breast carcinomas. Findings No DNA methylation was identified in the promoter regions of the SDHA, SDHB, SDHC, SDHD and FH genes in 72 breast carcinomas and 10 breast cancer cell lines using methylation-sensitive high resolution melting which detects both homogeneous and heterogeneous methylation. Conclusion These results show that inactivation via DNA methylation of the promoter CpG islands of SDH and FH is unlikely to play a major role in sporadic breast carcinomas.

  4. Linkage and radiation hybrid mapping of the porcine gene for subunit C of succinate dehydrogenase complex (SDHC)

    Czech Academy of Sciences Publication Activity Database

    Stratil, Antonín; Reiner, G.; Peelman, L. J.; Poucke, M.; Geldermann, H.

    2001-01-01

    Roč. 32, č. 2 (2001), s. 110-112 ISSN 0268-9146 R&D Projects: GA AV ČR KSK5052113; GA ČR GA523/00/0669 Keywords : succinate dehydrogenase complex Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.020, year: 2001

  5. Krebs cycle metabolite profiling for identification and stratification of pheochromocytomas/paragangliomas due to succinate dehydrogenase deficiency

    NARCIS (Netherlands)

    Richter, S; Peitzsch, M.; Rapizzi, E.; Lenders, J.W.M.; Qin, N.; Cubas, A.A. de; Schiavi, F.; Rao, J.U.; Beuschlein, F.; Quinkler, M.; Timmers, H.J.L.M.; Opocher, G.; Mannelli, M.; Pacak, K.; Robledo, M.; Eisenhofer, G.

    2014-01-01

    CONTEXT: Mutations of succinate dehydrogenase A/B/C/D genes (SDHx) increase susceptibility to development of pheochromocytomas and paragangliomas (PPGLs), with particularly high rates of malignancy associated with SDHB mutations. OBJECTIVE: We assessed whether altered succinate dehydrogenase

  6. A split and rearranged nuclear gene encoding the iron-sulfur subunit of mitochondrial succinate dehydrogenase in Euglenozoa

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    Gray Michael W

    2009-02-01

    Full Text Available Abstract Background Analyses based on phylogenetic and ultrastructural data have suggested that euglenids (such as Euglena gracilis, trypanosomatids and diplonemids are members of a monophyletic lineage termed Euglenozoa. However, many uncertainties are associated with phylogenetic reconstructions for ancient and rapidly evolving groups; thus, rare genomic characters become increasingly important in reinforcing inferred phylogenetic relationships. Findings We discovered that the iron-sulfur subunit (SdhB of mitochondrial succinate dehydrogenase is encoded by a split and rearranged nuclear gene in Euglena gracilis and trypanosomatids, an example of a rare genomic character. The two subgenic modules are transcribed independently and the resulting mRNAs appear to be independently translated, with the two protein products imported into mitochondria, based on the presence of predicted mitochondrial targeting peptides. Although the inferred protein sequences are in general very divergent from those of other organisms, all of the required iron-sulfur cluster-coordinating residues are present. Moreover, the discontinuity in the euglenozoan SdhB sequence occurs between the two domains of a typical, covalently continuous SdhB, consistent with the inference that the euglenozoan 'half' proteins are functional. Conclusion The discovery of this unique molecular marker provides evidence for the monophyly of Euglenozoa that is independent of evolutionary models. Our results pose questions about the origin and timing of this novel gene arrangement and the structure and function of euglenozoan SdhB.

  7. Natural history of succinic semialdehyde dehydrogenase deficiency through adulthood

    NARCIS (Netherlands)

    Lapalme-Remis, S.; Lewis, E.C.; De Meulemeester, C.; Chakraborty, P.; Gibson, K.M.; Torres, C.; Guberman, A.; Salomons, G.; Jakobs, C.; Ali-Ridha, A.; Parviz, M.; Pearl, P.L.

    2015-01-01

    Objective: The natural history of succinic semialdehyde dehydrogenase (SSADH) deficiency in adulthood is unknown; we elucidate the clinical manifestations of the disease later in life. Methods: A 63-year-old man with long-standing intellectual disability was diagnosed with SSADH deficiency following

  8. Identification of succinate dehydrogenase-deficient bladder paragangliomas.

    Science.gov (United States)

    Mason, Emily F; Sadow, Peter M; Wagner, Andrew J; Remillard, Stephen P; Flood, Trevor A; Belanger, Eric C; Hornick, Jason L; Barletta, Justine A

    2013-10-01

    A significant number of patients with paragangliomas harbor germline mutations in one of the succinate dehydrogenase (SDH) genes (SDHA, B, C, or D). Tumors with mutations in SDH genes can be identified using immunohistochemistry. Loss of SDHB staining is seen in tumors with a mutation in any one of the SDH genes, whereas loss of both SDHB and SDHA expression is seen only in the context of an SDHA mutation. Identifying an SDH-deficient tumor can be prognostically significant, as tumors with SDHB mutations are more likely to pursue a malignant course. Although the rate of SDH deficiency in paragangliomas in general is known to be approximately 30%, there are only rare reports of SDH-deficient bladder paragangliomas. Therefore, the aim of this study was to determine the rate of SDH deficiency in bladder paragangliomas. Eleven cases of bladder paragangliomas were identified. Hematoxylin and eosin-stained slides of all tumors were reviewed, and immunohistochemical analysis for SDHB and SDHA was performed. For cases with loss of SDHA expression by immunohistochemistry, mutation analysis of the SDHA gene was performed. Loss of SDHB staining was seen in 3 (27%) cases (2 with loss of SDHB only, 1 with loss of SDHB and SDHA). Patients with SDH-deficient tumors were younger than those with tumors with intact SDH expression (mean age at presentation 39 y and 58 y, respectively). Of the 2 patients with SDHB-deficient and SDHA-intact tumors, one was found to have a germline SDHB mutation, and the other had a family history of a malignant paraganglioma. Both patients developed metastatic disease. The one patient with a tumor that was deficient for both SDHB and SDHA had no family history of paragangliomas and no evidence of metastatic disease. Sequencing of this tumor revealed a deleterious heterozygous single-base pair substitution in exon 10 of SDHA (c.1340 A>G; p.His447Arg) in both the tumor and normal tissue, indicative of a germline SDHA mutation, and a deleterious single

  9. Circadian rhythm in succinate dehydrogenase activity in Neurospora crassa

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    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.

  10. Escherichia coli yjjPB genes encode a succinate transporter important for succinate production.

    Science.gov (United States)

    Fukui, Keita; Nanatani, Kei; Hara, Yoshihiko; Yamakami, Suguru; Yahagi, Daiki; Chinen, Akito; Tokura, Mitsunori; Abe, Keietsu

    2017-09-01

    Under anaerobic conditions, Escherichia coli produces succinate from glucose via the reductive tricarboxylic acid cycle. To date, however, no genes encoding succinate exporters have been established in E. coli. Therefore, we attempted to identify genes encoding succinate exporters by screening an E. coli MG1655 genome library. We identified the yjjPB genes as candidates encoding a succinate transporter, which enhanced succinate production in Pantoea ananatis under aerobic conditions. A complementation assay conducted in Corynebacterium glutamicum strain AJ110655ΔsucE1 demonstrated that both YjjP and YjjB are required for the restoration of succinate production. Furthermore, deletion of yjjPB decreased succinate production in E. coli by 70% under anaerobic conditions. Taken together, these results suggest that YjjPB constitutes a succinate transporter in E. coli and that the products of both genes are required for succinate export.

  11. Krebs cycle metabolite profiling for identification and stratification of pheochromocytomas/paragangliomas due to succinate dehydrogenase deficiency.

    Science.gov (United States)

    Richter, Susan; Peitzsch, Mirko; Rapizzi, Elena; Lenders, Jacques W; Qin, Nan; de Cubas, Aguirre A; Schiavi, Francesca; Rao, Jyotsna U; Beuschlein, Felix; Quinkler, Marcus; Timmers, Henri J; Opocher, Giuseppe; Mannelli, Massimo; Pacak, Karel; Robledo, Mercedes; Eisenhofer, Graeme

    2014-10-01

    Mutations of succinate dehydrogenase A/B/C/D genes (SDHx) increase susceptibility to development of pheochromocytomas and paragangliomas (PPGLs), with particularly high rates of malignancy associated with SDHB mutations. We assessed whether altered succinate dehydrogenase product-precursor relationships, manifested by differences in tumor ratios of succinate to fumarate or other metabolites, might aid in identifying and stratifying patients with SDHx mutations. PPGL tumor specimens from 233 patients, including 45 with SDHx mutations, were provided from eight tertiary referral centers for mass spectrometric analyses of Krebs cycle metabolites. Diagnostic performance of the succinate:fumarate ratio for identification of pathogenic SDHx mutations. SDH-deficient PPGLs were characterized by 25-fold higher succinate and 80% lower fumarate, cis-aconitate, and isocitrate tissue levels than PPGLs without SDHx mutations. Receiver-operating characteristic curves for use of ratios of succinate to fumarate or to cis-aconitate and isocitrate to identify SDHx mutations indicated areas under curves of 0.94 to 0.96; an optimal cut-off of 97.7 for the succinate:fumarate ratio provided a diagnostic sensitivity of 93% at a specificity of 97% to identify SDHX-mutated PPGLs. Succinate:fumarate ratios were higher in both SDHB-mutated and metastatic tumors than in those due to SDHD/C mutations or without metastases. Mass spectrometric-based measurements of ratios of succinate:fumarate and other metabolites in PPGLs offer a useful method to identify patients for testing of SDHx mutations, with additional utility to quantitatively assess functionality of mutations and metabolic factors responsible for malignant risk.

  12. Succinic semialdehyde dehydrogenase deficiency: lessons from mice and men.

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    Pearl, P L; Gibson, K M; Cortez, M A; Wu, Y; Carter Snead, O; Knerr, I; Forester, K; Pettiford, J M; Jakobs, C; Theodore, W H

    2009-06-01

    Succinic semialdehyde dehydrogenase (SSADH) deficiency, a disorder of GABA degradation with subsequent elevations in brain GABA and GHB, is a neurometabolic disorder with intellectual disability, epilepsy, hypotonia, ataxia, sleep disorders, and psychiatric disturbances. Neuroimaging reveals increased T2-weighted MRI signal usually affecting the globus pallidus, cerebellar dentate nucleus, and subthalamic nucleus, and often cerebral and cerebellar atrophy. EEG abnormalities are usually generalized spike-wave, consistent with a predilection for generalized epilepsy. The murine phenotype is characterized by failure-to-thrive, progressive ataxia, and a transition from generalized absence to tonic-clonic to ultimately fatal convulsive status epilepticus. Binding and electrophysiological studies demonstrate use-dependent downregulation of GABA(A) and (B) receptors in the mutant mouse. Translational human studies similarly reveal downregulation of GABAergic activity in patients, utilizing flumazenil-PET and transcranial magnetic stimulation for GABA(A) and (B) activity, respectively. Sleep studies reveal decreased stage REM with prolonged REM latencies and diminished percentage of stage REM. An ad libitum ketogenic diet was reported as effective in the mouse model, with unclear applicability to the human condition. Acute application of SGS-742, a GABA(B) antagonist, leads to improvement in epileptiform activity on electrocorticography. Promising mouse data using compounds available for clinical use, including taurine and SGS-742, form the framework for human trials.

  13. Review of succinate dehydrogenase-deficient renal cell carcinoma with focus on clinical and pathobiological aspects

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    Naoto Kuroda

    2016-05-01

    Full Text Available Succinate dehydrogenase (SDH-deficient renal cell carcinoma (RCC was first identified in 2004 and has been integrated into the 2016 WHO classification of RCC. Succinate dehydrogenase (SDH is an enzyme complex composed of four protein subunits (SDHA, SDHB, SDHC and SDHD. The tumor which presents this enzyme mutation accounts for 0.05 to 0.2% of all renal carcinomas. Multiple tumors may occur in approximately 30% of affected patients. SDHB-deficient RCC is the most frequent, and the tumor histologically consists of cuboidal cells with eosinophilic cytoplasm, vacuolization, flocculent intracytoplasmic inclusion and indistinct cell borders. Ultrastructurally, the tumor contains abundant mitochondria. Immunohistochemically, tumor cells are positive for SDHA, but negative for SDHB in SDHB-, SDHC- and SDHD-deficient RCCs. However, SDHA-deficient RCC shows negativity for both SDHA and SDHB. In molecular genetic analyses, a germline mutation in the SDHB , SDHC or SDHD gene (in keeping with most patients having germline mutations in an SDH gene has been identified in patients with or without a family history of renal tumors, paraganglioma/pheochromocytoma or gastrointestinal stromal tumor. While most tumors are low grade, some tumors may behave in an aggressive fashion, particularly if they are high nuclear grade, and have coagulative necrosis or sarcomatoid differentiation.

  14. The expression of succinate dehydrogenase in breast phyllodes tumor.

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    Choi, Junjeong; Kim, Do Hee; Jung, WooHee; Koo, Ja Seung

    2014-10-01

    The purpose of this study is to investigate the expression of succinate dehydrogenase (SDH)A, SDHB, and HIF-1α in phyllodes tumors and the association with clinic-pathologic factors. Using tissue microarray (TMA) for 206 phyllodes tumor cases, we performed immunohistochemical stains for SDHA, SDHB, and HIF-1α and analyzed their expression in regard to clinicopathologic parameters of each case. The cases were comprised of 156 benign, 34 borderline, and 16 malignant phyllodes tumors. The expression of stromal SDHA and epithelial- and stromal- SDHB increased as the tumor progressed from benign to malignant (P⟨0.001). There were five stromal SDHA-negative cases and 31 stromal SDHB-negative cases. SDHB negativity was associated with a lower histologic grade (P=0.054) and lower stromal atypia (P=0.048). Univariate analysis revealed that a shorter disease free survival (DFS) was associated with stromal SDHB high-positivity (P=0.013) and a shorter overall survival (OS) was associated with high-positivity of stromal SDHA and SDHB (P⟨0.001 and P⟨0.001, respectively). The multivariate Cox analysis with the variables stromal cellularity, stromal atypia, stromal mitosis, stromal overgrowth, tumor margin, stromal SDHA expression, and stromal SDHB expression revealed that stromal overgrowth was associated with a shorter DFS (hazard ratio: 24.78, 95% CI: 3.126-196.5, P=0.002) and a shorter OS (hazard ratio: 176.7, 95% CI: 8.466-3691, P=0.001). In conclusion, Tumor grade is positively correlated with SDHA and SDHB expression in the tumor stroma in phyllodes tumors of the breast. This result may be attributed to the increased metabolic demand in high grade tumors.

  15. Effect of gene disruptions of the TCA cycle on production of succinic acid in Saccharomyces cerevisiae.

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    Arikawa, Y; Kuroyanagi, T; Shimosaka, M; Muratsubaki, H; Enomoto, K; Kodaira, R; Okazaki, M

    1999-01-01

    Succinate is the main taste component produced by yeasts during sake (Japanese rice wine) fermentation. The pathway leading to accumulation of succinate was examined in liquid culture in the presence of a high concentration (15%) of glucose under aerobic and anaerobic conditions using a series of Saccharomyces cerevisiae strains in which various genes that encode the expression of enzymes required in TCA cycle were disrupted. When cultured in YPD medium containing 15% glucose under aerobic conditions, the KGD1 (alpha-ketoglutarate dehydrogenase) gene disrupted mutant produced a lower level of succinate than the wild-type strain, while the SDH1 (succinate dehydrogenase) gene-disrupted mutant produced an increased level of succinate. On the other hand, the FUM1 (fumarase) gene disrupted mutant produced significantly higher levels of fumarate but did not form malate at all. These results indicate that succinate, fumarate and malate are mainly synthesized through the TCA cycle (oxidative direction) even in the presence of glucose at a concentration as high as 15%. When the growth condition was shifted from aerobic to anaerobic, the increased level of succinate in SDH1 disruptants was no longer observed, whereas the decreased level of succinate in the KGD1 diruptant was still observed. A double mutant of the two fumarate reductase isozyme genes (OSM1 and FRDS) showed a succinate productivity of 50% as compared to the parent when cells were incubated in glucose-buffered solution. These results indicate that succinate could be synthesized through two pathways, namely, alpha-ketoglutarate oxidation via the TCA cycle and fumarate reduction under anaerobic conditions.

  16. Emergence of succinate dehydrogenase inhibitor resistance of Pyrenophora teres in Europe.

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    Rehfus, Alexandra; Miessner, Simone; Achenbach, Janosch; Strobel, Dieter; Bryson, Rosie; Stammler, Gerd

    2016-10-01

    Net blotch caused by Pyrenophora teres is an important disease of barley worldwide. In addition to strobilurins (quinone ouside inhibitors) and azoles (demethylation inhibitors), succinate dehydrogenase inhibitors (SDHIs) are very effective fungicides for net blotch control. Recently, SDHI-resistant isolates have been found in the field. Intensive sensitivity monitoring programmes across Europe were carried out to investigate the situation concerning SDHI resistance in P. teres. The first isolates with a lower sensitivity to SDHIs registered in barley were found in Germany in 2012 and carried the B-H277Y substitution in the succinate dehydrogenase enzyme. In 2013 and 2014, a significant increase in isolates with lower SDHI sensitivity was detected mainly in France and Germany, and the range of target-site mutations increased. Most of the resistant isolates carried the C-G79R substitution, which exhibits a strong impact on all SDHIs in microtitre tests. All SDHIs tested were shown to be cross-resistant. Other substitutions are gaining in importance, e.g. C-N75S in France and D-D145G in Germany. So far, no double mutants in SDH genes have been detected. Glasshouse tests showed that SDHI-resistant isolates were still controlled by the SDHI fluxapyroxad when applied preventively. To date, most isolates with C-G79R substitution have not simultaneously carried the F129L change in cytochrome b, which causes resistance towards QoI fungicides at low to moderate levels. Several target-site mutations in the genes of subunits SDH-B, SDH-C and SDH-D with different impact on SDHI fungicides were detected. The pattern of mutations varied from year to year and between different regions. Strict resistance management strategies are recommended to maintain SDHIs as effective tools for net blotch control, especially in areas with low frequencies of resistant isolates. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  17. Differences between the succinate dehydrogenase sequences of isopyrazam sensitive Zymoseptoria tritici and insensitive Fusarium graminearum strains.

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    Dubos, Tiphaine; Pasquali, Matias; Pogoda, Friederike; Casanova, Angèle; Hoffmann, Lucien; Beyer, Marco

    2013-01-01

    Forty-one Zymoseptoria tritici strains isolated in Luxembourg between 2009 and 2010 were highly sensitive towards the new succinate dehydrogenase inhibitor (SDHI) isopyrazam, with concentrations inhibiting fungal growth by 50% (EC50) ranging from 0.0281 to 4.53μM, whereas 41 Fusarium graminearum strains isolated in Europe and Northern America between 1969 and 2009 were insensitive with the average rate of inhibition converging towards 28% with increasing isopyrazam concentration. Seven isolates of both species covering the range of isopyrazam sensitivities observed in the present study were selected for the sequencing of the subunits B, C and D of the succinate dehydrogenase (sdh) gene. Predicted sdh amino acid sequences of subunits B, C and D were identical among F. graminearum strains. By comparing with fungal strains where resistance towards SDHIs was previously reported, three variations were unique to F. graminearum; B-D130N located in the iron-sulfur cluster [2Fe-2S], B-A275T located in the [3Fe-4S] cluster and an additional S at amino acid position 83-84 of sdhC, probably modifying structurally the ubiquinone binding site and therefore the biological activity of the fungicide. No variation was found among the Z. tritici strains in subunits B and D. Two variations were observed within the subunit C sequences of Z. tritici strains: C-N33T and C-N34T. The difference in EC50 values between Z. tritici strains with the NN and TT configuration was non-significant at P=0.289. Two outliers in the Z. tritici group with significantly higher EC50 values that were not related to mutations in the sdhB, sdhC, or sdhD were detected. The role of isopyrazam for the control of F. graminearum and Z. tritici in Luxembourg is discussed. Copyright © 2012 Elsevier Inc. All rights reserved.

  18. 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

  19. Crystallization and preliminary X-ray crystallographic studies of succinic semialdehyde dehydrogenase from Streptococcus pyogenes

    International Nuclear Information System (INIS)

    Jang, Eun Hyuk; Lim, Jong Eun; Chi, Young Min; Lee, Ki Seog

    2012-01-01

    Succinic semialdehyde dehydrogenase (SSADH) from S. pyogenes was purified and crystallized. Crystals of native and NAD + -complexed SSADH diffracted to resolutions of 1.6 and 1.7 Å, respectively. Succinic semialdehyde dehydrogenase (SSADH) plays a critical role in the metabolism of the inhibitory neurotransmitter γ-aminobutyric acid (GABA) and catalyzes the NAD(P) + -coupled oxidation of succinic semialdehyde (SSA) to succinic acid (SA). SSADH from Streptococcus pyogenes has been purified and crystallized as the apoenzyme and in a complex with NAD + . The crystals of native and NAD + -complexed SSADH diffracted to resolutions of 1.6 and 1.7 Å, respectively, using a synchrotron-radiation source. Both crystals belonged to the orthorhombic space group P2 1 2 1 2 1 , with unit-cell parameters a = 93.3, b = 100.3, c = 105.1 Å for the native crystal and a = 93.3, b = 100.3, c = 105.0 Å for the complex crystal. Preliminary molecular replacement confirmed the presence of one dimer in both crystals, corresponding to a Matthews coefficient (V M ) of 2.37 Å 3 Da −1 and a solvent content of 48.0%

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

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    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.

  2. Evidence of redox imbalance in a patient with succinic semialdehyde dehydrogenase deficiency

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    Anna-Kaisa Niemi

    2014-01-01

    Full Text Available The pathophysiology of succinic semialdehyde dehydrogenase (SSADH deficiency is not completely understood. Oxidative stress, mitochondrial pathology, and low reduced glutathione levels have been demonstrated in mice, but no studies have been reported in humans. We report on a patient with SSADH deficiency in whom we found low levels of blood reduced glutathione (GSH, and elevations of dicarboxylic acids in urine, suggestive of possible redox imbalance and/or mitochondrial dysfunction. Thus, targeting the oxidative stress axis may be a potential therapeutic approach if our findings are confirmed in other patients.

  3. In vitro kinetics of the rat brain succinate dehydrogenase inhibition by hexachlorophene.

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    Lokanatha, V; Sailaja, P; Rajendra, W

    1999-01-01

    Brain succinate dehydrogenase (SDH) activity was inhibited by in vitro hexachlorophene (HCP) with a half inhibitory concentration (IC50) of 0.65 x 10(-3) M. The HCP exerted noncompetitive inhibition at 0.5 mM (IC50) on SDH activity. The brain SDH demands more energy of activation (deltaE) in the presence of HCP. The ionizable groups of SDH such as the sulfhydral group of cysteine and alpha-amino groups of cysteine were not altered qualitatively in the presence of HCP.

  4. 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.)

  5. 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

  6. Molecular characterisation and detection of resistance to succinate dehydrogenase inhibitor fungicides in Botryotinia fuckeliana (Botrytis cinerea).

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    De Miccolis Angelini, Rita M; Masiello, Mario; Rotolo, Caterina; Pollastro, Stefania; Faretra, Francesco

    2014-12-01

    Succinate dehydrogenase inhibitors (SDHIs), interfering with fungal respiration, are considered to be fungicides at medium to high risk of resistance. Boscalid was the first molecule belonging to the SDHIs that was introduced for the control of Botryotinia fuckeliana. A range of different target-site mutations leading to boscalid resistance have been found in field populations of the fungus. The different types of mutation confer different cross-resistance profiles towards novel SDHIs, such as the recently introduced fungicide fluopyram. This study combines the determination of cross-resistance profiles and the setting-up of methods for fast molecular detection of the mutations. By means of in vitro tests, a range of SdhB mutations were characterised for resistance levels towards boscalid and fluopyram. SdhB mutations conferring P225L and P225F substitutions conferred high resistance to boscalid and high or moderate resistance to fluopyram respectively. Mutants carrying the N230I replacement were moderately resistant to both SDHIs. Substitutions at position H272 responsible for a high level of resistance to boscalid conferred sensitivity (H272R), hypersensitivity (H272Y) or moderate resistance (H272V) to fluopyram. Allele-specific (AS) PCR was developed and used for genotyping 135 B. fuckeliana isolates. The assay confirmed the strict association between resistance profiles and allelic variants of the SdhB gene. Real-time AS-PCR proved to be sensitive and specific for quantitative detection of different SDHI-resistant genotypes. Fluopyram-resistant mutants are currently rarely detected in the field sprayed with boscalid, but this may change with intensive exposure of the fungal population to fluopyram. PCR assays/methods developed in the study provide tools for fast monitoring of field populations and observing possible changes in population composition following fluopyram introduction, useful for the setting-up of appropriate preventive measures. © 2014 Society

  7. Mutants of GABA transaminase (POP2 suppress the severe phenotype of succinic semialdehyde dehydrogenase (ssadh mutants in Arabidopsis.

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    Frank Ludewig

    Full Text Available BACKGROUND: The gamma-aminubutyrate (GABA shunt bypasses two steps of the tricarboxylic acid cycle, and is present in both prokaryotes and eukaryotes. In plants, the pathway is composed of the calcium/calmodulin-regulated cytosolic enzyme glutamate decarboxylase (GAD, the mitochondrial enzymes GABA transaminase (GABA-T; POP2 and succinic semialdehyde dehydrogenase (SSADH. We have previously shown that compromising the function of the GABA-shunt, by disrupting the SSADH gene of Arabidopsis, causes enhanced accumulation of reactive oxygen intermediates (ROIs and cell death in response to light and heat stress. However, to date, genetic investigations of the relationships between enzymes of the GABA shunt have not been reported. PRINCIPAL FINDINGS: To elucidate the role of succinic semialdehyde (SSA, gamma-hydroxybutyrate (GHB and GABA in the accumulation of ROIs, we combined two genetic approaches to suppress the severe phenotype of ssadh mutants. Analysis of double pop2 ssadh mutants revealed that pop2 is epistatic to ssadh. Moreover, we isolated EMS-generated mutants suppressing the phenotype of ssadh revealing two new pop2 alleles. By measuring thermoluminescence at high temperature, the peroxide contents of ssadh and pop2 mutants were evaluated, showing that only ssadh plants accumulate peroxides. In addition, pop2 ssadh seedlings are more sensitive to exogenous SSA or GHB relative to wild type, because GHB and/or SSA accumulate in these plants. SIGNIFICANCE: We conclude that the lack of supply of succinate and NADH to the TCA cycle is not responsible for the oxidative stress and growth retardations of ssadh mutants. Rather, we suggest that the accumulation of SSA, GHB, or both, produced downstream of the GABA-T transamination step, is toxic to the plants, resulting in high ROI levels and impaired development.

  8. 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.

  9. Succinic semialdehyde dehydrogenase deficiency: Decrease in 4-OH-butyric acid levels with low doses of vigabatrin

    NARCIS (Netherlands)

    Escalera, G.I.; Ferrer, I.; Marina, L.C.; Sala, P.R.; Salomons, G.S.; Jakobs, C.; Perez-Cerda, C.

    2010-01-01

    Succinic semialdehyde dehydrogenase deficiency (gamma-hydroxybutyric aciduria) is a rare neurometabolic disease caused by a deficiency in gamma-aminobutyric degradation, resulting in an increase in gamma-hydroxybutyric acid in biological fluids. The clinical spectrum is heterogeneous, including a

  10. Discovering novel Alternaria solani succinate dehydrogenase inhibitors by in silico modeling and virtual screening strategies to combat early blight

    NARCIS (Netherlands)

    Iftikhar, Sehrish; Shahid, Ahmad A.; Halim, Sobia A.; Wolters, Pieter J.; Vleeshouwers, Vivianne G.A.A.; Khan, Ajmal; Al-Harrasi, Ahmed; Ahmad, Shahbaz

    2017-01-01

    Alternaria blight is an important foliage disease caused by Alternaria solani. The enzyme Succinate dehydrogenase (SDH) is a potential drug target because of its role in tricarboxylic acid cycle. Hence targeting Alternaria solani SDH enzyme could be efficient tool to design novel fungicides against

  11. A new point mutation in the iron-sulfur subunit of succinate dehydrogenase confers resistance to boscalid in Sclerotinia sclerotiorum.

    Science.gov (United States)

    Wang, Yong; Duan, Yabing; Wang, Jianxin; Zhou, Mingguo

    2015-09-01

    Research has established that mutations in highly conserved amino acids of the succinate dehydrogenase (SDH) complex in various fungi confer SDH inhibitor (SDHI) resistance. For Sclerotinia sclerotiorum (Lib.) de Bary, a necrotrophic fungus with a broad host range and a worldwide distribution, boscalid resistance has been attributed to the mutation H132R in the highly conserved SdhD subunit protein of the SDH complex. In our previous study, however, only one point mutation, A11V in SdhB (GCA to GTA change in SdhB), was detected in S. sclerotiorum boscalid-resistant (BR) mutants. In the current study, replacement of the SdhB gene in a boscalid-sensitive (BS) S. sclerotiorum strain with the mutant SdhB gene conferred resistance. Compared with wild-type strains, BR and GSM (SdhB gene in the wild-type strain replaced by the mutant SdhB gene) mutants were more sensitive to osmotic stress, lacked the ability to produce sclerotia and exhibited lower expression of the pac1 gene. Importantly, the point mutation was not located in the highly conserved sequence of the iron-sulfur subunit of SDH. These results suggest that resistance based on non-conserved vs. conserved protein domains differs in mechanism. In addition to increasing our understanding of boscalid resistance in S. sclerotiorum, the new information will be useful for the development of alternative antifungal drugs. © 2014 BSPP AND JOHN WILEY & SONS LTD.

  12. 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

  13. Immunolocalization of succinate dehydrogenase in the esophagus epithelium of domesticated mammals

    Directory of Open Access Journals (Sweden)

    W. Meyer

    2013-05-01

    Full Text Available Using immunohistochemistry and transmission electron microscopy (TEM, the esophagus epithelia of seven domesticated mammals (horse, cattle, goat, pig, dog, laboratory rat, cat of three nutrition groups (herbivorous, omnivorous, carnivorous were studied to get first information about energy generation, as demonstrated by succinate dehydrogenase (SDH activities. Distinct reaction intensities could be observed in all esophageal cell layers of the different species studied reflecting moderate to strong metabolic activities. The generally strong staining in the stratum basale indicated that new cells are continuously produced. The latter feature was confirmed by a thick, and in the horse generally highly active stratum spinosum. Only in the pig, reaction intensity variations occurred, obviously related to differences in physical feed quality or restricted feed allocation. The immunohistochemical results were corroborated by the presence of intact mitochondria in the esophageal cells of all species and nutrition types studied, except for the horse. Possible relationships between SDH reaction intensities and feed structure, mass or consistency are discussed.

  14. 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.

  15. 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.

  16. Fumarate and Succinate Regulate Expression of Hypoxia-inducible Genes via TET Enzymes.

    Science.gov (United States)

    Laukka, Tuomas; Mariani, Christopher J; Ihantola, Tuukka; Cao, John Z; Hokkanen, Juho; Kaelin, William G; Godley, Lucy A; Koivunen, Peppi

    2016-02-19

    The TET enzymes are members of the 2-oxoglutarate-dependent dioxygenase family and comprise three isoenzymes in humans: TETs 1-3. These TETs convert 5-methylcytosine to 5-hydroxymethylcytosine (5-hmC) in DNA, and high 5-hmC levels are associated with active transcription. The importance of the balance in these modified cytosines is emphasized by the fact that TET2 is mutated in several human cancers, including myeloid malignancies such as acute myeloid leukemia (AML). We characterize here the kinetic and inhibitory properties of Tets and show that the Km value of Tets 1 and 2 for O2 is 30 μm, indicating that they retain high activity even under hypoxic conditions. The AML-associated mutations in the Fe(2+) and 2-oxoglutarate-binding residues increased the Km values for these factors 30-80-fold and reduced the Vmax values. Fumarate and succinate, which can accumulate to millimolar levels in succinate dehydrogenase and fumarate hydratase-mutant tumors, were identified as potent Tet inhibitors in vitro, with IC50 values ∼400-500 μm. Fumarate and succinate also down-regulated global 5-hmC levels in neuroblastoma cells and the expression levels of some hypoxia-inducible factor (HIF) target genes via TET inhibition, despite simultaneous HIFα stabilization. The combination of fumarate or succinate treatment with TET1 or TET3 silencing caused differential effects on the expression of specific HIF target genes. Altogether these data show that hypoxia-inducible genes are regulated in a multilayered manner that includes epigenetic regulation via TETs and 5-hmC levels in addition to HIF stabilization. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. 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.

  18. Mutagenesis and functional studies with succinate dehydrogenase inhibitors in the wheat pathogen Mycosphaerella graminicola.

    Directory of Open Access Journals (Sweden)

    Gabriel Scalliet

    Full Text Available A range of novel carboxamide fungicides, inhibitors of the succinate dehydrogenase enzyme (SDH, EC 1.3.5.1 is currently being introduced to the crop protection market. The aim of this study was to explore the impact of structurally distinct carboxamides on target site resistance development and to assess possible impact on fitness. We used a UV mutagenesis approach in Mycosphaerella graminicola, a key pathogen of wheat to compare the nature, frequencies and impact of target mutations towards five subclasses of carboxamides. From this screen we identified 27 amino acid substitutions occurring at 18 different positions on the 3 subunits constituting the ubiquinone binding (Qp site of the enzyme. The nature of substitutions and cross resistance profiles indicated significant differences in the binding interaction to the enzyme across the different inhibitors. Pharmacophore elucidation followed by docking studies in a tridimensional SDH model allowed us to propose rational hypotheses explaining some of the differential behaviors for the first time. Interestingly all the characterized substitutions had a negative impact on enzyme efficiency, however very low levels of enzyme activity appeared to be sufficient for cell survival. In order to explore the impact of mutations on pathogen fitness in vivo and in planta, homologous recombinants were generated for a selection of mutation types. In vivo, in contrast to previous studies performed in yeast and other organisms, SDH mutations did not result in a major increase of reactive oxygen species levels and did not display any significant fitness penalty. However, a number of Qp site mutations affecting enzyme efficiency were shown to have a biological impact in planta.Using the combined approaches described here, we have significantly improved our understanding of possible resistance mechanisms to carboxamides and performed preliminary fitness penalty assessment in an economically important plant pathogen

  19. S-nitrosylation of the mitochondrial chaperone TRAP1 sensitizes hepatocellular carcinoma cells to inhibitors of succinate dehydrogenase

    DEFF Research Database (Denmark)

    Rizza, Salvatore; Montagna, Costanza; Cardaci, Simone

    2016-01-01

    degradation. As a result, GSNOR-deficient cells and tumors are highly sensitive to SDH inhibition, namely to α-tocopheryl succinate, an SDH-targeting molecule that induced RIP1/PARP1-mediated necroptosis and inhibited tumor growth. Our work provides a specific molecular signature of aberrant S....... We find that hepatocyte GSNOR deficiency is characterized by mitochondrial alteration and by marked increases in succinate dehydrogenase (SDH) levels and activity. We find that this depends on the selective S-nitrosylation of Cys(501) in the mitochondrial chaperone TRAP1, which mediates its......-nitrosylation in HCC, a novel molecular target in SDH, and a first-in-class therapy to treat the disease. Cancer Res; 76(14); 1-13. ©2016 AACR....

  20. Gamma-Hydroxybutyrate (GHB) Content in Hair Samples Correlates Negatively with Age in Succinic Semialdehyde Dehydrogenase Deficiency

    DEFF Research Database (Denmark)

    Johansen, S S; Wang, X.; Pedersen, Daniel Sejer

    2017-01-01

    Gamma-hydroxybutyrate (GHB) is a drug of abuse, an approved therapeutic for narcolepsy, an agent employed for facilitation of sexual assault, as well as a biomarker of succinic semialdehyde dehydrogenase deficiency (SSADHD). Our laboratory seeks to identify surrogate biomarkers in SSADHD that can...... shed light on the developmental course of this neurometabolic disease. Since GHB may be quantified in hair as a potential surrogate to identify victims of drug-related assault, we have opted to examine its level in SSADHD. We quantified GHB in hair derived from ten patients with SSADHD, and documented...

  1. 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.

  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. Inhibition of mitochondrial glycerol-3-phosphate dehydrogenase by alpha-tocopheryl succinate

    Czech Academy of Sciences Publication Activity Database

    Rauchová, Hana; Vokurková, Martina; Drahota, Zdeněk

    2014-01-01

    Roč. 53, AUG (2014), s. 409-413 ISSN 1357-2725 R&D Projects: GA ČR(CZ) GAP304/12/0259 Institutional support: RVO:67985823 Keywords : brown adipose tissue mitochondria * oxygen consumption * glycerol-3-phosphate * succinate * reactive oxygen species Subject RIV: ED - Physiology Impact factor: 4.046, year: 2014

  4. 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.

  5. Discovering Novel Alternaria solani Succinate Dehydrogenase Inhibitors by in Silico Modeling and Virtual Screening Strategies to Combat Early Blight

    Directory of Open Access Journals (Sweden)

    Sehrish Iftikhar

    2017-11-01

    Full Text Available Alternaria blight is an important foliage disease caused by Alternaria solani. The enzyme Succinate dehydrogenase (SDH is a potential drug target because of its role in tricarboxylic acid cycle. Hence targeting Alternaria solani SDH enzyme could be efficient tool to design novel fungicides against A. solani. We employed computational methodologies to design new SDH inhibitors using homology modeling; pharmacophore modeling and structure based virtual screening. The three dimensional SDH model showed good stereo-chemical and structural properties. Based on virtual screening results twelve commercially available compounds were purchased and tested in vitro and in vivo. The compounds were found to inhibit mycelial growth of A. solani. Moreover in vitro trials showed that inhibitory effects were enhanced with increase in concentrations. Similarly increased disease control was observed in pre-treated potato tubers. Hence the applied in silico strategy led us to identify novel fungicides.

  6. 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.

  7. The SDH mutation database: an online resource for succinate dehydrogenase sequence variants involved in pheochromocytoma, paraganglioma and mitochondrial complex II deficiency

    Directory of Open Access Journals (Sweden)

    Devilee Peter

    2005-11-01

    Full Text Available Abstract Background The SDHA, SDHB, SDHC and SDHD genes encode the subunits of succinate dehydrogenase (succinate: ubiquinone oxidoreductase, a component of both the Krebs cycle and the mitochondrial respiratory chain. SDHA, a flavoprotein and SDHB, an iron-sulfur protein together constitute the catalytic domain, while SDHC and SDHD encode membrane anchors that allow the complex to participate in the respiratory chain as complex II. Germline mutations of SDHD and SDHB are a major cause of the hereditary forms of the tumors paraganglioma and pheochromocytoma. The largest subunit, SDHA, is mutated in patients with Leigh syndrome and late-onset optic atrophy, but has not as yet been identified as a factor in hereditary cancer. Description The SDH mutation database is based on the recently described Leiden Open (source Variation Database (LOVD system. The variants currently described in the database were extracted from the published literature and in some cases annotated to conform to current mutation nomenclature. Researchers can also directly submit new sequence variants online. Since the identification of SDHD, SDHC, and SDHB as classic tumor suppressor genes in 2000 and 2001, studies from research groups around the world have identified a total of 120 variants. Here we introduce all reported paraganglioma and pheochromocytoma related sequence variations in these genes, in addition to all reported mutations of SDHA. The database is now accessible online. Conclusion The SDH mutation database offers a valuable tool and resource for clinicians involved in the treatment of patients with paraganglioma-pheochromocytoma, clinical geneticists needing an overview of current knowledge, and geneticists and other researchers needing a solid foundation for further exploration of both these tumor syndromes and SDHA-related phenotypes.

  8. 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.

  9. 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.

  10. 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.

  11. Mutations in Succinate Dehydrogenase Subunit C Increase Radiosensitivity and Bystander Responses

    Science.gov (United States)

    Zhou, Hongning; Hei, Tom K.

    Although radiation-induced bystander effect is well studied in the past decade, the precise mech-anisms are still unclear. It is likely that a combination of pathways involving both primary and secondary signaling processes is involved in producing a bystander effect. There is recent evidence that mitochondria play a critical role in bystander responses. Recently studies found that a mutation in succinate dehydrogenese subunit C (SDHC), an integral membrane protein in complex II of the electron transport chain, resulted in increased superoxide, oxidative stress, apoptosis, tumorigenesis, and genomic instability, indicating that SDHC play a critical role in maintaining mitochondrial function. In the present study, using Chinese hamster fibroblasts (B1 cells) and the mutants (B9 cells) containing a single base substitution that produced a premature stop codon resulting in a 33-amino acid COOH-terminal truncation of the SDHC protein, we found that B9 cells had an increase in intracellular superoxide content, nitric oxide species, and mitochondrial membrane potential when compared with wild type cells. After irradiated with a grade of doses of gamma rays, B9 cells show an increased radiosensitivity, especially at high doses. The HPRT- mutant yield after gamma-ray irradiation in B9 cells was significantly higher than that of B1 cells. A single, 3Gy dose of gamma-rays increased the background mutant level by more than 4 fold. In contrast, the mutant induction was less than 2 fold in B1 cells. In addition, B9 cells produced a higher bystander mutagenesis after alpha particle irradiation than the B1 cells. Furthermore, pretreated with carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO), a nitric oxide scavenger, significantly decreased the bystander effect. Our findings demonstrate that a mutation in SDHC increases radiosensitivity in both directly irradiated cells and in neighboring bystander cells, and mito-chondrial function play an essential role in

  12. Visualization of mitochondrial respiratory function using cytochrome c oxidase/succinate dehydrogenase (COX/SDH) double-labeling histochemistry.

    Science.gov (United States)

    Ross, Jaime M

    2011-11-23

    Mitochondrial DNA (mtDNA) defects are an important cause of disease and may underlie aging and aging-related alterations (1,2). The mitochondrial theory of aging suggests a role for mtDNA mutations, which can alter bioenergetics homeostasis and cellular function, in the aging process (3). A wealth of evidence has been compiled in support of this theory (1,4), an example being the mtDNA mutator mouse (5); however, the precise role of mtDNA damage in aging is not entirely understood (6,7). Observing the activity of respiratory enzymes is a straightforward approach for investigating mitochondrial dysfunction. Complex IV, or cytochrome c oxidase (COX), is essential for mitochondrial function. The catalytic subunits of COX are encoded by mtDNA and are essential for assembly of the complex (Figure 1). Thus, proper synthesis and function are largely based on mtDNA integrity (2). Although other respiratory complexes could be investigated, Complexes IV and II are the most amenable to histochemical examination (8,9). Complex II, or succinate dehydrogenase (SDH), is entirely encoded by nuclear DNA (Figure 1), and its activity is typically not affected by impaired mtDNA, although an increase might indicate mitochondrial biogenesis (10-12). The impaired mtDNA observed in mitochondrial diseases, aging, and age-related diseases often leads to the presence of cells with low or absent COX activity (2,12-14). Although COX and SDH activities can be investigated individually, the sequential double-labeling method (15,16) has proved to be advantageous in locating cells with mitochondrial dysfunction (12,17-21). Many of the optimal constitutions of the assay have been determined, such as substrate concentration, electron acceptors/donors, intermediate electron carriers, influence of pH, and reaction time (9,22,23). 3,3'-diaminobenzidine (DAB) is an effective and reliable electron donor (22). In cells with functioning COX, the brown indamine polymer product will localize in mitochondrial

  13. Hyperthermo-chemo-radiotherapy for patients with carcinoma of the esophagus and prediction of the clinical remedial value using the in vitro succinate dehydrogenase inhibition test

    International Nuclear Information System (INIS)

    Matsuda, Hiroyuki; Sugimachi, Keizo; Ohno, Shinji; Mori, Masaki; Kuwano, Hiroyuki; Maehara, Yoshihiko

    1989-01-01

    Hyperthermia combined with irradiation and chemotherapy was prescribed for patients with resectable (n=88) and unresectable (n=36) carcinoma of the esophagus. The histopathological effectiveness, and the long term results were compared between two groups of patients treated with hyperthermo-chemo-radiotherapy (HCR therapy) and those treated with chemoradiotherapy (CR therapy). A correlation between the hyperthermia sensitivity test, using the in vitro succinate dehydrogenase inhibition test, and the clinical remedial value was also examined in malignant tissues from 47 patients with esophageal cancer, and treated with HCR therapy. In the resected cases, preoperative HCR therapy resulted in a significantly higher histopathological effectiveness rate (69%) compared with that in cases treated by CR therapy (48%) (P<0.05). The long term results of patients with both resectable and unresectable carcinoma, given HCR therapy were significanly better than those given CR therapy (P<0.05). On the other hand, the clinical remedial value, determined radiographically showed a correlation rate of 77% to the hyperthermia sensitivity test. Thus, the HCR therapy resulted in not only a higher histopathological effectiveness rate but also a significantly longer survival without severe side effects, and this hyperthermia sensitivity test using the succinate dehydrogenase inhibition test facilitates prediction of the outcome of the HCR therapy. (author)

  14. CRISPR interference (CRISPRi) for gene regulation and succinate production in cyanobacterium S. elongatus PCC 7942.

    Science.gov (United States)

    Huang, Chun-Hung; Shen, Claire R; Li, Hung; Sung, Li-Yu; Wu, Meng-Ying; Hu, Yu-Chen

    2016-11-15

    Cyanobacterium Synechococcus elongatus PCC 7942 holds promise for biochemical conversion, but gene deletion in PCC 7942 is time-consuming and may be lethal to cells. CRISPR interference (CRISPRi) is an emerging technology that exploits the catalytically inactive Cas9 (dCas9) and single guide RNA (sgRNA) to repress sequence-specific genes without the need of gene knockout, and is repurposed to rewire metabolic networks in various procaryotic cells. To employ CRISPRi for the manipulation of gene network in PCC 7942, we integrated the cassettes expressing enhanced yellow fluorescent protein (EYFP), dCas9 and sgRNA targeting different regions on eyfp into the PCC 7942 chromosome. Co-expression of dCas9 and sgRNA conferred effective and stable suppression of EYFP production at efficiencies exceeding 99%, without impairing cell growth. We next integrated the dCas9 and sgRNA targeting endogenous genes essential for glycogen accumulation (glgc) and succinate conversion to fumarate (sdhA and sdhB). Transcription levels of glgc, sdhA and sdhB were effectively suppressed with efficiencies depending on the sgRNA binding site. Targeted suppression of glgc reduced the expression to 6.2%, attenuated the glycogen accumulation to 4.8% and significantly enhanced the succinate titer. Targeting sdhA or sdhB also effectively downregulated the gene expression and enhanced the succinate titer ≈12.5-fold to ≈0.58-0.63 mg/L. These data demonstrated that CRISPRi-mediated gene suppression allowed for re-directing the cellular carbon flow, thus paving a new avenue to rationally fine-tune the metabolic pathways in PCC 7942 for the production of biotechnological products.

  15. Glucose-dependent turnover of the mRNAs encoding succinate dehydrogenase peptides in Saccharomyces cerevisiae: sequence elements in the 5' untranslated region of the Ip mRNA play a dominant role.

    OpenAIRE

    Cereghino, G P; Atencio, D P; Saghbini, M; Beiner, J; Scheffler, I E

    1995-01-01

    We have demonstrated previously that glucose repression of mitochondrial biogenesis in Saccharomyces cerevisiae involves the control of the turnover of mRNAs for the iron protein (Ip) and flavoprotein (Fp) subunits of succinate dehydrogenase (SDH). Their half-lives are > 60 min in the presence of a nonfermentable carbon source (YPG medium) and 60 min in the presence of a nonfermentable carbon source...

  16. Enhancement of malate-production and increase in sensitivity to dimethyl succinate by mutation of the VID24 gene in Saccharomyces cerevisiae.

    Science.gov (United States)

    Negoro, Hiroaki; Kotaka, Atsushi; Matsumura, Kengo; Tsutsumi, Hiroko; Hata, Yoji

    2016-06-01

    Malate in sake (a Japanese alcoholic beverage) is an important component for taste that is produced by yeasts during alcoholic fermentation. To date, many researchers have developed methods for breeding high-malate-producing yeasts; however, genes responsible for the high-acidity phenotype are not known. We determined the mutated gene involved in high malate production in yeast, isolated as a sensitive mutant to dimethyl succinate. In the comparative whole genome analysis between high-malate-producing strain and its parent strain, one of the non-synonymous substitutions was identified in the VID24 gene. The mutation of VID24 resulted in enhancement of malate-productivity and sensitivity to dimethyl succinate. The mutation appeared to lead to a deficiency in Vid24p function. Furthermore, disruption of cytoplasmic malate dehydrogenase (Mdh2p) gene in the VID24 mutant inhibited the high-malate-producing phenotype. Vid24p is known as a component of the multisubunit ubiquitin ligase and participates in the degradation of gluconeogenic enzymes such as Mdh2p. We suggest that the enhancement of malate-productivity results from an accumulation of Mdh2p due to the loss of Vid24p function. These findings propose a novel mechanism for the regulation of organic acid production in yeast cells by the component of ubiquitin ligase, Vid24p. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  17. Isolation and characterization of the rat gene encoding glutamate dehydrogenase

    NARCIS (Netherlands)

    Das, A. T.; Arnberg, A. C.; Malingré, H.; Moerer, P.; Charles, R.; Moorman, A. F.; Lamers, W. H.

    1993-01-01

    The concentration of glutamate dehydrogenase (GDH) varies strongly between different organs and between different regions within organs. To permit further studies on the regulation of GDH expression, we isolated and characterized the rat gene encoding the GDH protein. This gene contains 13 exons and

  18. Succinate dehydrogenase (SDHx) mutations in pituitary tumors: could this be a new role for mitochondrial complex II and/or Krebs cycle defects?

    Science.gov (United States)

    Xekouki, Paraskevi; Stratakis, Constantine A

    2012-12-01

    Succinate dehydrogenase (SDH) or mitochondrial complex II is a multimeric enzyme that is bound to the inner membrane of mitochondria and has a dual role as it serves both as a critical step of the tricarboxylic acid or Krebs cycle and as a member of the respiratory chain that transfers electrons directly to the ubiquinone pool. Mutations in SDH subunits have been implicated in the formation of familial paragangliomas (PGLs) and/or pheochromocytomas (PHEOs) and in Carney-Stratakis syndrome. More recently, SDH defects were associated with predisposition to a Cowden disease phenotype, renal, and thyroid cancer. We recently described a kindred with the coexistence of familial PGLs and an aggressive GH-secreting pituitary adenoma, harboring an SDHD mutation. The pituitary tumor showed loss of heterozygosity at the SDHD locus, indicating the possibility that SDHD's loss was causatively linked to the development of the neoplasm. In total, 29 cases of pituitary adenomas presenting in association with PHEOs and/or extra-adrenal PGLs have been reported in the literature since 1952. Although a number of other genetic defects are possible in these cases, we speculate that the association of PHEOs and/or PGLs with pituitary tumors is a new syndromic association and a novel phenotype for SDH defects.

  19. 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.

  20. 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.

  1. 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.

  2. Mutant E. coli strain with increased succinic acid production

    Science.gov (United States)

    Donnelly, Mark; Millard, Cynthia S.; Stols, Lucy

    1998-01-01

    A method for isolating succinic acid producing bacteria is provided comprising increasing the biomass of an organism which lacks the ability to catabolize pyruvate, and then subjecting the biomass to glucose-rich medium in an anaerobic environment to enable pyruvate-catabolizing mutants to grow. The invention also provides for a mutant that produces high amounts of succinic acid, which as been derived from a parent which lacked the genes for pyruvate formate lyase and lactate dehydrogenase, and which belongs to the E.coli Group of Bacteria.

  3. Mutant E. coli strain with increased succinic acid production

    Science.gov (United States)

    Donnelly, Mark; Millard, Cynthia S.; Stols, Lucy

    2001-09-25

    A method for isolating succinic acid producing bacteria is provided comprising increasing the biomass of an organism which lacks the ability to catabolize pyruvate, and then subjecting the biomass to glucose-rich medium in an anaerobic environment to enable pyruvate-catabolizing mutants to grow. The invention also provides for a mutant that produces high amounts of succinic acid, which has been derived from a parent which lacked the genes for pyruvate formate lyase and lactate dehydrogenase, and which belongs to the E.coli Group of Bacteria.

  4. Succinate Dehydrogenase Subunit B (SDHB Is Expressed in Neurofibromatosis 1-Associated Gastrointestinal Stromal Tumors (Gists: Implications for the SDHB Expression Based Classification of Gists

    Directory of Open Access Journals (Sweden)

    Jeanny H. Wang, Jerzy Lasota, Markku Miettinen

    2011-01-01

    Full Text Available Gastrointestinal Stromal Tumor (GIST is the most common mesenchymal tumor of the digestive tract. GISTs develop with relatively high incidence in patients with Neurofibromatosis-1 syndrome (NF1. Mutational activation of KIT or PDGFRA is believed to be a driving force in the pathogenesis of familial and sporadic GISTs. Unlike those tumors, NF1-associated GISTs do not have KIT or PGDFRA mutations. Similarly, no mutational activation of KIT or PDGFRA has been identified in pediatric GISTs and in GISTs associated with Carney Triad and Carney-Stratakis Syndrome. KIT and PDGFRA-wild type tumors are expected to have lesser response to imatinib treatment. Recently, Carney Triad and Carney-Stratakis Syndrome -associated GISTs and pediatric GISTs have been shown to have a loss of expression of succinate dehydrogenase subunit B (SDHB, a Krebs cycle/electron transport chain interface protein. It was proposed that GISTs can be divided into SDHB- positive (type 1, and SDHB-negative (type 2 tumors because of similarities in clinical features and response to imatinib treatment. In this study, SDHB expression was examined immunohistochemically in 22 well-characterized NF1-associated GISTs. All analyzed tumors expressed SDHB. Based on SDHB-expression status, NF1-associated GISTs belong to type 1 category; however, similarly to SDHB type 2 tumors, they do not respond well to imatinib treatment. Therefore, a simple categorization of GISTs into SDHB-positive and-negative seems to be incomplete. A classification based on both SDHB expression status and KIT and PDGFRA mutation status characterize GISTs more accurately and allow subdivision of SDHB-positive tumors into different clinico-genetic categories.

  5. [Control of acetic acid metabolism of recombinant Yarrowia lipolytica for efficient succinic acid production].

    Science.gov (United States)

    Gao, Cuijuan; Zheng, Yaqin

    2018-03-25

    Succinic acid is a high value-added organic acid widely used in food, chemical and pesticide industries. As a new robust non-conventional yeast, Yarrowia lipolytica attracts more and more attention due to its potential for industrial applications. Previously, we obtained a succinic acid-producing strain through gene deletion of succinic acid dehydrogenase subunit encoding gene Ylsdh5, resulting in the strain of PGC01003. However, the recombinant strain produced large amount of acetic acid due to imbalance between glycolysis and TCA cycle which hindered the efficient production of succinic acid. PDH bypass was interfered to decrease the overflow of acetic acid and produce succinic acid under natural pH. Acetic acid was reduced to 4.6 g/L through heterologous expression of acetyl coenzyme A synthase from Salmonella enteric, which was 75.4% of the control strain. Deletion of CoA-transferase gene Ylach1 eliminated acetate formation and improved succinic acid production, and the resulting strain produced as high as 7.0 g/L succinic acid. Our study provides foundation for further construction of efficient cell factory of succinic acid production.

  6. Cationic lipids bearing succinic-based, acyclic and macrocyclic hydrophobic domains: Synthetic studies and in vitro gene transfer.

    Science.gov (United States)

    Jubeli, Emile; Maginty, Amanda B; Khalique, Nada Abdul; Raju, Liji; Nicholson, David G; Larsen, Helge; Pungente, Michael D; Goldring, William P D

    2017-01-05

    In this communication we describe the construction of four succinic-based cationic lipids, their formulation with plasmid DNA (pDNA), and an evaluation of their in vitro gene delivery into Chinese hamster ovarian (CHO-K1) cells. The cationic lipids employed in this work possess either a dimethylamine or trimethylamine headgroup, and a macrocyclic or an acyclic hydrophobic domain composed of, or derived from two 16-atom, succinic-based acyl chains. The synthesized lipids and a co-lipid of neutral charge, either cholesterol or 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), were formulated in an overall 3:2 cationic-to-neutral lipid molar ratio, then complexed with plasmid DNA (pDNA). The relative transfection performance was evaluated via a comparison between matched versus mismatched formulations defined by the rigidity relationship between the lipids employed. Gel electrophoresis was used to characterize the binding of the lipid formulations with plasmid DNA and the relative degree of plasmid degradation using a DNase I degradation assay. Small angle X-ray diffraction (SAXD) was employed to characterize the packing morphology of the lipid-DNA complexes. In general, the succinic unit embedded within the hydrophobic domain of the cationic lipids was found to improve lipid hydration. The transfection assays revealed a general trend in which mismatched formulations that employed a rigid lipid combined with a non-rigid (or flexible) lipid, outperformed the matched formulations. The results from this work suggest that the design of the cationic lipid structure and the composition of the lipoplex formulation play key roles in governing the transfection performance of nonviral gene delivery agents. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  7. Glucose-dependent turnover of the mRNAs encoding succinate dehydrogenase peptides in Saccharomyces cerevisiae: sequence elements in the 5' untranslated region of the Ip mRNA play a dominant role.

    Science.gov (United States)

    Cereghino, G P; Atencio, D P; Saghbini, M; Beiner, J; Scheffler, I E

    1995-09-01

    We have demonstrated previously that glucose repression of mitochondrial biogenesis in Saccharomyces cerevisiae involves the control of the turnover of mRNAs for the iron protein (Ip) and flavoprotein (Fp) subunits of succinate dehydrogenase (SDH). Their half-lives are > 60 min in the presence of a nonfermentable carbon source (YPG medium) and YPD medium). This is a rare example in yeast in which the half-lives are > 60 min in the presence of a nonfermentable carbon source (YPG medium) and YPD medium). This is a rare example in yeast in which the half-life of an mRNA can be controlled by manipulating external conditions. In our current studies, a series of Ip transcripts with internal deletions as well as chimeric transcripts with heterologous sequences (internally or at the ends) have been examined, and we established that the 5'-untranslated region (5' UTR) of the Ip mRNA contains a major determinant controlling its differential turnover in YPG and YPD. Furthermore, the 5' exonuclease encoded by the XRN1 gene is required for the rapid degradation of the Ip and Fp mRNAs upon the addition of glucose. In the presence of cycloheximide the nucleolytic degradation of the Ip mRNA can be slowed down by stalled ribosomes to allow the identification of intermediates. Such intermediates have lost their 5' ends but still retain their 3' UTRs. If protein synthesis is inhibited at an early initiation step by the use of a prt1 mutation (affecting the initiation factor eIF3), the Ip and Fp mRNAs are very rapidly degraded even in YPG. Significantly, the arrest of translation by the introduction of a stable hairpin loop just upstream of the initiation codon does not alter the differential stability of the transcript in YPG and YPD. These observations suggest that a signaling pathway exists in which the external carbon source can control the turnover of mRNAs of specific mitochondrial proteins. Factors must be present that control either the activity or more likely the access of a

  8. Cloning and Polymorphisms of Yak Lactate Dehydrogenase b Gene

    Directory of Open Access Journals (Sweden)

    Yaou Xu

    2013-06-01

    Full Text Available The main objective of this work was to study the unique polymorphisms of the lactate dehydrogenase-1 (LDH1 gene in yak (Bos grunniens. Native polyacrylamide gel electrophoresis revealed three phenotypes of LDH1 (a tetramer of H subunit in yak heart and longissimus muscle extracts. The corresponding gene, ldhb, encoding H subunits of three LDH1 phenotypes was obtained by RT-PCR. A total of six nucleotide differences were detected in yak ldhb compared with that of cattle, of which five mutations cause amino acid substitutions. Sequence analysis shows that the G896A and C689A, mutations of ldhb gene, result in alterations of differently charged amino acids, and create the three phenotypes (F, M, and S of yak LDH1. Molecular modeling of the H subunit of LDH indicates that the substituted amino acids are not located within NAD+ or substrate binding sites. PCR-RFLP examination of G896A mutation demonstrated that most LDH1-F samples are actually heterozygote at this site. These results help to elucidate the molecular basis and genetic characteristic of the three unique LDH1 phenotypes in yak.

  9. Isolation and Expression of Lactate Dehydrogenase Genes from Rhizopus oryzae

    Science.gov (United States)

    Skory, Christopher D.

    2000-01-01

    Rhizopus oryzae is used for industrial production of lactic acid, yet little is known about the genetics of this fungus. In this study I cloned two genes, ldhA and ldhB, which code for NAD+-dependent l-lactate dehydrogenases (LDH) (EC 1.1.1.27), from a lactic acid-producing strain of R. oryzae. These genes are similar to each other and exhibit more than 90% nucleotide sequence identity and they contain no introns. This is the first description of ldh genes in a fungus, and sequence comparisons revealed that these genes are distinct from previously isolated prokaryotic and eukaryotic ldh genes. Protein sequencing of the LDH isolated from R. oryzae during lactic acid production confirmed that ldhA codes for a 36-kDa protein that converts pyruvate to lactate. Production of LdhA was greatest when glucose was the carbon source, followed by xylose and trehalose; all of these sugars could be fermented to lactic acid. Transcripts from ldhB were not detected when R. oryzae was grown on any of these sugars but were present when R. oryzae was grown on glycerol, ethanol, and lactate. I hypothesize that ldhB encodes a second NAD+-dependent LDH that is capable of converting l-lactate to pyruvate and is produced by cultures grown on these nonfermentable substrates. Both ldhA and ldhB restored fermentative growth to Escherichia coli (ldhA pfl) mutants so that they grew anaerobically and produced lactic acid. PMID:10831409

  10. Industrial Systems Biology of Saccharomyces cerevisiae Enables Novel Succinic Acid Cell Factory

    DEFF Research Database (Denmark)

    Otero, José Manuel; Cimini, Donatella; Patil, Kiran Raosaheb

    2013-01-01

    production. Glycine and serine, both essential amino acids required for biomass formation, are formed from both glycolytic and TCA cycle intermediates. Succinate formation results from the isocitrate lyase catalyzed conversion of isocitrate, and from the α-keto-glutarate dehydrogenase catalyzed conversion...... of α-keto-glutarate. Succinate is subsequently depleted by the succinate dehydrogenase complex. The metabolic engineering strategy identified included deletion of the primary succinate consuming reaction, Sdh3p, and interruption of glycolysis derived serine by deletion of 3-phosphoglycerate...

  11. Evolution of glutamate dehydrogenase genes: evidence for lateral gene transfer within and between prokaryotes and eukaryotes

    Directory of Open Access Journals (Sweden)

    Roger Andrew J

    2003-06-01

    Full Text Available Abstract Background Lateral gene transfer can introduce genes with novel functions into genomes or replace genes with functionally similar orthologs or paralogs. Here we present a study of the occurrence of the latter gene replacement phenomenon in the four gene families encoding different classes of glutamate dehydrogenase (GDH, to evaluate and compare the patterns and rates of lateral gene transfer (LGT in prokaryotes and eukaryotes. Results We extend the taxon sampling of gdh genes with nine new eukaryotic sequences and examine the phylogenetic distribution pattern of the various GDH classes in combination with maximum likelihood phylogenetic analyses. The distribution pattern analyses indicate that LGT has played a significant role in the evolution of the four gdh gene families. Indeed, a number of gene transfer events are identified by phylogenetic analyses, including numerous prokaryotic intra-domain transfers, some prokaryotic inter-domain transfers and several inter-domain transfers between prokaryotes and microbial eukaryotes (protists. Conclusion LGT has apparently affected eukaryotes and prokaryotes to a similar extent within the gdh gene families. In the absence of indications that the evolution of the gdh gene families is radically different from other families, these results suggest that gene transfer might be an important evolutionary mechanism in microbial eukaryote genome evolution.

  12. Interaction between alcohol dehydrogenase II gene, alcohol consumption, and risk for breast cancer

    OpenAIRE

    St?rmer, T; Wang-Gohrke, S; Arndt, V; Boeing, H; Kong, X; Kreienberg, R; Brenner, H

    2002-01-01

    MaeIII Restriction Fragment Length Polymorphism in exon 3 of the alcohol dehydrogenase II was assessed in serum from 467 randomly selected German women and 278 women with invasive breast cancer to evaluate the interaction between a polymorphism of the alcohol dehydrogenase II gene, alcohol consumption and risk for breast cancer. In both groups, usual consumption of different alcoholic beverages was asked for using semiquantitative food frequency questionnaires. We used multivariable logistic ...

  13. Influence of long-term hyper-gravity on the reactivity of succinic acid dehydrogenase and NADPH-diaphorase in the central nervous system of fish: a histochemical study

    Science.gov (United States)

    Anken, R. H.; Rahmann, H.

    In the course of a densitometric evaluation, the histochemically demonstrated reactivity of succinic acid dehydrogenase (SDH) and of NADPH-diaphorase (NADPHD) was determined in different brain nuclei of two teleost fish (cichlid fish Oreochromis mossambicus, swordtail fish Xiphophorus helleri), which had been kept under 3g hyper-gravity for 8 days. SDH was chosen since it is a rate limiting enzyme of the Krebs cycle and therefore it is regarded as a marker for metabolic and neuronal activity. NADPHD reactivity reflects the activity of nitric oxide synthase. Nitric oxide (NO) is a gaseous intercellular messenger that has been suggested to play a major role in several different in vivo models of neuronal plasticity including learning. Within particular vestibulum-connected brain centers, significant effects of hyper-gravity were obtained, e.g., in the magnocellular nucleus, a primary vestibular relay ganglion of the brain stem octavolateralis area, in the superior rectus subdivision of the oculomotoric nucleus and within cerebellar eurydendroid cells, which in teleosts possibly resemble the deep cerebellar nucleus of higher vertebrates. Non-vestibulum related nuclei did not respond to hypergravity in a significant way. The effect of hyper-gravity found was much less distinct in adult animals as compared to the circumstances seen in larval fish (Anken et al., Adv. Space Res. 17, 1996), possibly due to a development correlated loss of neuronal plasticity.

  14. ISOLATION AND CHARACTERIZATION OF THE RAT GENE ENCODING GLUTAMATE-DEHYDROGENASE

    NARCIS (Netherlands)

    DAS, AT; ARNBERG, AC; MALINGRE, H; MOERER, P; CHARLES, R; MOORMAN, AFM; LAMERS, WH

    1993-01-01

    The concentration of glutamate dehydrogenase (GDH) varies strongly between different organs and between different regions within organs. To permit further studies on the regulation of GDH expression, we isolated and characterized the rat gene encoding the GDH protein. This gene contains 13 exons and

  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. Alcoholism and alcohol drinking habits predicted from alcohol dehydrogenase genes

    DEFF Research Database (Denmark)

    Tolstrup, J.S.; Nordestgaard, Børge; Rasmussen, S.

    2008-01-01

    /1 genotype. Results for ADH1B and ADH1C genotypes among men and women were similar. Finally, because slow ADH1B alcohol degradation is found in more than 90% of the white population compared to less than 10% of East Asians, the population attributable risk of heavy drinking and alcoholism by ADH1B.1......Alcohol is degraded primarily by alcohol dehydrogenase (ADH) wherein genetic variation that affects the rate of alcohol degradation is found in ADH1B and ADH1C. It is biologically plausible that these variations may be associated with alcohol drinking habits and alcoholism. By genotyping 9080 white...... men and women from the general population, we found that men and women with ADH1B slow vs fast alcohol degradation drank more alcohol and had a higher risk of everyday drinking, heavy drinking, excessive drinking and of alcoholism. For example, the weekly alcohol intake was 9.8 drinks (95% confidence...

  17. Transgenic barley overexpressing a cytokinin dehydrogenase gene shows greater tolerance to drought stress

    Czech Academy of Sciences Publication Activity Database

    Pospíšilová, H.; Jiskrová, E.; Vojta, P.; Mrízová, K.; Kokáš, F.; Majeská Čudějková, M.; Bergougnoux, V.; Plíhal, O.; Klimešová, J.; Novák, Ondřej; Dzurová, L.; Frébort, I.; Galuszka, P.

    2016-01-01

    Roč. 33, č. 5 (2016), s. 692-705 ISSN 1871-6784 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : ROOT-GROWTH * OXIDASE/DEHYDROGENASE GENES * BETA-GLUCOSIDASE Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.813, year: 2016

  18. Molecular and comparative analysis of the hyperthermostable pyrococcus furiosus glutamate dehydrogenase and its gene

    NARCIS (Netherlands)

    Eggen, Rik I L; Geerling, Ans C M; Voorhorst, Wilfried G B; Kort, Remco; de Vos, Willem M.

    1994-01-01

    The gene for glutamate dehydrogenase (GDH) from Pyrococcus furiosus has been cloned, sequenced and expressed in Escherichia coli. Significant GDH activity could be detected in this host, allowing the further structure-function analysis of this hyperthermostable hexameric enzyme. The deduced primary

  19. Alcoholism and alcohol drinking habits predicted from alcohol dehydrogenase genes

    DEFF Research Database (Denmark)

    Tolstrup, J.S.; Nordestgaard, Børge; Rasmussen, S.

    2008-01-01

    Alcohol is degraded primarily by alcohol dehydrogenase (ADH) wherein genetic variation that affects the rate of alcohol degradation is found in ADH1B and ADH1C. It is biologically plausible that these variations may be associated with alcohol drinking habits and alcoholism. By genotyping 9080 white...... men and women from the general population, we found that men and women with ADH1B slow vs fast alcohol degradation drank more alcohol and had a higher risk of everyday drinking, heavy drinking, excessive drinking and of alcoholism. For example, the weekly alcohol intake was 9.8 drinks (95% confidence......, individuals with ADH1C slow vs fast alcohol degradation had a higher risk of heavy and excessive drinking. For example, the OR for heavy drinking was 1.4 (95% CI: 1.1-1.8) among men with the ADH1C.1/2 genotype and 1.4 (95% CI: 1.0-1.9) among men with the ADH1B.2/2 genotype, compared with men with the ADH1C.1...

  20. Structural organization of the human short-chain acyl-CoA dehydrogenase gene

    DEFF Research Database (Denmark)

    Corydon, M J; Andresen, B S; Bross, P

    1997-01-01

    Short-chain acyl-CoA dehydrogenase (SCAD) is a homotetrameric mitochondrial flavoenzyme that catalyzes the initial reaction in short-chain fatty acid beta-oxidation. Defects in the SCAD enzyme are associated with failure to thrive, often with neuromuscular dysfunction and elevated urinary excreti....... The evolutionary relationship between SCAD and five other members of the acyl-CoA dehydrogenase family was investigated by two independent approaches that gave similar phylogenetic trees....... shown to be associated with ethylmalonic aciduria. From analysis of 18 unrelated Danish families, we show that the four SCAD gene polymorphisms constitute five allelic variants of the SCAD gene, and that the 625A variant together with the less frequent variant form of the three other polymorphisms (321C...

  1. Association of restriction fragment length polymorphism in alcohol dehydrogenase 2 gene with alcohol induced liver damage.

    OpenAIRE

    Sherman, D I; Ward, R J; Warren-Perry, M; Williams, R; Peters, T J

    1993-01-01

    OBJECTIVE--To investigate the role of genetically determined differences in the enzymes of alcohol metabolism in susceptibility to liver damage from misusing alcohol. DESIGN--Use of pADH36 probe to study PVU II restriction length fragment polymorphism in alcohol dehydrogenase 2 gene in white alcohol misusers and controls. SETTING--Teaching hospital referral centres for liver disease and alcohol misuse. SUBJECTS--45 white alcohol misusers (38 with alcoholic liver disease) and 23 healthy contro...

  2. Efficient production of succinic acid from Palmaria palmata hydrolysate by metabolically engineered Escherichia coli.

    Science.gov (United States)

    Olajuyin, Ayobami Matthew; Yang, Maohua; Liu, Yilan; Mu, Tingzhen; Tian, Jiangnan; Adaramoye, Oluwatosin Adekunle; Xing, Jianmin

    2016-08-01

    Succinic acid, a C4 dicarboxylic acid is used in many fields such as food, agriculture, pharmaceutical and polymer industries. In this study, microbial production of succinic acid from Palmaria palmata was investigated for the first time. In engineered Escherichia coli KLPPP, lactate dehydrogenase, pyruvate formate lyase, phosphotransacetylase-acetate kinase and pyruvate oxidase genes were deleted while phosphoenolpyruvate carboxykinase was overexpressed. The recombinant exhibited higher molar yield of succinic acid on galactose (1.20±0.02mol/mol) than glucose (0.48±0.03mol/mol). The concentration and molar yield of succinic acid were 22.40±0.12g/L and 1.13±0.02mol/mol total sugar respectively after 72h dual phase fermentation from P. palmata hydrolysate which composed of glucose (12.57±0.17g/L) and galactose (18.03±0.10g/L). The results demonstrate that P. palmata red macroalgae biomass represents a novel and an economically alternative feedstock for biochemicals production. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Mutations in the medium chain acyl-CoA dehydrogenase (MCAD) gene

    DEFF Research Database (Denmark)

    Tanaka, K; Yokota, I; Coates, P M

    1992-01-01

    Medium chain acyl-CoA dehydrogenase (MCAD) catalyzes the first reaction of the beta-oxidation cycle for 4-10-carbon fatty acids. MCAD deficiency is one of the most frequent inborn metabolic disorders in populations of northwestern European origin. In the compilation of data from a worldwide study...... of 172 unrelated patients each representing an independent pedigree, a total of 8 different mutations have been identified. Among them, a single prevalent mutation, 985A-->G, was found in 90% of 344 variant alleles. 985A-->G causes glutamate substitution for lysine-304 in the mature MCAD subunit, which...... causes impairment of tetramer assembly and instability of the protein. Three of 7 rarer mutations have been identified in a few unrelated patients, while the remaining 4 have each been found in only a single pedigree. In addition to tabulating the mutations, the acyl-CoA dehydrogenase gene family...

  4. 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.

  5. 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.

  6. Identification of stable normalization genes for quantitative real-time PCR in porcine articular cartilage.

    Science.gov (United States)

    McCulloch, Ryan S; Ashwell, Melissa S; O'Nan, Audrey T; Mente, Peter L

    2012-11-12

    Expression levels for genes of interest must be normalized with an appropriate reference, or housekeeping gene, to make accurate comparisons of quantitative real-time PCR results. The purpose of this study was to identify the most stable housekeeping genes in porcine articular cartilage subjected to a mechanical injury from a panel of 10 candidate genes. Ten candidate housekeeping genes were evaluated in three different treatment groups of mechanically impacted porcine articular cartilage. The genes evaluated were: beta actin, beta-2-microglobulin, glyceraldehyde-3-phosphate dehydrogenase, hydroxymethylbilane synthase, hypoxanthine phosphoribosyl transferase, peptidylprolyl isomerase A (cyclophilin A), ribosomal protein L4, succinate dehydrogenase flavoprotein subunit A, TATA box binding protein, and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein-zeta polypeptide. The stability of the genes was measured using geNorm, BestKeeper, and NormFinder software. The four most stable genes measured via geNorm were (most to least stable) succinate dehydrogenase flavoprotein, subunit A, peptidylprolyl isomerase A, glyceraldehyde-3-phosphate dehydrogenase, beta actin; the four most stable genes measured via BestKeeper were glyceraldehyde-3-phosphate dehydrogenase, peptidylprolyl isomerase A, beta actin, succinate dehydrogenase flavoprotein, subunit A; and the four most stable genes measured via NormFinder were peptidylprolyl isomerase A, succinate dehydrogenase flavoprotein, subunit A, glyceraldehyde-3-phosphate dehydrogenase, beta actin. BestKeeper, geNorm, and NormFinder all generated similar results for the most stable genes in porcine articular cartilage. The use of these appropriate reference genes will facilitate accurate gene expression studies of porcine articular cartilage and suggest appropriate housekeeping genes for articular cartilage studies in other species.

  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. The Succinated Proteome

    Energy Technology Data Exchange (ETDEWEB)

    Merkley, Eric D.; Metz, Thomas O.; Smith, Richard D.; Baynes, John; Frizell, Norma

    2014-03-30

    Succination is a chemical modification of cysteine in protein by the Krebs cycle intermediate, fumarate, yielding S-(2-succino)cysteine (2SC). Intracellular fumarate concentration and succination of proteins are increased by hyperpolarization of the inner mitochondrial membrane, in concert with mitochondrial, endoplasmic reticulum (ER) and oxidative stress in adipocytes grown in high glucose medium and in adipose tissue in obesity and diabetes. Increased succination of proteins is also detected in the kidney of a fumarase conditional knock-out mouse which develops renal tumors. Keap1, the gatekeeper of the antioxidant response, was identified as a major succinated protein in renal cancer cells, suggesting that succination may play a role in activation of the antioxidant response. A wide range of proteins is subject to succination, including enzymes, adipokines, cytoskeletal proteins and ER chaperones with functional cysteine residues. There is also significant overlap between succinated and glutathionylated proteins, and with proteins containing cysteine residues that are readily oxidized to the sulfenic (cysteic) acid. Succination of adipocyte proteins is inhibited by uncouplers, which discharge the mitochondrial membrane potential (Δψm) and by ER stress inhibitors. 2SC serves as a biomarker of mitochondrial stress or dysfunction in chronic diseases, such as obesity, diabetes and cancer, and recent studies suggest that succination is a mechanistic link between mitochondrial dysfunction, oxidative and ER stress, and cellular progression toward apoptosis. In this article, we review the history of the succinated proteome and the challenges associated with measuring this non-enzymatic post-translational modification of proteins by proteomics approaches.

  9. Cloning, structure, and chromosome localization of the mouse glutaryl-CoA dehydrogenase gene

    Energy Technology Data Exchange (ETDEWEB)

    Koeller, D.M.; DiGiulio, A.; Frerman, F.E. [Univ. of Colorado Health Sciences Center, Denver, CO (United States)] [and others

    1995-08-10

    Glutaryl-CoA dehydrogenase (GCDH) is a nuclear-encoded, mitochondrial matrix enzyme. In humans, deficiency of GCDH leads to glutaric acidemia type I, and inherited disorder of amino acid metabolism characterized by a progressive neurodegenerative disease. In this report we describe the cloning and structure of the mouse GCDH (Gcdh) gene and cDNA and its chromosomal localization. The mouse Gcdh cDNA is 1.75 kb long and contains and open reading frame of 438 amino acids. The amino acid sequences of mouse, human, and pig GCDH are highly conserved. The mouse Gcdh gene contains 11 exons and spans 7 kb of genomic DNA. Gcdh was mapped by backcross analysis to mouse chromosome 8 within a region that is homologous to a region of human chromosome 19, where the human gene was previously mapped. 14 refs., 3 figs.

  10. [Application of created restriction site PCR-RFLP to identify alcohol dehydrogenase 2 gene polymorphism].

    Science.gov (United States)

    Jiao, Jie; Wang, Wei; Liu, Jing; Xia, Zhaolin

    2009-01-01

    To develop a appropriate assay for identifying single nucleotide polymorphism (SNP) of alcohol dehydrogenase 2 (ADH2) gene. According to base substitution situation of one single base mutational site, we designed the present study primers. One of the primers was designed on the basis of neighbourhood sequence of the mutational site, that is, we made one mismatch base to let product a new enzyme site between the 3' end of the primer and the single base mutation type after the PCR amplification. Then PCR-RFLP was adopted to identify the SNP in ADH2 gene. One primer pair can get target products containing ADH2 SNP site by PCR, restriction enzymes Bsh1236I were adopted to identify the SNP site. The expected results were reached. It suggested that the method of detecting the SNP of ADH2 based on CRS-PCR-RFLP theory is facilitated, economic, and rapid.

  11. 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.

  12. Selenium controls transcription of paralogous formate dehydrogenase genes in the termite gut acetogen, Treponema primitia.

    Science.gov (United States)

    Matson, Eric G; Zhang, Xinning; Leadbetter, Jared R

    2010-08-01

    The termite gut spirochete, Treponema primitia, is a CO(2)-reductive acetogen that is phylogenetically distinct from other distantly related and more extensively studied acetogens such as Moorella thermoacetica. Research on T. primitia has revealed details about the role of spirochetes in CO(2)-reductive acetogenesis, a process important to the mutualism occurring between termites and their gut microbial communities. Here, a locus of the T. primitia genome containing Wood-Ljungdahl pathway genes for CO(2)-reductive acetogenesis was sequenced. This locus contained methyl-branch genes of the pathway (i.e. for the reduction of CO(2) to the level of methyl-tetrahydrofolate) including paralogous genes for cysteine and selenocysteine (Sec) variants of formate dehydrogenase (FDH) and genes for Sec incorporation. The FDH variants affiliated phylogenetically with hydrogenase-linked FDH enzymes, suggesting that T. primitia FDH enzymes utilize electrons derived directly from molecular H(2). Sub-nanomolar concentrations of selenium decreased transcript levels of the cysteine variant FDH gene. Selenium concentration did not markedly influence the level of mRNA upstream of the Sec-codon in the Sec variant FDH; however, the level of transcript extending downstream of the Sec-codon increased incrementally with increasing selenium concentrations. The features and regulation of these FDH genes are an indication that T. primitia may experience dynamic selenium availability in its H(2)-rich gut environment. © 2010 Society for Applied Microbiology and Blackwell Publishing Ltd.

  13. Cloning, characterization, and regulation of the human type II IMP dehydrogenase gene

    Energy Technology Data Exchange (ETDEWEB)

    Glesne, D.A.; Huberman, E. [Argonne National Lab., IL (United States). Center for Mechanistic Biology and Biotechnology]|[Univ. of Chicago, IL (United States)

    1997-01-01

    Human type II inosine 5{prime}-monophosphate dehydrogenase (IMPDH, EC 1.1.1.205) is the rate-limiting enzyme in de novo guanine nucleotide biosynthesis. Regulated IMPDH activity is associated with cellular proliferation, transformation, and differentiation. The authors cloned and sequenced the entire gene for type II IMPDH and here provide details regarding the organization of the gene and the characterization of its promoter. The gene spans approximately 5 kb and is disrupted by 12 introns. The transcriptional start sites were determined by S1 nuclease mapping to be somewhat heterogeneous but predominated at 102 and 85 nucleotides from the translational initiation codon. Through the use of heterologous gene constructs and transient transfection assays, a minimal promoter from {minus}206 to {minus}85 was defined. This promoter is TATA-less and contains several transcription factor motifs including four potential Sp 1 binding sites. The minimal promoter is GC-rich (69%) and resembles a CpG island. Through the use of gel mobility shift assays, nuclear proteins were shown to specifically interact with this minimal promoter. Stable transfectants were used to demonstrate that the down-regulation of IMPDH gene expression in response to reduced cellular proliferation occurs by a transcriptional mechanism.

  14. Lactic acid production by Saccharomyces cerevisiae expressing a Rhizopus oryzae lactate dehydrogenase gene.

    Science.gov (United States)

    Skory, Christopher D

    2003-01-01

    This work demonstrates the first example of a fungal lactate dehydrogenase (LDH) expressed in yeast. A L(+)-LDH gene, ldhA, from the filamentous fungus Rhizopus oryzae was modified to be expressed under control of the Saccharomyces cerevisiae adh1 promoter and terminator and then placed in a 2 micro -containing yeast-replicating plasmid. The resulting construct, pLdhA68X, was transformed and tested by fermentation analyses in haploid and diploid yeast containing similar genetic backgrounds. Both recombinant strains utilized 92 g glucose/l in approximately 30 h. The diploid isolate accumulated approximately 40% more lactic acid with a final concentration of 38 g lactic acid/l and a yield of 0.44 g lactic acid/g glucose. The optimal pH for lactic acid production by the diploid strain was pH 5. LDH activity in this strain remained relatively constant at 1.5 units/mg protein throughout the fermentation. The majority of carbon was still diverted to the ethanol fermentation pathway, as indicated by ethanol yields between 0.25-0.33 g/g glucose. S. cerevisiae mutants impaired in ethanol production were transformed with pLdhA68X in an attempt to increase the lactic acid yield by minimizing the conversion of pyruvate to ethanol. Mutants with diminished pyruvate decarboxylase activity and mutants with disrupted alcohol dehydrogenase activity did result in transformants with diminished ethanol production. However, the efficiency of lactic acid production also decreased.

  15. Regulation of the ald Gene Encoding Alanine Dehydrogenase by AldR in Mycobacterium smegmatis

    Science.gov (United States)

    Jeong, Ji-A; Baek, Eun-Young; Kim, Si Wouk; Choi, Jong-Soon

    2013-01-01

    The regulatory gene aldR was identified 95 bp upstream of the ald gene encoding l-alanine dehydrogenase in Mycobacterium smegmatis. The AldR protein shows sequence similarity to the regulatory proteins of the Lrp/AsnC family. Using an aldR deletion mutant, we demonstrated that AldR serves as both activator and repressor for the regulation of ald gene expression, depending on the presence or absence of l-alanine. The purified AldR protein exists as a homodimer in the absence of l-alanine, while it adopts the quaternary structure of a homohexamer in the presence of l-alanine. The binding affinity of AldR for the ald control region was shown to be increased significantly by l-alanine. Two AldR binding sites (O1 and O2) with the consensus sequence GA-N2-ATC-N2-TC and one putative AldR binding site with the sequence GA-N2-GTT-N2-TC were identified upstream of the ald gene. Alanine and cysteine were demonstrated to be the effector molecules directly involved in the induction of ald expression. The cellular level of l-alanine was shown to be increased in M. smegmatis cells grown under hypoxic conditions, and the hypoxic induction of ald expression appears to be mediated by AldR, which senses the intracellular level of alanine. PMID:23749971

  16. LACTATE DEHYDROGENASE GENE GENOTYPIZATION FOR SPECIES IDENTIFICATION IN A FISH FARM ON THE RIVER NERETVA

    Directory of Open Access Journals (Sweden)

    Polonca Margeta

    2015-09-01

    Full Text Available There are severale Salmonid species, found in the river Neretva basin, among which S. trutta and S. obtusirostris. Also, natural hybrids such as S. obtusirostris x S. trutta have been observed. In one fish farm on the river Neretva, S. trutta and S. obtusirostris were decided to breed separately. Parental fishes were separated phenotypicaly on the basis of the morphological signs. PCR-RFLP analysis of the exon 3 to exon 4 part of the lactate dehydrogenase (LDH C1* gene with restriction endonuclease RsaI was employed to identify the presence of other species representatives or intercrosses in two groups of juvenille fishes. Using this method, we were able to identify two S. trutta representatives in the S. obtusirostris group.

  17. Exploring the potential of the glycerol-3-phosphate dehydrogenase 2 (GPD2) promoter for recombinant gene expression in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Knudsen, Jan Dines; Johanson, Ted; Eliasson Lantz, Anna

    2015-01-01

    A control point for keeping redox homeostasis in Saccharomyces cerevisiae during fermentative growth is the dynamic regulation of transcription for the glycerol-3-phosphate dehydrogenase 2 (GPD2) gene. In this study, the possibility to steer the activity of the GPD2 promoter was investigated by p...

  18. The Alcohol Dehydrogenase Gene Family in Melon (Cucumis melo L.: Bioinformatic Analysis and Expression Patterns

    Directory of Open Access Journals (Sweden)

    Yazhong eJin

    2016-05-01

    Full Text Available Alcohol dehydrogenases (ADH, encoded by multigene family in plants, play a critical role in plant growth, development, adaptation, fruit ripening and aroma production. Thirteen ADH genes were identified in melon genome, including 12 ADHs and one formaldehyde dehydrogenease (FDH, designated CmADH1-12 and CmFDH1, in which CmADH1 and CmADH2 have been isolated in Cantaloupe. ADH genes shared a lower identity with each other at the protein level and had different intron-exon structure at nucleotide level. No typical signal peptides were found in all CmADHs, and CmADH proteins might locate in the cytoplasm. The phylogenetic tree revealed that 13 ADH genes were divided into 3 groups respectively, namely long-, medium- and short-chain ADH subfamily, and CmADH1,3-11, which belongs to the medium-chain ADH subfamily, fell into 6 medium-chain ADH subgroups. CmADH12 may belong to the long-chain ADH subfamily, while CmFDH1 may be a Class III ADH and serve as an ancestral ADH in melon. Expression profiling revealed that CmADH1, CmADH2, CmADH10 and CmFDH1 were moderately or strongly expressed in different vegetative tissues and fruit at medium and late developmental stages, while CmADH8 and CmADH12 were highly expressed in fruit after 20 days. CmADH3 showed preferential expression in young tissues. CmADH4 only had slight expression in root. Promoter analysis revealed several motifs of CmADH genes involved in the gene expression modulated by various hormones, and the response pattern of CmADH genes to ABA, IAA and ethylene were different. These CmADHs were divided into ethylene-sensitive and –insensitive groups, and the functions of CmADHs were discussed.

  19. Improvement of salt tolerance in transgenic potato plants by glyceraldehyde-3 phosphate dehydrogenase gene transfer.

    Science.gov (United States)

    Jeong, M J; Park, S C; Byun, M O

    2001-10-31

    In the previous experiment, we isolated and characterized glyceraldehyde-3-phosphate dehydrogenase (GPD) gene of the oyster mushroom, Pleurotus sajor-caju. Expression levels of the GPD gene in the mycelia of P sajor-caju was significantly increased by exposing the mycelia to abiotic stresses, such as salt, cold, heat, and drought. We also showed that GPD confers abiotic stress resistance when introduced into yeast cells. The survival rate of the transgenic yeast cell that harbored the GPD gene was significantly higher when the yeast cells were subjected to salt, cold, heat, and drought stresses, compared with the yeast that was transformed with the pYES2 vector alone. In order to investigate the functional role of the P. sajor-caju GPD gene in higher plant cells, the complete P. sajor-caju GPD cDNA was fused into the CaMV35S promoter and then introduced into potato plants. Putative potato transformants were screened by using PCR. Twenty-one transformants were further analyzed with RT-PCR to confirm the expression of P. sajor-caju GPD. A RT-PCR Southern blot analysis revealed that 12 transgenics induced the P. sajor-caju GPD gene expression. A bioassay of these transformants revealed that the P. sajor-caju GPD gene was enough to confer salt stress resistance in the potato plant cell system. Results showed that P. sajor-caju GPD, which was continuously expressed in transgenic potato plants under normal growing conditions, resulted in improved tolerance against salt loading.

  20. Exogenous Gene Transmission of Isocitrate Dehydrogenase 2 Mimics Ischemic Preconditioning Protection.

    Science.gov (United States)

    Kolb, Alexander L; Corridon, Peter R; Zhang, Shijun; Xu, Weimin; Witzmann, Frank A; Collett, Jason A; Rhodes, George J; Winfree, Seth; Bready, Devin; Pfeffenberger, Zechariah J; Pomerantz, Jeremy M; Hato, Takashi; Nagami, Glenn T; Molitoris, Bruce A; Basile, David P; Atkinson, Simon J; Bacallao, Robert L

    2018-04-01

    Ischemic preconditioning confers organ-wide protection against subsequent ischemic stress. A substantial body of evidence underscores the importance of mitochondria adaptation as a critical component of cell protection from ischemia. To identify changes in mitochondria protein expression in response to ischemic preconditioning, we isolated mitochondria from ischemic preconditioned kidneys and sham-treated kidneys as a basis for comparison. The proteomic screen identified highly upregulated proteins, including NADP+-dependent isocitrate dehydrogenase 2 (IDH2), and we confirmed the ability of this protein to confer cellular protection from injury in murine S3 proximal tubule cells subjected to hypoxia. To further evaluate the role of IDH2 in cell protection, we performed detailed analysis of the effects of Idh2 gene delivery on kidney susceptibility to ischemia-reperfusion injury. Gene delivery of IDH2 before injury attenuated the injury-induced rise in serum creatinine ( P <0.05) observed in controls and increased the mitochondria membrane potential ( P <0.05), maximal respiratory capacity ( P <0.05), and intracellular ATP levels ( P <0.05) above those in controls. This communication shows that gene delivery of Idh2 can confer organ-wide protection against subsequent ischemia-reperfusion injury and mimics ischemic preconditioning. Copyright © 2018 by the American Society of Nephrology.

  1. Comparative evolutionary genomics of the HADH2 gene encoding Aβ-binding alcohol dehydrogenase/17β-hydroxysteroid dehydrogenase type 10 (ABAD/HSD10

    Directory of Open Access Journals (Sweden)

    Fernandes Pedro A

    2006-08-01

    Full Text Available Abstract Background The Aβ-binding alcohol dehydrogenase/17β-hydroxysteroid dehydrogenase type 10 (ABAD/HSD10 is an enzyme involved in pivotal metabolic processes and in the mitochondrial dysfunction seen in the Alzheimer's disease. Here we use comparative genomic analyses to study the evolution of the HADH2 gene encoding ABAD/HSD10 across several eukaryotic species. Results Both vertebrate and nematode HADH2 genes showed a six-exon/five-intron organization while those of the insects had a reduced and varied number of exons (two to three. Eutherian mammal HADH2 genes revealed some highly conserved noncoding regions, which may indicate the presence of functional elements, namely in the upstream region about 1 kb of the transcription start site and in the first part of intron 1. These regions were also conserved between Tetraodon and Fugu fishes. We identified a conserved alternative splicing event between human and dog, which have a nine amino acid deletion, causing the removal of the strand βF. This strand is one of the seven strands that compose the core β-sheet of the Rossman fold dinucleotide-binding motif characteristic of the short chain dehydrogenase/reductase (SDR family members. However, the fact that the substrate binding cleft residues are retained and the existence of a shared variant between human and dog suggest that it might be functional. Molecular adaptation analyses across eutherian mammal orthologues revealed the existence of sites under positive selection, some of which being localized in the substrate-binding cleft and in the insertion 1 region on loop D (an important region for the Aβ-binding to the enzyme. Interestingly, a higher than expected number of nonsynonymous substitutions were observed between human/chimpanzee and orangutan, with six out of the seven amino acid replacements being under molecular adaptation (including three in loop D and one in the substrate binding loop. Conclusion Our study revealed that HADH

  2. Global sequence diversity of the lactate dehydrogenase gene in Plasmodium falciparum.

    Science.gov (United States)

    Simpalipan, Phumin; Pattaradilokrat, Sittiporn; Harnyuttanakorn, Pongchai

    2018-01-09

    Antigen-detecting rapid diagnostic tests (RDTs) have been recommended by the World Health Organization for use in remote areas to improve malaria case management. Lactate dehydrogenase (LDH) of Plasmodium falciparum is one of the main parasite antigens employed by various commercial RDTs. It has been hypothesized that the poor detection of LDH-based RDTs is attributed in part to the sequence diversity of the gene. To test this, the present study aimed to investigate the genetic diversity of the P. falciparum ldh gene in Thailand and to construct the map of LDH sequence diversity in P. falciparum populations worldwide. The ldh gene was sequenced for 50 P. falciparum isolates in Thailand and compared with hundreds of sequences from P. falciparum populations worldwide. Several indices of molecular variation were calculated, including the proportion of polymorphic sites, the average nucleotide diversity index (π), and the haplotype diversity index (H). Tests of positive selection and neutrality tests were performed to determine signatures of natural selection on the gene. Mean genetic distance within and between species of Plasmodium ldh was analysed to infer evolutionary relationships. Nucleotide sequences of P. falciparum ldh could be classified into 9 alleles, encoding 5 isoforms of LDH. L1a was the most common allelic type and was distributed in P. falciparum populations worldwide. Plasmodium falciparum ldh sequences were highly conserved, with haplotype and nucleotide diversity values of 0.203 and 0.0004, respectively. The extremely low genetic diversity was maintained by purifying selection, likely due to functional constraints. Phylogenetic analysis inferred the close genetic relationship of P. falciparum to malaria parasites of great apes, rather than to other human malaria parasites. This study revealed the global genetic variation of the ldh gene in P. falciparum, providing knowledge for improving detection of LDH-based RDTs and supporting the candidacy of

  3. An Alcohol Dehydrogenase Gene from Synechocystis sp. Confers Salt Tolerance in Transgenic Tobacco

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    So Young Yi

    2017-11-01

    Full Text Available Synechocystis salt-responsive gene 1 (sysr1 was engineered for expression in higher plants, and gene construction was stably incorporated into tobacco plants. We investigated the role of Sysr1 [a member of the alcohol dehydrogenase (ADH superfamily] by examining the salt tolerance of sysr1-overexpressing (sysr1-OX tobacco plants using quantitative real-time polymerase chain reactions, gas chromatography-mass spectrometry, and bioassays. The sysr1-OX plants exhibited considerably increased ADH activity and tolerance to salt stress conditions. Additionally, the expression levels of several stress-responsive genes were upregulated. Moreover, airborne signals from salt-stressed sysr1-OX plants triggered salinity tolerance in neighboring wild-type (WT plants. Therefore, Sysr1 enhanced the interconversion of aldehydes to alcohols, and this occurrence might affect the quality of green leaf volatiles (GLVs in sysr1-OX plants. Actually, the Z-3-hexenol level was approximately twofold higher in sysr1-OX plants than in WT plants within 1–2 h of wounding. Furthermore, analyses of WT plants treated with vaporized GLVs indicated that Z-3-hexenol was a stronger inducer of stress-related gene expression and salt tolerance than E-2-hexenal. The results of the study suggested that increased C6 alcohol (Z-3-hexenol induced the expression of resistance genes, thereby enhancing salt tolerance of transgenic plants. Our results revealed a role for ADH in salinity stress responses, and the results provided a genetic engineering strategy that could improve the salt tolerance of crops.

  4. A rare disease-associated mutation in the medium-chain acyl-CoA dehydrogenase (MCAD) gene changes a conserved arginine, previously shown to be functionally essential in short-chain acyl-CoA dehydrogenase (SCAD)

    DEFF Research Database (Denmark)

    Andresen, B S; Bross, P; Jensen, T G

    1993-01-01

    Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is a serious and potentially fatal inherited defect in the beta-oxidation of fatty acids. Approximately 80% of patients with MCAD deficiency are homozygous for a single disease-causing mutation (G985). The remaining patients (except for a few ......-chain acyl-CoA dehydrogenase (SCAD) gene of a patient with SCAD deficiency, suggesting that the conserved arginine is crucial for formation of active enzyme in the straight-chain acyl-CoA dehydrogenases....

  5. Identification of some ectomycorrhizal basidiomycetes by PCR amplification of their gpd (glyceraldehyde-3-phosphate dehydrogenase) genes.

    Science.gov (United States)

    Kreuzinger, N; Podeu, R; Gruber, F; Göbl, F; Kubicek, C P

    1996-01-01

    Degenerated oligonucleotide primers designed to flank an approximately 1.2-kb fragment of the gene encoding glyceraldehyde-3-phosphate dehydrogenase (gpd) from ascomycetes and basidiomycetes were used to amplify the corresponding gpd fragments from several species of the ectomycorrhizal fungal taxa Boletus, Amanita, and Lactarius. Those from B. edulis, A. muscaria, and L. deterrimus were cloned and sequenced. The respective nucleotide sequences of these gene fragments showed a moderate degree of similarity (72 to 76%) in the protein-encoding regions and only a low degree of similarity in the introns (56 to 66%). Introns, where present, occurred at conserved positions, but the respective positions and numbers of introns in a given taxon varied. The amplified fragment from a given taxon could be distinguished from that of others by both restriction nuclease cleavage analysis and Southern hybridization. A procedure for labeling DNA probes with fluorescein-12-dUTP by PCR was developed. These probes were used in a nonradioactive hybridization assay, with which the gene could be detected in 2 ng of chromosomal DNA of L. deterrimus on slot blots. Taxon-specific amplification was achieved by the design of specific oligonucleotide primers. The application of the gpd gene for the identification of mycorrhizal fungi under field conditions was demonstrated, with Picea abies (spruce) mycorrhizal roots harvested from a northern alpine forest area as well as from a plant-breeding nursery. The interference by inhibitory substances, which sometimes occurred in the DNA extracted from the root-fungus mixture, could be overcome by using very diluted concentrations of template DNA for a first round of PCR amplification followed by a second round with nested oligonucleotide primers. We conclude that gpd can be used to detect ectomycorrhizal fungi during symbiotic interaction. PMID:8795234

  6. Gene transcript accumulation and in situ mRNA hybridization of two putative glutamate dehydrogenase genes in etiolated Glycine max seedlings.

    Science.gov (United States)

    Dimou, M; Tsaniklidis, G; Aivalakis, G; Katinakis, P

    2015-01-01

    Glutamate dehydrogenase (EC 1.4.1.2) is a multimeric enzyme that catalyzes the reversible amination of α-ketoglutarate to form glutamate. We characterized cDNA clones of two Glycine max sequences, GmGDH1 and GmGDH2, that code for putative α- and β-subunits, respectively, of the NADH dependent enzyme. Temporal and spatial gene transcript accumulation studies using semiquantitative RT-PCR and in situ hybridization have shown an overlapping gene transcript accumulation pattern with differences in relative gene transcript accumulation in the organs examined. Detection of NADH-dependent glutamate dehydrogenase activity in situ using a histochemical method showed concordance with the spatial gene transcript accumulation patterns. Our findings suggest that although the two gene transcripts are co-localized in roots of etiolated soybean seedlings, the ratio of the two subunits of the active holoenzyme may vary among tissues.

  7. Novel glutamate dehydrogenase genes show increased transcript and protein abundances in mature tomato fruits.

    Science.gov (United States)

    Ferraro, Gisela; Bortolotti, Santiago; Mortera, Pablo; Schlereth, Armin; Stitt, Mark; Carrari, Fernando; Kamenetzky, Laura; Valle, Estela M

    2012-06-15

    NAD(P)H-glutamate dehydrogenase (GDH, EC 1.4.1.3) contributes to the control of glutamate homeostasis in all living organisms. In bacteria and animals, GDH is a homohexamer allosterically regulated, whereas in plants NADH-GDH (EC 1.4.1.2) is also found as heterohexamer of α- and β-subunits, but its regulation remains undefined. In tomato (Solanum lycopersicum), GDH activity increases during the fruit ripening along with the content of free glutamate, the most abundant amino acid of ripe fruit involved in conferring the genuine tomato flavour. In this work, novel Slgdh-NAD genes were identified in the recently deciphered tomato genome: three encoding the α-subunit (Slgdh-NAD;A1-3) and one additional gene encoding the β-subunit of GDH (Slgdh-NAD;B1) isolated from a genomic library. These genes are located in different chromosomes. Slgdh-NAD;A1-3 show conserved structures, whereas Slgdh-NAD;B1 includes a novel 5'-untranslated exon. Slgdh-NAD;A1-3 transcripts were detected in all tomato tissues examined, showing the highest levels in mature green fruits, contrasting with Slgdh-NAD;B1 transcripts which were detected mainly in roots or in mature fruits when treated with glutamate, NaCl or salicylic acid. Analyses of GDH activity and protein distribution in different tissues of the Micro-Tom cultivar showed that only the active homohexamer of GDH β-subunits was detected in roots while heterohexamers of GDH α- and β-subunits were found in fruits. These results indicate that GDH β-subunit could modulate the heteromeric isoforms of GDH in response to the environment and physiology of the tomato fruit. This information is relevant to manipulate glutamate contents in tomato fruits genetically. Copyright © 2012 Elsevier GmbH. All rights reserved.

  8. Isocitrate dehydrogenase 1 and 2 genes mutations and MGMT methylation in gliomas

    Directory of Open Access Journals (Sweden)

    D. V. Tabakov

    2017-01-01

    Full Text Available Gliomas are the most common brain tumors. It is difficult to detect them at early stages of disease and there is a few available therapies providing significant improvement in survival. Mutations of isocitrate dehydrogenase 1 and 2 genes (IDH1 and IDH2 play significant role in gliomogenesis, diagnostics and selection of patient therapy. We tested the distribution of IDH1 and IDH2 mutations in gliomas of different histological types and grades of malignancy by DNA melting analysis using our protocol with a sensitivity of 5 %. The results of this assay were confirmed by conventional Sanger sequencing. IDH1/2 mutations were detected in 74 % of lower grade gliomas (II and III, World Health Organization and in 14 % of glioblastomas (IV, World Health Organization. Mutation rate in gliomas with oligodendroglioma component were significantly higher then in other glioma types (р = 0.014. The IDH1 mutations was the most common (79 % of general mutation number. IDH1/2 mutations can induce aberrant gene methylation. Detection of methylation rate of the gene encoding for O6-methylguanine-DNA-methyltransferase (MGMT, predictive biomarker for treatment of gliomas with the alkylating agents, has demonstrated a partial association with IDH1/2 mutations. In 73 % of IDH1/2-mutant tumors MGMT promoter methylation were observed. At the same time IDH1/2 mutations were not revealed in 67 % tumors with MGMT promoter methylation. These results indicate existence of another mechanism of MGMT methylation in gliomas. Our data strong support for necessity of both markers testing when patient therapy is selected.

  9. Molecular Analysis of Glucose-6-Phosphate Dehydrogenase Gene Mutations in Bangladeshi Individuals.

    Directory of Open Access Journals (Sweden)

    Suprovath Kumar Sarker

    Full Text Available Glucose-6-phosphate dehydrogenase (G6PD deficiency is a common X-linked human enzyme defect of red blood cells (RBCs. Individuals with this gene defect appear normal until exposed to oxidative stress which induces hemolysis. Consumption of certain foods such as fava beans, legumes; infection with bacteria or virus; and use of certain drugs such as primaquine, sulfa drugs etc. may result in lysis of RBCs in G6PD deficient individuals. The genetic defect that causes G6PD deficiency has been identified mostly as single base missense mutations. One hundred and sixty G6PD gene mutations, which lead to amino acid substitutions, have been described worldwide. The purpose of this study was to detect G6PD gene mutations in hospital-based settings in the local population of Dhaka city, Bangladesh. Qualitative fluorescent spot test and quantitative enzyme activity measurement using RANDOX G6PDH kit were performed for analysis of blood specimens and detection of G6PD-deficient participants. For G6PD-deficient samples, PCR was done with six sets of primers specific for G6PD gene. Automated Sanger sequencing of the PCR products was performed to identify the mutations in the gene. Based on fluorescence spot test and quantitative enzyme assay followed by G6PD gene sequencing, 12 specimens (11 males and one female among 121 clinically suspected patient-specimens were found to be deficient, suggesting a frequency of 9.9% G6PD deficiency. Sequencing of the G6PD-deficient samples revealed c.C131G substitution (exon-3: Ala44Gly in six samples, c.G487A substitution (exon-6:Gly163Ser in five samples and c.G949A substitution (exon-9: Glu317Lys of coding sequence in one sample. These mutations either affect NADP binding or disrupt protein structure. From the study it appears that Ala44Gly and Gly163Ser are the most common G6PD mutations in Dhaka, Bangladesh. This is the first study of G6PD mutations in Bangladesh.

  10. Molecular cloning and expression analysis of the gene encoding proline dehydrogenase from Jatropha curcas L.

    Science.gov (United States)

    Wang, Haibo; Ao, Pingxing; Yang, Shuanglong; Zou, Zhurong; Wang, Shasha; Gong, Ming

    2015-03-01

    Proline dehydrogenase (ProDH) (EC 1.5.99.8) is a key enzyme in the catabolism of proline. The enzyme JcProDH and its complementary DNA (cDNA) were isolated from Jatropha curcas L., an important woody oil plant used as a raw material for biodiesels. It has been classified as a member of the Pro_dh superfamily based on multiple sequence alignment, phylogenetic characterization, and its role in proline catabolism. Its cDNA is 1674 bp in length with a complete open reading frame of 1485 bp, which encodes a polypeptide chain of 494 amino acids with a predicted molecular mass of 54 kD and a pI of 8.27. Phylogenetic analysis indicated that JcProDH showed high similarity with ProDH from other plants. Reverse transcription PCR (RT-PCR) analysis revealed that JcProDH was especially abundant in the seeds and flowers but scarcely present in the stems, roots, and leaves. In addition, the expression of JcProDH increased in leaves experiencing environmental stress such as cold (5 °C), heat (42 °C), salt (300 mM), and drought (30 % PEG6000). The JcProDH protein was successfully expressed in the yeast strain INVSc1 and showed high enzyme activity in proline catabolism. This result confirmed that the JcProDH gene negatively participated in the stress response.

  11. 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.

  12. Associations between a polymorphism in the hydroxysteroid (11-beta) dehydrogenase 1 gene, neuroticism and postpartum depression.

    Science.gov (United States)

    Iliadis, S I; Comasco, E; Hellgren, C; Kollia, N; Sundström Poromaa, I; Skalkidou, A

    2017-01-01

    This study examined the association between a single nucleotide polymorphism in the hydroxysteroid (11-beta) dehydrogenase 1 gene and neuroticism, as well as the possible mediatory role of neuroticism in the association between the polymorphism and postpartum depressive symptoms. 769 women received questionnaires containing the Edinburgh Postnatal Depression Scale (EPDS) at six weeks postpartum and demographic data at pregnancy week 17 and 32 and at six weeks postpartum, as well as the Swedish universities Scales of Personality at pregnancy week 32. Linear regression models showed an association between the GG genotype and depressive symptoms. When neuroticism was introduced in the model, it was associated with EPDS score, whereas the association between the GG genotype and EPDS became borderline significant. A path analysis showed that neuroticism had a mediatory role in the association between the polymorphism and EPDS score. The use of the EPDS, which is a self-reporting instrument. Neuroticism was associated with the polymorphism and had a mediatory role in the association between the polymorphism and postpartum depression. This finding elucidates the genetic background of neuroticism and postpartum depression. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. 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.

  14. 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

  15. Clinical importance of risk variants in the dihydropyrimidine dehydrogenase gene for the prediction of early-onset fluoropyrimidine toxicity

    OpenAIRE

    Froehlich Tanja K.; Amstutz Ursula; Aebi Stefan; Joerger Markus; Largiader Carlo R.

    2015-01-01

    We investigated the clinical relevance of dihydropyrimidine dehydrogenase gene (DPYD) variants to predict severe early onset fluoropyrimidine (FP) toxicity in particular of a recently discovered haplotype hapB3 and a linked deep intronic splice site mutation c.1129 5923C>G. Selected regions of DPYD were sequenced in prospectively collected germline DNA of 500 patients receiving FP based chemotherapy. Associations of DPYD variants and haplotypes with hematologic gastrointestinal infectious and...

  16. Molecular characterization of lipoamide dehydrogenase gene in Trypanosoma cruzi populations susceptible and resistant to benznidazole.

    Science.gov (United States)

    Dos Santos, Paula F; Moreira, Douglas S; Baba, Elio H; Volpe, Caroline M O; Ruiz, Jerônimo C; Romanha, Alvaro J; Murta, Silvane M F

    2016-11-01

    Lipoamide dehydrogenase (LipDH) is a flavin-containing disulfide oxidoreductase from the same group of thioredoxin reductase, glutathione reductase and trypanothione reductase. This enzyme is found in the mitochondria of all aerobic organisms where it takes part in at least three important multienzyme complexes from the citric acid cycle. In this study, we performed a phylogenetic analysis comparing the amino acid sequence of the LipDH from Trypanosoma cruzi (TcLipDH) with the LipDH from other organisms. Subsequently, the copy number of the TcLipDH gene, the mRNA and protein levels, and the enzymatic activity of the LipDH were determined in populations and strains of T. cruzi that were either resistant or susceptible to benznidazole (BZ). In silico analysis showed the presence of two TcLipDH alleles in the T. cruzi genome. It also showed that TcLipDH protein has less than 55% of identity in comparison to the human LipDH, but the active site is conserved in both of them. Southern blot results suggest that the TcLipDH is a single copy gene in the genome of the T. cruzi samples analyzed. Northern blot assays showed one transcript of 2.4 kb in all T. cruzi populations. Northern blot and Real Time RT-PCR data revealed that the TcLipDH mRNA levels were 2-fold more expressed in the BZ-resistant T. cruzi population (17LER) than in its susceptible pair (17WTS). Western blot results revealed that the TcLipDH protein level is 2-fold higher in 17LER sample in comparison to 17WTS sample. In addition, LipDH activity was higher in the 17LER population than in the 17WTS. Sequencing analysis revealed that the amino acid sequences of the TcLipDH from 17WTS and 17LER populations are identical. Our findings show that one of the mechanisms associated with in vitro-induced BZ resistance to T. cruzi correlates with upregulation of LipDH enzyme. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. [Cloning of genes coding for 3-alpha-hydroxysteroid dehydrogenase and for (3-17)-beta-hydroxysteroid dehydrogenase from Pseudomonas testosteroni].

    Science.gov (United States)

    Floch, H H; Abalain, J H; Di Stefano, S; Carré, J L; Abalain-Colloc, M L

    1995-01-01

    A genomic library of Pseudomonas testosteroi total DNA constructed from SauIIIA digests ligated to a lambda gt11 vector was probed with different polyclonal antibodies raised against purified 3 alpha-HSD and (3 beta-17 beta)-HSD. Two different clones reacting with one antibody were selected. The clone reacting with (3-17)beta-HSD antibody contained a 2,661-base pair insert and was found to contained an open reading frame of 765 base pair that corresponds to a protein of 254 amino-acid residues. A 1,492-base pair was inserte in pBR 322 plasmid vector; the recombinant bacterie over expressed the (3-17)beta-HSD gene. The clone reacting with 3 alpha-HSD antibody contained a 1746 base pair insert which contained an open reading frame of 696 base pairs that corresponds to a protein of 231 amino-acid residues. A search for homologous proteins was performed. Distant similarities were found between (3-17)beta-HSD and members of the short-chain alcool dehydrogenase (SCAD) family but no similarity was observed between 3 alpha-HSD and proteins of this family.

  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. PCR-based amplification and heterologous expression of Pseudomonas alcohol dehydrogenase genes from the soil metagenome for biocatalysis.

    Science.gov (United States)

    Itoh, Nobuya; Isotani, Kentaro; Makino, Yoshihide; Kato, Masaki; Kitayama, Kouta; Ishimota, Tuyoshi

    2014-02-05

    The amplification of useful genes from metagenomes offers great biotechnological potential. We employed this approach to isolate alcohol dehydrogenase (adh) genes from Pseudomonas to aid in the synthesis of optically pure alcohols from various ketones. A PCR primer combination synthesized by reference to the adh sequences of known Pseudomonas genes was used to amplify full-length adh genes directly from 17 samples of DNA extracted from soil. Three such adh preparations were used to construct Escherichia coli plasmid libraries. Of the approximately 2800 colonies obtained, 240 putative adh-positive clones were identified by colony-PCR. Next, 23 functional adh genes named using the descriptors HBadh and HPadh were analyzed. The adh genes obtained via this metagenomic approach varied in their DNA and amino acid sequences. Expression of the gene products in E. coli indicated varying substrate specificity. Two representative genes, HBadh-1 and HPadh-24, expressed in E. coli and Pseudomonas putida, respectively, were purified and characterized in detail. The enzyme products of these genes were confirmed to be useful for producing anti-Prelog chiral alcohols. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. Cytokinin oxidase/dehydrogenase genes in barley and wheat. Cloning and heterologous expression

    Czech Academy of Sciences Publication Activity Database

    Galuszka, P.; Frébortová, Jitka; Werner, T.; Yamada, M.; Strnad, Miroslav; Schmülling, T.; Frébort, I.

    2004-01-01

    Roč. 271, č. 20 (2004), s. 3990-4002 ISSN 0014-2956 Institutional research plan: CEZ:AV0Z5038910 Keywords : cereals * cloning * cytokinin oxidase/dehydrogenase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.260, year: 2004

  1. Mutations in the isocitrate dehydrogenase genes IDH1 and IDH2 in tumors

    NARCIS (Netherlands)

    Schaap, Frank G.; French, Pim J.; Bovée, Judith V. M. G.

    2013-01-01

    Heterozygous hotspot mutations in isocitrate dehydrogenases (IDH) IDH1 or IDH2 are frequently observed in specific types of cartilaginous tumors, gliomas, and leukemias. Mutant IDH enzyme loses its normal activity to convert isocitrate into α-ketoglutarate (αKG) and instead acquires the ability to

  2. Structural organization of the human short-chain acyl-CoA dehydrogenase gene

    DEFF Research Database (Denmark)

    Corydon, M J; Andresen, B S; Bross, P

    1997-01-01

    Short-chain acyl-CoA dehydrogenase (SCAD) is a homotetrameric mitochondrial flavoenzyme that catalyzes the initial reaction in short-chain fatty acid beta-oxidation. Defects in the SCAD enzyme are associated with failure to thrive, often with neuromuscular dysfunction and elevated urinary excreti...

  3. Several homozygous mutations in the gene for 11{beta}-hydroxysteroid dehydrogenase type 2 in patients with apparent mineralocorticoid excess

    Energy Technology Data Exchange (ETDEWEB)

    Wilson, R.C.; Harbison, M.D.; Hanauske-Abel, H.M.; Licholai, T. [New York Hospital-Cornell Medical Center, New York, NY (United States)] [and others

    1995-10-01

    Four deleterious mutations are described in the gene for HSD11B2, which encodes the type 2 isoenzyme of 11{beta}-hydroxysteroid dehydrogenase (11{beta}HSD2). In seven families with one or more members affected by apparent mineralocortiocoid excess, this disorder is shown to be the result of a deficiency in 11{beta}HSD2. Surprisingly, the patients are all homozygous for their mutation. This results from consanguinity in two families and possibly from endogamy or a founder effect in four of the other five families. The absence of compound heterozygotes remains to be investigated. 25 refs., 3 figs., 2 tabs.

  4. Sorbitol synthesis by an engineered Lactobacillus casei strain expressing a sorbitol-6-phosphate dehydrogenase gene within the lactose operon.

    Science.gov (United States)

    Nissen, Lorenzo; Pérez-Martínez, Gaspar; Yebra, María J

    2005-08-01

    Sorbitol is claimed to have important health-promoting effects and Lactobacillus casei is a lactic acid bacterium relevant as probiotic and used as a cheese starter culture. A sorbitol-producing L. casei strain might therefore be of considerable interest in the food industry. A recombinant strain of L. casei was constructed by the integration of a d-sorbitol-6-phosphate dehydrogenase-encoding gene (gutF) in the chromosomal lactose operon (strain BL232). gutF expression in this strain followed the same regulation as that of the lac genes, that is, it was repressed by glucose and induced by lactose. (13)C-nuclear magnetic resonance analysis of supernatants of BL232 resting cells demonstrated that, when pre-grown on lactose, cells were able to synthesize sorbitol from glucose. Inactivation of the l-lactate dehydrogenase gene in BL232 led to an increase in sorbitol production, suggesting that the engineered route provides an alternative pathway for NAD(+) regeneration.

  5. New recombinant bacterium comprises a heterologous gene encoding glycerol dehydrogenase and/or an up-regulated native gene encoding glycerol dehydrogenase, useful for producing ethanol

    DEFF Research Database (Denmark)

    2010-01-01

    TECHNOLOGY FOCUS - BIOTECHNOLOGY - Preparation (claimed): Producing recombinant bacterium having enhanced ethanol production characteristics when cultivated in growth medium comprising glycerol comprises: (a) transforming a parental bacterium by (i) the insertion of a heterologous gene encoding...

  6. 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.

  7. Overexpression of the methanol dehydrogenase gene mxaF in Methylobacterium sp. MB200 enhances L-serine production.

    Science.gov (United States)

    Chao, H; Wu, B; Shen, P

    2015-10-01

    Increase in L-serine production is of interest for industry. Here, we describe a metabolic engineering approach to increase the production of L-serine in a methylotrophic bacterium. mxaF, the gene encoding the large subunit of a methanol dehydrogenase, was cloned from Methylobacterium sp. MB200 through transposon mutagenesis. Deletion of mxaF gene prevented the strain to grow on methanol, suggesting that mxaF is involved in methanol metabolism. Overexpression of mxaF gene in the strain MB200 resulted in a fivefold increase in methanol dehydrogenase activity compared to the wild-type. Resting cell assays showed that the recombinant strain accumulated 6·6 mg ml(-1) L-serine in 72 h with 30 mg ml(-1) wet cells from 50 mg ml(-1) glycine and 50 mg ml(-1) methanol, representing a 1·5-fold increment for L-serine production in contrast to the wild-type strain. These results demonstrate that the potential for improving the production of L-serine can be achieved by overexpressing mxaF gene in methylotrophic bacteria. The amount of L-serine produced each year worldwide is relatively small compared with the amounts of the other amino acids and hence it is in great demand. Here, we describe a metabolic engineering approach to increase the production of L-serine in a methylotrophic bacterium Methylobacterium sp. MB200. The result demonstrates that raising the output of L-serine can be achieved by overexpressing mxaF gene in methylotrophic bacteria. © 2015 The Society for Applied Microbiology.

  8. [Cloning and prokaryotic expression of malate dehydrogenase gene of Taenia saginata asiatica and immunogenicity analysis of the recombinant protein].

    Science.gov (United States)

    Huang, Jiang; Hu, Xu-Chu; Wu, Xuan; Xu, Jin; Yu, Xin-Bing; Bao, Huai-En; Lang, Shu-Yuan; Liao, Xing-Jiang

    2008-08-01

    To clone and express the lactate dehydrogenase (LDH) gene of Taenia saginata asiatica and analyze the immunogenicity of the recombinant protein. By screening the full length cDNA plasmid library, the coding region of LDH was amplified with PCR, and cloned into the prokaryotic expression vector pET-30a (+), then expressed in E. coli BL21 with IPTG induction. The recombinant protein was detected by SDS-PAGE and purified by Ni-IDA affinity chromatography, and its immunogenicity was analyzed by Western blotting. PCR, double enzyme digestion and DNA sequencing confirmed that the recombinant expression plasmid was constructed. The expression products were obtained and purified by Ni-IDA affinity chromatography. Western blotting analysis of LDH recombinant protein testified that the recombinant protein could be recognized by sera of the Taenia saginata asiatica infected swine and the patient. The LDH gene of Taenia saginata asiatica has been cloned and expressed, and the purified protein has been confirmed with immunogenicity.

  9. Combined inactivation of the Clostridium cellulolyticum lactate and malate dehydrogenase genes substantially increases ethanol yield from cellulose and switchgrass fermentations

    Energy Technology Data Exchange (ETDEWEB)

    Li, Yongchao [ORNL; Tschaplinski, Timothy J [ORNL; Engle, Nancy L [ORNL; Hamilton, Choo Yieng [ORNL; Rodriguez, Jr., Miguel [ORNL; Liao, James C [ORNL; Schadt, Christopher Warren [ORNL; Guss, Adam M [ORNL; Yang, Yunfeng [ORNL; Graham, David E [ORNL

    2012-01-01

    Background: The model bacterium Clostridium cellulolyticum efficiently hydrolyzes crystalline cellulose and hemicellulose, using cellulosomes to degrade lignocellulosic biomass. Although it imports and ferments both pentose and hexose sugars to produce a mixture of ethanol, acetate, lactate, H2 and CO2, the proportion of ethanol is low, which impedes its use in consolidated bioprocessing for biofuels. Therefore genetic engineering will likely be required to improve the ethanol yield. Random mutagenesis, plasmid transformation, and heterologous expression systems have previously been developed for C. cellulolyticum, but targeted mutagenesis has not been reported for this organism. Results: The first targeted gene inactivation system was developed for C. cellulolyticum, based on a mobile group II intron originating from the Lactococcus lactis L1.LtrB intron. This markerless mutagenesis system was used to disrupt both the paralogous L-lactate dehydrogenase (Ccel_2485; ldh) and L-malate dehydrogenase (Ccel_0137; mdh) genes, distinguishing the overlapping substrate specificities of these enzymes. Both mutations were then combined in a single strain. This double mutant produced 8.5-times more ethanol than wild-type cells growing on crystalline cellulose. Ethanol constituted 93% of the major fermentation products (by molarity), corresponding to a molar ratio of ethanol to organic acids of 15, versus 0.18 in wild-type cells. During growth on acid-pretreated switchgrass, the double mutant also produced four-times as much ethanol as wild-type cells. Detailed metabolomic analyses identified increased flux through the oxidative branch of the mutant s TCA pathway. Conclusions: The efficient intron-based gene inactivation system produced the first gene-targeted mutations in C. cellulolyticum. As a key component of the genetic toolbox for this bacterium, markerless targeted mutagenesis enables functional genomic research in C. cellulolyticum and rapid genetic engineering to

  10. 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.

  11. Quantification of the expression of reference and alcohol dehydrogenase genes of some acetic acid bacteria in different growth conditions.

    Science.gov (United States)

    Quintero, Y; Poblet, M; Guillamón, J M; Mas, A

    2009-02-01

    The aim of this study was to develop a reliable system to analyse the expression of the pyrroloquinoline quinone (PQQ)-alcohol dehydrogenase (ADH) and test its ability to predict the growth and oxidative activity of some acetic acid bacteria (AAB). Specific primers were designed for use in RT-PCR to quantify ADH expression and several housekeeping genes in four species of AAB. 16S rRNA gene was selected as an internal control. The relative expression of adhA was measured in Acetobacter aceti, Acetobacter pasteurianus, Gluconacetobacter hansenii and Gluconobacter oxydans grown in two media that had glucose or ethanol as the carbon source. AAB adhA expression was shown to be related to the two Acetobacter species' ability to oxidise and grow on ethanol, whereas G. oxydans were unable to grow on ethanol and the growth of Ga. hansenii was not related to adhA expression. The differential expression of ADH could be a marker to analyse both growth and oxidation ability in some AAB, especially those of the genus Acetobacter. Several housekeeping genes were tested in AAB and after growth in different media and it was evident that only the ribosomal coding genes were adequate as reference genes for RT-PCR.

  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. 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.

  14. 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.

  15. Regulatory region with putA gene of proline dehydrogenase that links to the lum and the lux operons in Photobacterium leiognathi.

    Science.gov (United States)

    Lin, J W; Yu, K Y; Chen, H Y; Weng, S F

    1996-02-27

    Nucleotide sequence of regulatory region (R & R) with putA gene (EMBL Accession No. U39227) from Photobacterium leiognathi PL741 has been determined, and the putA gene encoded amino acid sequence of proline dehydrogenase is deduced. Alignment and comparison of proline dehydrogenase of P. leiognathi with the proline dehydrogenase domain in the PutA protein of Escherichia coli and Salmonella typhimurium show that they are homologous. Nucleotide sequence reveals that regulatory region with the putA gene is linked to the lum and lux operons in genome; the gene order is putA--R & R(I) (R & R: regulatory region; ter:transcriptional terminator), whereas the R & R is the regulatory region for the lum and the lux operons, ter is the transcriptional terminator for the lum operon, and R & R(I) apparently is the regulatory region for the putA and related genes. Nucleotide sequence analysis illustrates the specific inverted repeat (SIR), cAMP-CRP consensus sequence, canonical -10/-35 promoter, putative operator and Shine-Dalgarno (SD) sequence on the regulatory region R & R(I) for the putA and related genes; it suggests that the putA and related genes are simply linked to the lum and the lux operons in genome, the regulatory region R & R(I) is independent for the putA and related genes.

  16. Molecular and functional characterization of the kstD2 gene of Rhodococcus erythropolis SQ1 encoding a second 3-ketosteroid Δ1-dehydrogenase isoenzyme

    NARCIS (Netherlands)

    Geize, Robert van der; Hessels, Gerda I.; Dijkhuizen, Lubbert

    2002-01-01

    Previously, Rhodococcus erythropolis SQ1 kstD, encoding ketosteroid Δ1-dehydrogenase (KSTD1) was characterized. Surprisingly, a kstD gene deletion mutant (strain RG1) grew normally on steroids. UV mutagenesis of strain RG1 allowed isolation of strains (e.g. strain RG1-UV29) unable to perform the

  17. Effect of deletion of 2,3-butanediol dehydrogenase gene (bdhA) on acetoin production of Bacillus subtilis.

    Science.gov (United States)

    Zhang, Junjiao; Zhao, Xiangying; Zhang, Jiaxiang; Zhao, Chen; Liu, Jianjun; Tian, Yanjun; Yang, Liping

    2017-09-14

    The present work aims to block 2,3-butanediol synthesis in acetoin fermentation of Bacillus subtilis. First, we constructed a recombinant strain BS168D by deleting the 2,3-butanediol dehydrogenase gene bdhA of the B. subtilis168, and there was almost no 2,3-butanediol production in 20 g/L of glucose media. The acetoin yield of BS168D reached 6.61 g/L, which was about 1.5 times higher than that of the control B. subtilis168 (4.47 g/L). Then, when the glucose concentration was increased to 100 g/L, the acetoin yield reached 24.6 g/L, but 2.4 g/L of 2,3-butanediol was detected at the end of fermentation. The analysis of 2,3-butanediol chiral structure indicated that the main 2,3-butanediol production of BS168D was meso-2,3-butanediol, and the bdhA gene was only responsible for (2R,3R)-2,3-butanediol synthesis. Therefore, we speculated that there may exit another pathway relating to the meso-2,3-butanediol synthesis in the B. subtilis. In addition, the results of low oxygen condition fermentation showed that deletion of bdhA gene successfully blocked the reversible transformation between acetoin and 2,3-butanediol and eliminated the effect of dissolved oxygen on the transformation.

  18. Receptor structure-based discovery of non-metabolite agonists for the succinate receptor GPR91.

    Science.gov (United States)

    Trauelsen, Mette; Rexen Ulven, Elisabeth; Hjorth, Siv A; Brvar, Matjaz; Monaco, Claudia; Frimurer, Thomas M; Schwartz, Thue W

    2017-12-01

    Besides functioning as an intracellular metabolite, succinate acts as a stress-induced extracellular signal through activation of GPR91 (SUCNR1) for which we lack suitable pharmacological tools. Here we first determined that the cis conformation of the succinate backbone is preferred and that certain backbone modifications are allowed for GPR91 activation. Through receptor modeling over the X-ray structure of the closely related P2Y1 receptor, we discovered that the binding pocket is partly occupied by a segment of an extracellular loop and that succinate therefore binds in a very different mode than generally believed. Importantly, an empty side-pocket is identified next to the succinate binding site. All this information formed the basis for a substructure-based search query, which, combined with molecular docking, was used in virtual screening of the ZINC database to pick two serial mini-libraries of a total of only 245 compounds from which sub-micromolar, selective GPR91 agonists of unique structures were identified. The best compounds were backbone-modified succinate analogs in which an amide-linked hydrophobic moiety docked into the side-pocket next to succinate as shown by both loss- and gain-of-function mutagenesis. These compounds displayed GPR91-dependent activity in altering cytokine expression in human M2 macrophages similar to succinate, and importantly were devoid of any effect on the major intracellular target, succinate dehydrogenase. These novel, synthetic non-metabolite GPR91 agonists will be valuable both as pharmacological tools to delineate the GPR91-mediated functions of succinate and as leads for the development of GPR91-targeted drugs to potentially treat low grade metabolic inflammation and diabetic complications such as retinopathy and nephropathy. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.

  19. Isolation, characterization, and nucleotide sequence of the Streptococcus mutans mannitol-phosphate dehydrogenase gene and the mannitol-specific factor III gene of the phosphoenolpyruvate phosphotransferase system.

    Science.gov (United States)

    Honeyman, A L; Curtiss, R

    1992-08-01

    Streptococcus mutans, the causative agent of dental caries, utilizes carbohydrates by means of the phosphoenolpyruvate-dependent phosphotransferase system (PTS). The PTS facilitates vectorial translocation of metabolizable carbohydrates to form the corresponding sugar-phosphates, which are subsequently converted to glycolytic intermediates. The PTS consists of both sugar-specific and sugar-independent components. Complementation of an Escherichia coli mtlD mutation with a streptococcal recombinant DNA library allowed isolation of the mannitol-1-phosphate dehydrogenase gene (mtlD) and the adjacent sugar-specific mannitol factor III gene (mtlF) from S. mutans. Subsequent transposon mutagenesis of the complementing DNA fragment with Tn5seq1 defined the region that encodes the mtlD-complementing activity, the streptococcal mtlD gene. Nucleotide sequence analysis of this region revealed two complete open reading frames (ORFs) from within the streptococcal mannitol PTS operon. One ORF encodes the mtlD gene product, a 43.0-kDa protein which exhibits similarity to the E. coli and Enterococcus faecalis mannitol-1-phosphate dehydrogenases. The second ORF encodes a 15.8-kDa protein which exhibits similarity to mannitol factor III proteins from several bacterial species. In vitro transcription-translation assays were used to produce proteins of the sizes predicted by the streptococcal ORFs. These data indicate that the S. mutans mannitol PTS utilizes an enzyme II-factor III complex similar to the mannitol system found in other gram-positive organisms, as opposed to that of E. coli, which utilizes an independent enzyme II system.

  20. Disparate sequence characteristics of the Erysiphe graminis f.sp. hordei glyceraldehyde-3-phosphate dehydrogenase gene

    DEFF Research Database (Denmark)

    Christiansen, S.K.; Justesen, A.F.; Giese, H.

    1997-01-01

    , Egh falls into the group of Ascomycetes located at a basal position. The regulatory region of the Egh gpd gene has no homology to corresponding sequences in other filamentous Ascomycetes. Codon usage was determined for the four characterized Egh genes (tub2, Egh7, Egh16 and gpd) and found...

  1. Cloning, Expression, and Characterization of budC Gene Encoding meso-2,3-Butanediol Dehydrogenase from Bacillus licheniformis.

    Science.gov (United States)

    Xu, Guo-Chao; Bian, Ya-Qian; Han, Rui-Zhi; Dong, Jin-Jun; Ni, Ye

    2016-02-01

    The budC gene encoding a meso-2,3-butanediol dehydrogenase (BlBDH) from Bacillus licheniformis was cloned and overexpressed in Escherichia coli BL21(DE3). Sequence analysis reveals that this BlBDH belongs to short-chain dehydrogenase/reductase (SDR) superfamily. In the presence of NADH, BlBDH catalyzes the reduction of diacetyl to (3S)-acetoin (97.3% ee), and further to (2S,3S)-2,3-butanediol (97.3% ee and 96.5% de). Similar to other meso-2,3-BDHs, it shows oxidative activity to racemic 2,3-butanediol whereas no activity toward racemic acetoin in the presence of NAD(+). For diacetyl reduction and 2,3-butanediol oxidation, the pH optimum of BlBDH is 5.0 and 10.0, respectively. Unusually, it shows relatively high activity over a wide pH range from 5.0 to 8.0 for racemic acetoin reduction. BlBDH shows lower K m and higher catalytic efficiency toward racemic acetoin (K m = 0.47 mM, k cat /K m = 432 s(-1)·mM(-1)) when compared with 2,3-butanediol (K m = 7.25 mM, k cat /K m = 81.5 s(-1)·mM(-1)), indicating its physiological role in favor of reducing racemic acetoin into 2,3-butanediol. The enzymatic characterization of BlBDH provides evidence for the directed engineering of B. licheniformis for producing enantiopure 2,3-butanediol.

  2. Synthesis of carbon-14 labeled doxylamine succinate

    Energy Technology Data Exchange (ETDEWEB)

    Rao, P.N.; Damodaran, K.M.

    1986-05-01

    Doxylamine succinate, N,N-dimethyl-2-(1-phenyl-1-(2-pyridinyl)-ethoxy)ethanamine succinate is an antihistamine used primarily as a sedative. Carbon-14 labeled doxylamine succinate, required for toxicological studies, was synthesized in two steps starting from 2-benzoyl pyridine.

  3. 21 CFR 582.1091 - Succinic acid.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Succinic acid. 582.1091 Section 582.1091 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS....1091 Succinic acid. (a) Product. Succinic acid. (b) Conditions of use. This substance is generally...

  4. Feeding hydroalcoholic extract powder of Lepidium meyenii (maca) enhances testicular gene expression of 3β-hydroxysteroid dehydrogenase in rats.

    Science.gov (United States)

    Ohta, Y; Kawate, N; Inaba, T; Morii, H; Takahashi, K; Tamada, H

    2017-12-01

    Although feeding diets containing the extract powder of Lepidium meyenii (maca), a plant growing in Peru's Central Andes, increases serum testosterone concentration associated with enhanced ability of testosterone production by Leydig cells in male rats, changes in testicular steroidogenesis-related factors by the maca treatment are not known. This study examined the effects of maca on testicular gene expressions for luteinizing hormone receptor, steroidogenic acute regulatory protein and steroidogenic enzymes. Eight-week-old male rats were given the diets with or without (control) the maca extract powder (2%) for 6 weeks, and mRNA levels were determined by reverse transcription quantitative real-time PCR. The results showed that the testicular mRNA level of HSD3B1 (3β-hydroxysteroid dehydrogenase; 3β-HSD) increased by the treatment, whereas the levels of the other factors examined did not change. These results suggest that increased expression of 3β-HSD gene may be involved in the enhanced steroidogenic ability by the maca treatment in rat testes. © 2017 Blackwell Verlag GmbH.

  5. Identification of four new mutations in the short-chain acyl-CoA dehydrogenase (SCAD) gene in two patients

    DEFF Research Database (Denmark)

    Gregersen, N; Winter, V S; Corydon, M J

    1998-01-01

    We have shown previously that a variant allele of the short-chain acyl-CoA dehydrogenase ( SCAD ) gene, 625G-->A, is present in homozygous form in 7% of control individuals and in 60% of 135 patients with elevated urinary excretion of ethylmalonic acid (EMA). We have now characterized three disease......-causing mutations (confirmed by lack of enzyme activity after expression in COS-7 cells) and a new susceptibility variant in the SCAD gene of two patients with SCAD deficiency, and investigated their frequency in patients with elevated EMA excretion. The first SCAD-deficient patient was a compound heterozygote...... that the allele 511T-625G-like 511C-625A-confers susceptibility to ethylmalonic aciduria. Expression of the variant R147W SCAD protein, encoded by the 511T-625G allele, in COS-7 cells showed 45% activity at 37 degrees C in comparison with the wild-type protein, comparable levels of activity at 26 degrees C...

  6. Identification of point mutations in Glucose-6-Phosphate Dehydrogenase gene in Timor Island people : A preliminary report

    Directory of Open Access Journals (Sweden)

    Widanto Hardjowasito

    2001-12-01

    Full Text Available Glucose 6 phosphate dehydrogenase (G6PD deficiency is common in malaria endemic region, however no molecular study has been performed on G6PD deficiency in Timor Island, Indonesia a malarial hyperendemic area which Proto Malay is the majority of the people in that island. To observe the frequency and molecular type of mutations in G6PD deficient Proto Malay people, 118 native people were screened using formazan ring test. Mutation in the G6PD gene were determined by MPTP (Multiple PCR using Multiple Tandem Forward Primers and a common Reserve Pimer method and confirmed by automatic sequencer. This study shows that three males have lower G6PD activity. Using MPTP method, a point mutation could be indicated in the two cases. Sequencing of the amplified products in 2 G6PD patients disclosed mutations of T383C in exon 5 and C 592 T in exon 6 in respective case. Our result documents point mutations in exon 5 and exon 6 in the G6PD gene of two Proto Malay people in Timor. These mutations are common in Asia region. (Med J Indones 2001; 10: 210-3Keywords: mutations, G6PD, Proto Malay.

  7. Metabolite Fingerprinting in Transgenic Nicotiana tabacum Altered by the Escherichia coli Glutamate Dehydrogenase Gene

    Directory of Open Access Journals (Sweden)

    R. Mungur

    2005-01-01

    Full Text Available With about 200 000 phytochemicals in existence, identifying those of biomedical significance is a mammoth task. In the postgenomic era, relating metabolite fingerprints, abundances, and profiles to genotype is also a large task. Ion analysis using Fourier transformed ion cyclotron resonance mass spectrometry (FT-ICR-MS may provide a high-throughput approach to measure genotype dependency of the inferred metabolome if reproducible techniques can be established. Ion profile inferred metabolite fingerprints are coproducts. We used FT-ICR-MS-derived ion analysis to examine gdhA (glutamate dehydrogenase (GDH; EC 1.4.1.1 transgenic Nicotiana tabacum (tobacco carrying out altered glutamate, amino acid, and carbon metabolisms, that fundamentally alter plant productivity. Cause and effect between gdhA expression, glutamate metabolism, and plant phenotypes was analyzed by 13NH4+ labeling of amino acid fractions, and by FT-ICR-MS analysis of metabolites. The gdhA transgenic plants increased 13N labeling of glutamate and glutamine significantly. FT-ICR-MS detected 2 012 ions reproducible in 2 to 4 ionization protocols. There were 283 ions in roots and 98 ions in leaves that appeared to significantly change abundance due to the measured GDH activity. About 58% percent of ions could not be used to infer a corresponding metabolite. From the 42% of ions that inferred known metabolites we found that certain amino acids, organic acids, and sugars increased and some fatty acids decreased. The transgene caused increased ammonium assimilation and detectable ion variation. Thirty-two compounds with biomedical significance were altered in abundance by GDH including 9 known carcinogens and 14 potential drugs. Therefore, the GDH transgene may lead to new uses for crops like tobacco.

  8. Improved Production of 2,3-Butanediol in Bacillus amyloliquefaciens by Over-Expression of Glyceraldehyde-3-Phosphate Dehydrogenase and 2,3-butanediol Dehydrogenase

    Science.gov (United States)

    Yang, Taowei; Rao, Zhiming; Zhang, Xian; Xu, Meijuan; Xu, Zhenghong; Yang, Shang-Tian

    2013-01-01

    Background Previously, a safe strain, Bacillus amyloliquefaciens B10-127 was identified as an excellent candidate for industrial-scale microbial fermentation of 2,3-butanediol (2,3-BD). However, B. amyloliquefaciens fermentation yields large quantities of acetoin, lactate and succinate as by-products, and the 2,3-BD yield remains prohibitively low for commercial production. Methodology/Principal Findings In the 2,3-butanediol metabolic pathway, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyzes the conversion of 3-phosphate glyceraldehyde to 1,3-bisphosphoglycerate, with concomitant reduction of NAD+ to NADH. In the same pathway, 2,3-BD dehydrogenase (BDH) catalyzes the conversion of acetoin to 2,3-BD with concomitant oxidation of NADH to NAD+. In this study, to improve 2,3-BD production, we first over-produced NAD+-dependent GAPDH and NADH-dependent BDH in B. amyloliquefaciens. Excess GAPDH reduced the fermentation time, increased the 2,3-BD yield by 12.7%, and decreased the acetoin titer by 44.3%. However, the process also enhanced lactate and succinate production. Excess BDH increased the 2,3-BD yield by 16.6% while decreasing acetoin, lactate and succinate production, but prolonged the fermentation time. When BDH and GAPDH were co-overproduced in B. amyloliquefaciens, the fermentation time was reduced. Furthermore, in the NADH-dependent pathways, the molar yield of 2,3-BD was increased by 22.7%, while those of acetoin, lactate and succinate were reduced by 80.8%, 33.3% and 39.5%, relative to the parent strain. In fed-batch fermentations, the 2,3-BD concentration was maximized at 132.9 g/l after 45 h, with a productivity of 2.95 g/l·h. Conclusions/Significance Co-overexpression of bdh and gapA genes proved an effective method for enhancing 2,3-BD production and inhibiting the accumulation of unwanted by-products (acetoin, lactate and succinate). To our knowledge, we have attained the highest 2,3-BD fermentation yield thus far reported for safe

  9. Improved production of 2,3-butanediol in Bacillus amyloliquefaciens by over-expression of glyceraldehyde-3-phosphate dehydrogenase and 2,3-butanediol dehydrogenase.

    Directory of Open Access Journals (Sweden)

    Taowei Yang

    Full Text Available Previously, a safe strain, Bacillus amyloliquefaciens B10-127 was identified as an excellent candidate for industrial-scale microbial fermentation of 2,3-butanediol (2,3-BD. However, B. amyloliquefaciens fermentation yields large quantities of acetoin, lactate and succinate as by-products, and the 2,3-BD yield remains prohibitively low for commercial production.In the 2,3-butanediol metabolic pathway, glyceraldehyde-3-phosphate dehydrogenase (GAPDH catalyzes the conversion of 3-phosphate glyceraldehyde to 1,3-bisphosphoglycerate, with concomitant reduction of NAD(+ to NADH. In the same pathway, 2,3-BD dehydrogenase (BDH catalyzes the conversion of acetoin to 2,3-BD with concomitant oxidation of NADH to NAD(+. In this study, to improve 2,3-BD production, we first over-produced NAD(+-dependent GAPDH and NADH-dependent BDH in B. amyloliquefaciens. Excess GAPDH reduced the fermentation time, increased the 2,3-BD yield by 12.7%, and decreased the acetoin titer by 44.3%. However, the process also enhanced lactate and succinate production. Excess BDH increased the 2,3-BD yield by 16.6% while decreasing acetoin, lactate and succinate production, but prolonged the fermentation time. When BDH and GAPDH were co-overproduced in B. amyloliquefaciens, the fermentation time was reduced. Furthermore, in the NADH-dependent pathways, the molar yield of 2,3-BD was increased by 22.7%, while those of acetoin, lactate and succinate were reduced by 80.8%, 33.3% and 39.5%, relative to the parent strain. In fed-batch fermentations, the 2,3-BD concentration was maximized at 132.9 g/l after 45 h, with a productivity of 2.95 g/l·h.Co-overexpression of bdh and gapA genes proved an effective method for enhancing 2,3-BD production and inhibiting the accumulation of unwanted by-products (acetoin, lactate and succinate. To our knowledge, we have attained the highest 2,3-BD fermentation yield thus far reported for safe microorganisms.

  10. Identification and Characterization of the Glucose-6-Phosphate Dehydrogenase Gene Family in the Para Rubber Tree, Hevea brasiliensis.

    Science.gov (United States)

    Long, Xiangyu; He, Bin; Fang, Yongjun; Tang, Chaorong

    2016-01-01

    As a key enzyme in the pentose phosphate pathway (PPP), glucose-6-phosphate dehydrogenase (G6PDH) provides nicotinamide adenine dinucleotide phosphate (NADPH) and intermediary metabolites for rubber biosynthesis, and plays an important role in plant development and stress responses. In this study, four Hevea brasiliensis (Para rubber tree) G6PDH genes (HbG6PDH1 to 4) were identified and cloned using a genome-wide scanning approach. All four HbG6PDH genes encode functional G6PDH enzymes as shown by heterologous expression in E. coli. Phylogeny analysis and subcellular localization prediction show that HbG6PDH3 is a cytosolic isoform, while the other three genes (HbG6PDH1, 2 and 4) are plastidic isoforms. The subcellular locations of HbG6PDH3 and 4, two latex-abundant isoforms were further verified by transient expression in rice protoplasts. Enzyme activity assay and expression analysis showed HbG6PDH3 and 4 were implicated in PPP during latex regeneration, and to influence rubber production positively in rubber tree. The cytosolic HbG6PDH3 is a predominant isoform in latex, implying a principal role for this isoform in controlling carbon flow and NADPH production in the PPP during latex regeneration. The expression pattern of plastidic HbG6PDH4 correlates well with the degree of tapping panel dryness, a physiological disorder that stops the flow of latex from affected rubber trees. In addition, the four HbG6PDHs responded to temperature and drought stresses in root, bark, and leaves, implicating their roles in maintaining redox balance and defending against oxidative stress.

  11. Heterologous expression of Spathaspora passalidarum xylose reductase and xylitol dehydrogenase genes improved xylose fermentation ability of Aureobasidium pullulans.

    Science.gov (United States)

    Guo, Jian; Huang, Siyao; Chen, Yefu; Guo, Xuewu; Xiao, Dongguang

    2018-04-30

    Aureobasidium pullulans is a yeast-like fungus that can ferment xylose to generate high-value-added products, such as pullulan, heavy oil, and melanin. The combinatorial expression of two xylose reductase (XR) genes and two xylitol dehydrogenase (XDH) genes from Spathaspora passalidarum and the heterologous expression of the Piromyces sp. xylose isomerase (XI) gene were induced in A. pullulans to increase the consumption capability of A. pullulans on xylose. The overexpression of XYL1.2 (encoding XR) and XYL2.2 (encoding XDH) was the most beneficial for xylose utilization, resulting in a 17.76% increase in consumed xylose compared with the parent strain, whereas the introduction of the Piromyces sp. XI pathway failed to enhance xylose utilization efficiency. Mutants with superior xylose fermentation performance exhibited increased intracellular reducing equivalents. The fermentation performance of all recombinant strains was not affected when glucose or sucrose was utilized as the carbon source. The strain with overexpression of XYL1.2 and XYL2.2 exhibited excellent fermentation performance with mimicked hydrolysate, and pullulan production increased by 97.72% compared with that of the parent strain. The present work indicates that the P4 mutant (using the XR/XDH pathway) with overexpressed XYL1.2 and XYL2.2 exhibited the best xylose fermentation performance. The P4 strain showed the highest intracellular reducing equivalents and XR and XDH activity, with consequently improved pullulan productivity and reduced melanin production. This valuable development in aerobic fermentation by the P4 strain may provide guidance for the biotransformation of xylose to high-value products by A. pullulans through genetic approach.

  12. Isoepoxydon dehydrogenase (idh) gene expression in relation to patulin production by Penicillium expansum under different temperature and atmosphere.

    Science.gov (United States)

    De Clercq, N; Vlaemynck, G; Van Pamel, E; Van Weyenberg, S; Herman, L; Devlieghere, F; De Meulenaer, B; Van Coillie, E

    2016-03-02

    Penicillium expansum growth and patulin production occur mainly at post-harvest stage during the long-term storage of apples. Low temperature in combination with reduced oxygen concentrations is commonly applied as a control strategy to extend apple shelf life and supply the market throughout the year. Our in vitro study investigated the effect of temperature and atmosphere on expression of the idh gene in relation to the patulin production by P. expansum. The idh gene encodes the isoepoxydon dehydrogenase enzyme, a key enzyme in the patulin biosynthesis pathway. First, a reverse transcription real-time PCR (RT-qPCR) method was optimized to measure accurately the P. expansum idh mRNA levels relative to the mRNA levels of three reference genes (18S, β-tubulin, calmodulin), taking into account important parameters such as PCR inhibition and multiple reference gene stability. Subsequently, two P. expansum field isolates and one reference strain were grown on apple puree agar medium (APAM) under three conditions of temperature and atmosphere: 20 °C - air, 4 °C - air and 4 °C - controlled atmosphere (CA; 3% O2). When P. expansum strains reached a 0.5 and 2.0 cm colony diameter, idh expression and patulin concentrations were determined by means of the developed RT-qPCR and an HPLC-UV method, respectively. The in vitro study showed a clear reduction in patulin production and down-regulation of the idh gene expression when P. expansum was grown under 4 °C - CA. The results suggest that stress (low temperature and oxygen level) caused a delay of the fungal metabolism rather than a complete inhibition of toxin biosynthesis. A good correlation was found between the idh expression and patulin production, corroborating that temperature and atmosphere affected patulin production by acting at the transcriptional level of the idh gene. Finally, a reliable RT-qPCR can be considered as an alternative tool to investigate the effect of control strategies on the toxin formation in

  13. Clonal diversity of the glutamate dehydrogenase gene in Giardia duodenalis from Thai Isolates: evidence of genetic exchange or Mixed Infections?

    Directory of Open Access Journals (Sweden)

    Saksirisampant Wilai

    2011-09-01

    Full Text Available Abstract Background The glutamate dehydrogenase gene (gdh is one of the most popular and useful genetic markers for the genotypic analysis of Giardia duodenalis (syn. G. lamblia, G. intestinalis, the protozoan that widely causes enteric disease in humans. To determine the distribution of genotypes of G. duodenalis in Thai populations and to investigate the extent of sequence variation at this locus, 42 fecal samples were collected from 3 regions of Thailand i.e., Central, Northern, and Eastern regions. All specimens were analyzed using PCR-based genotyping and recombinant subcloning methods. Results The results showed that the prevalence of assemblages A and B among these populations was approximately equal, 20 (47.6% and 22 (52.4%, respectively. Sequence analysis revealed that the nucleotide diversity of assemblage B was significantly greater than that in assemblage A. Among all assemblage B positive specimens, the allelic sequence divergence within isolates was detected. Nine isolates showed mixed alleles, ranged from three to nine distinct alleles per isolate. Statistical analysis demonstrated the occurrence of genetic recombination within subassemblages BIII and BIV was likely. Conclusion This study supports increasing evidence that G. duodenalis has the potential for genetic exchange.

  14. 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.

  15. Improved sugar-free succinate production by Synechocystis sp. PCC 6803 following identification of the limiting steps in glycogen catabolism

    Directory of Open Access Journals (Sweden)

    Tomohisa Hasunuma

    2016-12-01

    Full Text Available Succinate produced by microorganisms can replace currently used petroleum-based succinate but typically requires mono- or poly-saccharides as a feedstock. The cyanobacterium Synechocystis sp. PCC6803 can produce organic acids such as succinate from CO2 not supplemented with sugars under dark anoxic conditions using an unknown metabolic pathway. The TCA cycle in cyanobacteria branches into oxidative and reductive routes. Time-course analyses of the metabolome, transcriptome and metabolic turnover described here revealed dynamic changes in the metabolism of Synechocystis sp. PCC6803 cultivated under dark anoxic conditions, allowing identification of the carbon flow and rate-limiting steps in glycogen catabolism. Glycogen biosynthesized from CO2 assimilated during periods of light exposure is catabolized to succinate via glycolysis, the anaplerotic pathway, and the reductive TCA cycle under dark anoxic conditions. Expression of the phosphoenolpyruvate (PEP carboxylase gene (ppc was identified as a rate-limiting step in succinate biosynthesis and this rate limitation was alleviated by ppc overexpression, resulting in improved succinate excretion. The sugar-free succinate production was further enhanced by the addition of bicarbonate. In vivo labeling with NaH13CO3 clearly showed carbon incorporation into succinate via the anaplerotic pathway. Bicarbonate is in equilibrium with CO2. Succinate production by Synechocystis sp. PCC6803 therefore holds significant promise for CO2 capture and utilization. Keywords: Autofermentation, Cyanobacteria, Dynamic metabolic profiling, Metabolomics, Succinate, Synechocystis

  16. 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.

  17. Targeted Disruption of the kstD Gene Encoding a 3-Ketosteroid Δ1-Dehydrogenase Isoenzyme of Rhodococcus erythropolis Strain SQ1

    Science.gov (United States)

    van der Geize, R.; Hessels, G. I.; van Gerwen, R.; Vrijbloed, J. W.; van der Meijden, P.; Dijkhuizen, L.

    2000-01-01

    Microbial phytosterol degradation is accompanied by the formation of steroid pathway intermediates, which are potential precursors in the synthesis of bioactive steroids. Degradation of these steroid intermediates is initiated by Δ1-dehydrogenation of the steroid ring structure. Characterization of a 2.9-kb DNA fragment of Rhodococcus erythropolis SQ1 revealed an open reading frame (kstD) showing similarity with known 3-ketosteroid Δ1-dehydrogenase genes. Heterologous expression of kstD yielded 3-ketosteroid Δ1-dehydrogenase (KSTD) activity under the control of the lac promoter in Escherichia coli. Targeted disruption of the kstD gene in R. erythropolis SQ1 was achieved, resulting in loss of more than 99% of the KSTD activity. However, growth on the steroid substrate 4-androstene-3,17-dione or 9α-hydroxy-4-androstene-3,17-dione was not abolished by the kstD gene disruption. Bioconversion of phytosterols was also not blocked at the level of Δ1-dehydrogenation in the kstD mutant strain, since no accumulation of steroid pathway intermediates was observed. Thus, inactivation of kstD is not sufficient for inactivation of the Δ1-dehydrogenase activity. Native polyacrylamide gel electrophoresis of cell extracts stained for KSTD activity showed that R. erythropolis SQ1 in fact harbors two activity bands, one of which is absent in the kstD mutant strain. PMID:10788377

  18. IRE1 KNOCKDOWN MODIFIES THE GLUTAMINE AND GLUCOSE DEPRIVATION EFFECT ON THE EXPRESSION OF NUCLEAR GENES ENCODING MITOCHONDRIAL PROTEINS IN U87 GLIOMA CELLS

    Directory of Open Access Journals (Sweden)

    O. O.

    2016-04-01

    Full Text Available We have studied the glucose and glutamine deprivation effect on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells in relation to inhibition of inositol requiring enzyme-1 (IRE1. It was shown that glutamine deprivation down-regulated the expression of mitochondrial (NADP+-dependent isocitrate dehydrogenase 2 (IDH2, malic enzyme 2 (ME2, mitochondrial aspartate aminotransferase (GOT2, and subunit B of succinate dehydrogenase (SDHB genes in control glioma cells in gene specific manner. At the same time, the expression level of malate dehydrogenase 2 (MDH2 and subunit D of succinate dehydrogenase (SDHD genes in these cells was not changed upon glutamine deprivation. It was also shown that inhibition of ІRE1 signaling enzyme function in U87 glioma cells modified the glutamine deprivation effect on the expression of all studied genes. Furthermore, the expression of the majority of studied genes was resistant to glucose deprivation, except IDH2 and SDHB genes, which expression levels were slightly down-regulated. Inhibition of IRE1 modified the effect of glucose deprivation on ME2, SDHB, SDHD, and GOT2 genes expression. Therefore, glucose and glutamine deprivation affected the expression level of the majority of nuclear genes encoding mitochondrial proteins in relation to the functional activity of IRE1 enzyme, which is a central mediator of endoplasmic reticulum stress and controls cell proliferation and tumor growth.

  19. 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

  20. Poly(Butylene Succinate) and Poly[(Butylene Succinate)-co-Adipate] Nanocomposites

    CSIR Research Space (South Africa)

    Ojijo, Vincent O

    2012-01-01

    Full Text Available , and properties of nanoclay-containing composites of biodegradable poly(butylene succinate) (PBS) and poly[(butylene succinate)-co-adipate] (PBSA). Various nanocomposite structures arising from the incorporation of layered silicate particles, both pristine...

  1. Activite succino-deshydrogenasique des microsomes et mode d'incorporation du succinate 2,3-(14)C dans les acides gras des microsomes de foie de rat in vitro.

    Science.gov (United States)

    Rous, S; Aubry, L; Bonini, F

    1970-03-16

    The incorporations of 2,3-(14)C-succinate 2-(14)C-acetate into fatty acids of different cellular fractions of rat liver were studied. Acetate was incorporated mainly into supernatant and succinate into microsomal fatty acids. Mitochondria only could intensively decarboxylate pyruvate. Avidine inhibited fatty acid synthesis from succinate mainly in the supernatant. It is suggested that succinate is an important physiological precursor of fatty acids in the liver and that an active succino-dehydrogenase is present in microsomes.

  2. Markerless deletion of putative alanine dehydrogenase genes in Bacillus licheniformis using a codBA-based counterselection technique.

    Science.gov (United States)

    Kostner, David; Rachinger, Michael; Liebl, Wolfgang; Ehrenreich, Armin

    2017-11-01

    Bacillus licheniformis strains are used for the large-scale production of industrial exoenzymes from proteinaceous substrates, but details of the amino acid metabolism involved are largely unknown. In this study, two chromosomal genes putatively involved in amino acid metabolism of B. licheniformis were deleted to clarify their role. For this, a convenient counterselection system for markerless in-frame deletions was developed for B. licheniformis. A deletion plasmid containing up- and downstream DNA segments of the chromosomal deletion target was conjugated to B. licheniformis and integrated into the genome by homologous recombination. Thereafter, the counterselection was done by using a codBA cassette. The presence of cytosine deaminase and cytosine permease exerted a conditionally lethal phenotype on B. licheniformis cells in the presence of the cytosine analogue 5-fluorocytosine. Thereby clones were selected that lost the integrated vector sequence and the anticipated deletion target after a second recombination step. This method allows the construction of markerless mutants in Bacillus strains in iterative cycles. B. licheniformis MW3 derivatives lacking either one of the ORFs BL03009 or BL00190, encoding a putative alanine dehydrogenase and a similar putative enzyme, respectively, retained the ability to grow in minimal medium supplemented with alanine as the carbon source. In the double deletion mutant MW3 ΔBL03009 ΔBL00190, however, growth on alanine was completely abolished. These data indicate that the two encoded enzymes are paralogues fulfilling mutually replaceable functions in alanine utilization, and suggest that in B. licheniformis MW3 alanine utilization is initiated by direct oxidative transamination to pyruvate and ammonium.

  3. 21 CFR 184.1091 - Succinic acid.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Succinic acid. 184.1091 Section 184.1091 Food and... Substances Affirmed as GRAS § 184.1091 Succinic acid. (a) Succinic acid (C4H6O4, CAS Reg. No. 110-15-6), also referred to as amber acid and ethylenesuccinic acid, is the chemical 1,4-butanedioic acid. It is...

  4. 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.

  5. Functional assignment of gene AAC16202.1 from Rhodobacter capsulatus SB1003: new insights into the bacterial SDR sorbitol dehydrogenases family.

    Science.gov (United States)

    Sola-Carvajal, Agustín; García-García, María Inmaculada; Sánchez-Carrón, Guiomar; García-Carmona, Francisco; Sánchez-Ferrer, Alvaro

    2012-11-01

    Short-chain dehydrogenases/reductases (SDR) constitute one of the largest enzyme superfamilies with over 60,000 non-redundant sequences in the database, many of which need a correct functional assignment. Among them, the gene AAC16202.1 (NCBI) from Rhodobacter capsulatus SB1003 has been assigned in Uniprot both as a sorbitol dehydrogenase (#D5AUY1) and, as an N-acetyl-d-mannosamine dehydrogenase (#O66112), both enzymes being of biotechnological interest. When the gene was overexpressed in Escherichia coli Rosetta (DE3)pLys, the purified enzyme was not active toward N-acetyl-d-mannosamine, whereas it was active toward d-sorbitol and d-fructose. However, the relative activities toward xylitol and l-iditol (0.45 and 6.9%, respectively) were low compared with that toward d-sorbitol. Thus, the enzyme could be considered sorbitol dehydrogenase (SDH) with very low activity toward xylitol, which could increase its biotechnological interest for determining sorbitol without the unspecific cross-determination of added xylitol in food and pharma compositions. The tetrameric enzyme (120 kDa) showed similar catalytic efficiency (2.2 × 10(3) M(-1) s(-1)) to other sorbitol dehydrogenases for d-sorbitol, with an optimum pH of 9.0 and an optimum temperature of 37 °C. The enzyme was also more thermostable than other reported SDH, ammonium sulfate being the best stabilizer in this respect, increasing the melting temperature (T(m)) up to 52.9 °C. The enzyme can also be considered as a new member of the Zn(2+) independent SDH family since no effect on activity was detected in the presence of divalent cations or chelating agents. Finally, its in silico analysis enabled the specific conserved sequence blocks that are the fingerprints of bacterial sorbitol dehydrogenases and mainly located at C-terminal of the protein, to be determined for the first time. This knowledge will facilitate future data curation of present databases and a better functional assignment of newly described

  6. Functionality of allelic variations in human alcohol dehydrogenase gene family: assessment of a functional window for protection against alcoholism.

    Science.gov (United States)

    Lee, Shou-Lun; Höög, Jan-Olov; Yin, Shih-Jiun

    2004-11-01

    Alcohol dehydrogenase (ADH) catalyses the rate-determining reaction in ethanol metabolism. Genetic association studies of diverse ethnic groups have firmly demonstrated that the allelic variant ADH1B*2 significantly protects against alcoholism but that ADH1C*1, which is in linkage with ADH1B*2, produces a negligible protection. The influence of other potential candidate genes/alleles within the human ADH family, ADH1B*3 and ADH2, remains unclear or controversial. To address this question, functionalities of ADH1B3 and ADH2 were assessed at a physiological level of coenzyme and substrate range. Ethanol-oxidizing activities of recombinant ADH1B1, ADH1B2, ADH1B3, ADH1C1, ADH1C2 and ADH2 were determined at pH 7.5 in the presence of 0.5 mm NAD with 2-50 mm ethanol. The activity differences between ADH1B2 and ADH1B1 were taken as a threshold for effective protection against alcoholism and those between ADH1C1 and ADH1C2 as a threshold for null protection. Over 2-50 mm ethanol, the activities of ADH1B3 were found 2.9-23-fold lower than those of ADH1B2, largely attributed to the Km effect (ADH1B2, 1.8 mm; ADH1B3, 61 mm). Strikingly, the ADH1B3 activity was only 84% that of ADH1B1 at a low ethanol concentration, 2 mm, but increased 10-fold at 50 mm. Corrected for relative expression levels of the enzyme in liver, the hepatic ADH2 activities were estimated to be 18-97% those of ADH1B1 over 2-50 mm ethanol and were 28-140% of the activity differences between ADH1C1 and ADH1C2. The assessment based on the proposed functional window for the human ADH gene family indicates that ADH1B*3 may show some degree of protection against alcoholism and that the ADH2 functional variants appear to be negligible for this protection.

  7. Acyl-CoA reductase PGN_0723 utilizes succinyl-CoA to generate succinate semialdehyde in a butyrate-producing pathway of Porphyromonas gingivalis.

    Science.gov (United States)

    Yoshida, Yasuo; Sato, Mitsunari; Kezuka, Yuichiro; Hasegawa, Yoshiaki; Nagano, Keiji; Takebe, Jun; Yoshimura, Fuminobu

    2016-04-15

    The molecular basis of butyrate production in Porphyromonas gingivalis has not been fully elucidated, even though butyrate, a short chain fatty acid (SCFA), can exert both beneficial and harmful effects on a mammalian host. A database search showed that the amino acid sequence of PGN_0723 protein was 50.6% identical with CoA-dependent succinate semialdehyde dehydrogenase (SSADH) in Clostridium kluyveri. By contrast, the protein has limited identity (19.1%) with CoA-independent SSADH in Escherichia coli. Compared with the wild type, growth speed, and final turbidity were lower in the PGN_0723 deletion strain that was constructed by replacing the PGN_0723 gene with an erythromycin resistance cassette. Gas chromatography mass spectrometry revealed the supernatant concentrations of the SCFAs butyrate, isobutyrate, and isovalerate, but not propionate, in the PGN_0723 deletion strain were also lower than those in the wild type. The wild-type phenotype was restored in a complemented strain. We cloned the PGN_0723 gene, purified the recombinant protein, and computationally constructed its three-dimensional model. A colorimetric assay and liquid chromatography-tandem mass spectrometry analysis demonstrated that the recombinant PGN_0723 produces succinate semialdehyde, which is an intermediate in the P. gingivalis butyrate synthesis pathway, not from succinate but from succinyl-CoA in the presence of NAD(P)H via a ping-pong bi-bi mechanism. Asn110Ala and Cys239Ala mutations resulted in a significant loss of the CoA-dependent PGN_0723 enzymatic activity. The study provides new insights into butyrate production, which constitutes a virulence factor in P. gingivalis. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. Improved sugar-free succinate production bySynechocystissp. PCC 6803 following identification of the limiting steps in glycogen catabolism.

    Science.gov (United States)

    Hasunuma, Tomohisa; Matsuda, Mami; Kondo, Akihiko

    2016-12-01

    Succinate produced by microorganisms can replace currently used petroleum-based succinate but typically requires mono- or poly-saccharides as a feedstock. The cyanobacterium Synechocystis sp. PCC6803 can produce organic acids such as succinate from CO 2 not supplemented with sugars under dark anoxic conditions using an unknown metabolic pathway. The TCA cycle in cyanobacteria branches into oxidative and reductive routes. Time-course analyses of the metabolome, transcriptome and metabolic turnover described here revealed dynamic changes in the metabolism of Synechocystis sp. PCC6803 cultivated under dark anoxic conditions, allowing identification of the carbon flow and rate-limiting steps in glycogen catabolism. Glycogen biosynthesized from CO 2 assimilated during periods of light exposure is catabolized to succinate via glycolysis, the anaplerotic pathway, and the reductive TCA cycle under dark anoxic conditions. Expression of the phospho enol pyruvate (PEP) carboxylase gene ( ppc ) was identified as a rate-limiting step in succinate biosynthesis and this rate limitation was alleviated by ppc overexpression, resulting in improved succinate excretion. The sugar-free succinate production was further enhanced by the addition of bicarbonate. In vivo labeling with NaH 13 CO 3 clearly showed carbon incorporation into succinate via the anaplerotic pathway. Bicarbonate is in equilibrium with CO 2 . Succinate production by Synechocystis sp. PCC6803 therefore holds significant promise for CO 2 capture and utilization.

  9. The effect of short/branched chain acyl-coenzyme A dehydrogenase gene on triglyceride synthesis of bovine mammary epithelial cells

    Directory of Open Access Journals (Sweden)

    P. Jiang

    2018-03-01

    Full Text Available Short/branched chain acyl-CoA dehydrogenase (ACADSB is a member of the acyl-CoA dehydrogenase family of enzymes that catalyze the dehydrogenation of acyl-CoA derivatives in the metabolism of fatty acids. Our previous transcriptome analysis in dairy cattle showed that ACADSB was differentially expressed and was associated with milk fat metabolism. The aim of this study was to elucidate the background of this differential expression and to evaluate the role of ACADSB as a candidate for fat metabolism in dairy cattle. After analysis of ACADSB mRNA abundance by qRT-PCR and Western blot, overexpression and RNA interference (RNAi vectors of ACADSB gene were constructed and then transfected into bovine mammary epithelial cells (bMECs to examine the effects of ACADSB on milk fat synthesis. The results showed that the ACADSB was differentially expressed in mammary tissue of low and high milk fat dairy cattle. Overexpression of ACADSB gene could significantly increase the level of intracellular triglyceride (TG, while ACADSB gene knockdown could significantly reduce the TG synthesis in bMECs. This study suggested that the ACADSB was important in TG synthesis in bMECs, and it could be a candidate gene to regulate the metabolism of milk fat in dairy cattle.

  10. Succinic Acid: Technology Development and Commercialization

    Directory of Open Access Journals (Sweden)

    Nhuan P. Nghiem

    2017-06-01

    Full Text Available Succinic acid is a precursor of many important, large-volume industrial chemicals and consumer products. It was once common knowledge that many ruminant microorganisms accumulated succinic acid under anaerobic conditions. However, it was not until the discovery of Anaerobiospirillum succiniciproducens at the Michigan Biotechnology Institute (MBI, which was capable of producing succinic acid up to about 50 g/L under optimum conditions, that the commercial feasibility of producing the compound by biological processes was realized. Other microbial strains capable of producing succinic acid to high final concentrations subsequently were isolated and engineered, followed by development of fermentation processes for their uses. Processes for recovery and purification of succinic acid from fermentation broths were simultaneously established along with new applications of succinic acid, e.g., production of biodegradable deicing compounds and solvents. Several technologies for the fermentation-based production of succinic acid and the subsequent conversion to useful products are currently commercialized. This review gives a summary of the development of microbial strains, their fermentation, and the importance of the down-stream recovery and purification efforts to suit various applications in the context of their current commercialization status for biologically derived succinic acid.

  11. Expression and characterization of a class III alcohol dehydrogenase gene from Gluconobacter frateurii in the presence of methanol during glyceric acid production from glycerol.

    Science.gov (United States)

    Sato, Shun; Morita, Naoki; Kitamoto, Dai; Habe, Hiroshi

    2013-01-01

    Some acetic acid bacteria have been shown to produce large amounts of glyceric acid (GA) from glycerol, which is a by-product of biodiesel fuel (BDF) production. Previously, a Gluconobacter strain was found that produced decreased amounts of GA from glycerol in the presence of methanol, a major ingredient of raw glycerol derived from the BDF industry. Thus, a comparative transcriptome analysis of Gluconobacter frateurii NBRC103465 was performed to investigate changes in gene expression during GA production from glycerol in the presence of methanol. Cells grown with methanol showed upregulated expression of a class III alcohol dehydrogenase homolog (adhC(Gf)) and decreased GA production. adhC(Gf) was cloned and expressed heterologously in Escherichia coli, and the presence of an additional protein with an approximate molecular mass of 39 kDa in the cytosol of the recombinant E. coli cells was identified by SDS-PAGE. Activity measurements of the cytosol revealed that the translational product of adhC(Gf) exhibited formaldehyde dehydrogenase activity in the presence of nicotinamide adenine dinucleotide and glutathione. Gluconobacter frateurii cells grown in 1% methanol-containing glycerol were found to have fivefold higher formaldehyde dehydrogenase activity than cells grown without methanol, suggesting that adhC(Gf) in G. frateurii cells functions in the dissimilation of methanol-derived formaldehyde.

  12. Probing cytokinin homeostasis in Arabidopsis thaliana by constitutively overexpressing two forms of the maize cytokinin oxidase/dehydrogenase 1 gene

    Czech Academy of Sciences Publication Activity Database

    Kopečný, D.; Tarkowski, Petr; Majira, M.; Bouchez-Mahiout, I.; Nogué, F.; Laurière, M.; Sandberg, G.; Laloue, M.; Houba-Hérin, N.

    2006-01-01

    Roč. 171, č. 1 (2006), s. 114-122 ISSN 0168-9452 Institutional research plan: CEZ:AV0Z50380511 Keywords : Arabidopsis thaliana * Cytokinin oxidase/dehydrogenase * Homeostasis Subject RIV: CE - Biochemistry Impact factor: 1.631, year: 2006

  13. Role of common gene variations in the molecular pathogenesis of short-chain acyl-CoA dehydrogenase deficiency.

    NARCIS (Netherlands)

    Corydon, M.J.; Vockley, J.; Rinaldo, P.; Rhead, W.J.; Kjeldsen, M.; Winter, V.; Riggs, C.; Babovic-Vuksanovic, D.; Smeitink, J.A.M.; Jong, J. de; Levy, H.L.; Sewell, A.C.; Roe, C.; Matern, D.; Dasouki, M.; Gregersen, N.

    2001-01-01

    ABSTRACT Short-chain acyl-CoA dehydrogenase (SCAD) deficiency is considered a rare inherited mitochondrial fatty acid oxidation disorder. Less than 10 patients have been reported, diagnosed on the basis of ethylmalonic aciduria and low SCAD activity in cultured fibroblast. However, mild ethylmalonic

  14. Human polymorphism in drug metabolism: mutation in the dihydropyrimidine dehydrogenase gene results in exon skipping and thymine uracilurea

    NARCIS (Netherlands)

    Meinsma, R.; Fernandez-Salguero, P.; van Kuilenburg, A. B.; van Gennip, A. H.; Gonzalez, F. J.

    1995-01-01

    A condition called thymine uracilurea has been described that is due to a lack of dihydropyrimidine dehydrogenase (DPD) activity. Cancer patients experiencing acute 5-fluorouracil toxicity also have lower-than-normal DPD activities. However, to date, the molecular basis of this disorder has not been

  15. Production of Succinic Acid for Lignocellulosic Hydrolysates

    Energy Technology Data Exchange (ETDEWEB)

    Davison, B.H.; Nghiem, J.

    2002-06-01

    The purpose of this Cooperative Research and Development Agreement (CRADA) is to add and test new metabolic activities to existing microbial catalysts for the production of succinic acid from renewables. In particular, they seek to add to the existing organism the ability to utilize xylose efficiently and simultaneously with glucose in mixtures of sugars or to add succinic acid production to another strain and to test the value of this new capability for production of succinic acid from industrial lignocellulosic hydrolyasates. The Contractors and Participant are hereinafter jointly referred to as the 'Parties'. Research to date in succinic acid fermentation, separation and genetic engineering has resulted in a potentially economical process based on the use of an Escherichia coli strain AFP111 with suitable characteristics for the production of succinic acid from glucose. Economic analysis has shown that higher value commodity chemicals can be economically produced from succinic acid based on repliminary laboratory findings and predicted catalytic parameters. The initial target markets include succinic acid itself, succinate salts, esters and other derivatives for use as deicers, solvents and acidulants. The other commodity products from the succinic acid platform include 1,4-butanediol, {gamma}-butyrolactone, 2-pyrrolidinone and N-methyl pyrrolidinone. Current economic analyses indicate that this platform is competitive with existing petrochemical routes, especially for the succinic acid and derivatives. The report presents the planned CRADA objectives followed by the results. The results section has a combined biocatalysis and fermentation section and a commercialization section. This is a nonproprietary report; additional proprietary information may be made available subject to acceptance of the appropriate proprietary information agreements.

  16. Cloning, sequencing, and characterization of the gene encoding the smallest subunit of the three-component membrane-bound alcohol dehydrogenase from Acetobacter pasteurianus.

    Science.gov (United States)

    Kondo, K; Beppu, T; Horinouchi, S

    1995-09-01

    The membrane-bound alcohol dehydrogenase (ADH) of Acetobacter pasteurianus NCI1452 consists of three different subunits, a 78-kDa dehydrogenase subunit, a 48-kDa cytochrome c subunit, and a 20-kDa subunit of unknown function. For elucidation of the function of the smallest subunit, this gene was cloned from this strain by the oligonucleotide-probing method, and its nucleotide sequence was determined. Comparison of the deduced amino acid sequence and the NH2-terminal sequence determined for the purified protein indicated that the smallest subunit contained a typical signal peptide of 28 amino acids, as did the larger two subunits. This gene complemented the ADH activity of a mutant strain which had lost the smallest subunit. Disruption of this gene on the chromosome resulted in loss of ADH activity in Acetobacter aceti, indicating that the smallest subunit was essential for ADH activity. Immunoblot analyses of cell lysates prepared from various ADH mutants suggested that the smallest subunit was concerned with the stability of the 78-kDa subunit and functioned as a molecular coupler of the 78-kDa subunit to the 48-kDa subunit on the cytoplasmic membrane.

  17. Chemoreactome analysis of ethylmethylhydroxypyridine succinate

    Directory of Open Access Journals (Sweden)

    O. A. Gromova

    2016-01-01

    Full Text Available Ethylmethylhydroxypyridine succinate (EMHPS is used in the therapy of ischemic stroke. A more complete understanding of the conditions that can affect the clinical efficacy of EMHPS needs the most complete information about its molecular mechanisms of action.Objective: to comparatively analyze the properties of EMHPS using the newest area of postgenomic pharmacology – chemoreactome simulation. Succinic acid and citicoline were used as the molecules of comparison (control molecules.Material and methods. Chemoreactome analysis was employed to assess the biological activity of a test molecule (simulation of the affinity profile of the examined molecular structure for various proteome proteins. A new mathematical method based on the combinatorial theory of solvability was devised for chemoinformational analysis.Results and discussion. Chemoreactome simulation has shown that EMHPS may be an agonist of acetylcholine and GABA receptors, as well as that of cannabinoid receptor type 2. The anti-inflammatory effect of an EMHPS molecule may be due to the inhibition of synthesis of proinflammatory prostaglandins. Its higher safety (a weaker effect on serotonin and opioid receptors and lower interaction with Erg channels and the enzymes monoamine oxidase and cytochrome CYP1A1 distinguish EMHPS from the molecules of comparison (citicolne and succinic acid. The predicted properties of the molecule of EMHPS make a substantial contribution to its neuroprotective effect.Conclusion. The results of chemoreactome analysis could reveal not only quite unexpected mechanisms of action of EMHPS, but also its mechanisms of synergic action with pyridoxine and magnesium. Owing to a combination of EMHPS with pyridoxine and magnesium, there is an increase in the antioxidant, anticonvulsant, stress- and neuroprotective, nootropic, and anxiolytic effects of EMHPS via activation of pyridoxine-dependent and magnesium-dependent proteins. The combination of EMHPS with

  18. Succinate-induced neuronal mitochondrial fission and hexokinase II malfunction in ischemic stroke: Therapeutical effects of kaempferol.

    Science.gov (United States)

    Wu, Bin; Luo, Hong; Zhou, Xu; Cheng, Cai-Yi; Lin, Lin; Liu, Bao-Lin; Liu, Kang; Li, Ping; Yang, Hua

    2017-09-01

    Mitochondrial dysfunction is known as one of causative factors in ischemic stroke, leading to neuronal cell death. The present work was undertaken to investigate whether succinate induces neuron apoptosis by regulating mitochondrial morphology and function. In neurons, oxygen-glucose deprivation induced succinate accumulation due to the reversal of succinate dehydrogenase (SDH) activation, leading to mitochondrial fission. Kaempferol inhibited mitochondrial fission and maintained mitochondrial HK-II through activation of Akt, and thereby protected neurons from succinate-mediated ischemi injury. Knockdown of Akt2 with siRNA diminished the effect of kaempferol, indicating that kaempferol suppressed dynamin-related protein 1 (Drp1) activation and promoted HK-II mitochondrial binding dependently on Akt. Moreover, we demonstrated that kaempferol potentiated autophagy during oxygen and glucose deprivation, contributing to protecting neuron survival against succinate insult. In vivo, oral administration of kaempferol in mice attenuated the infract volume after ischemic and reperfusion (I/R) injury and reproduced the similar mitochondrial protective effect in the brain infract area. This study indicates that succinate accumulation plays a pivotal role in I/R injury-induced neuronal mitochondrial dysfunction, and suggests that modulation of Drp1 phosphorylation might be potential therapeutic strategy to protect neuron mitochondrial integrity and treat ischemic stroke. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Regulation of succinate-fuelled mitochondrial respiration in liver and skeletal muscle of hibernating thirteen-lined ground squirrels.

    Science.gov (United States)

    Brown, Jason C L; Chung, Dillon J; Cooper, Alex N; Staples, James F

    2013-05-01

    Hibernating ground squirrels (Ictidomys tridecemlineatus) alternate between two distinct metabolic states throughout winter: torpor, during which metabolic rate (MR) and body temperature (Tb) are considerably suppressed, and interbout euthermia (IBE), during which MR and Tb briefly return to euthermic levels. Previous studies showed suppression of succinate-fuelled respiration during torpor in liver and skeletal muscle mitochondria; however, these studies used only a single, saturating succinate concentration. Therefore, they could not address whether mitochondrial metabolic suppression occurs under physiological substrate concentrations or whether differences in the kinetics of mitochondrial responses to changing substrate concentration might also contribute to mitochondrial metabolic regulation during torpor. The present study confirmed that succinate oxidation is reduced during torpor in liver and skeletal muscle at 37 and 10°C over a 100-fold range of succinate concentrations. At 37°C, this suppression resulted from inhibition of succinate dehydrogenase (SDH), which had a greater affinity for oxaloacetate (an SDH inhibitor) during torpor. At 10°C, SDH was not inhibited, suggesting that SDH inhibition initiates but does not maintain mitochondrial suppression during torpor. Moreover, in both liver and skeletal muscle, mitochondria from torpid animals maintained relatively higher respiration rates at low succinate concentrations, which reduces the extent of energy savings that can be achieved during torpor, but may also maintain mitochondrial oxidative capacity above some lower critical threshold, thereby preventing cellular and/or mitochondrial injury during torpor and facilitating rapid recruitment of oxidative capacity during arousal.

  20. Restoring of Glucose Metabolism of Engineered Yarrowia lipolytica for Succinic Acid Production via a Simple and Efficient Adaptive Evolution Strategy.

    Science.gov (United States)

    Yang, Xiaofeng; Wang, Huaimin; Li, Chong; Lin, Carol Sze Ki

    2017-05-24

    Succinate dehydrogenase inactivation in Yarrowia lipolytica has been demonstrated for robust succinic acid production, whereas the inefficient glucose metabolism has hindered its practical application. In this study, a simple and efficient adaptive evolution strategy via cell immobilization was conducted in shake flasks, with an aim to restore the glucose metabolism of Y. lipolytica mutant PGC01003. After 21 days with 14 generations evolution, glucose consumption rate increased to 0.30 g/L/h in YPD medium consisting of 150 g/L initial glucose concentration, while poor yeast growth was observed in the same medium using the initial strain without adaptive evolution. Succinic acid productivity of the evolved strain also increased by 2.3-fold, with stable cell growth in YPD medium with high initial glucose concentration. Batch fermentations resulted in final succinic acid concentrations of 65.7 g/L and 87.9 g/L succinic acid using YPD medium and food waste hydrolysate, respectively. The experimental results in this study show that a simple and efficient strategy could facilitate the glucose uptake rate in succinic acid fermentation using glucose-rich substrates.

  1. Cloning and characterization of human very-long-chain acyl-CoA dehydrogenase cDNA, chromosomal assignment of the gene and identification in four patients of nine different mutations within the VLCAD gene

    DEFF Research Database (Denmark)

    Andresen, B S; Bross, P; Vianey-Saban, C

    1996-01-01

    Very-long-chain acyl-CoA dehydrogenase (VLCAD) is one of four straight-chain acyl-CoA dehydrogenase (ACD) enzymes, which are all nuclear encoded mitochondrial flavoproteins catalyzing the initial step in fatty acid beta-oxidation. We have used the very fast, Rapid Amplification of cDNA Ends (RACE......) based strategy to obtain the sequence of cDNAs encoding human VLCAD from placenta and fibroblasts. Alignment of the predicted amino acid sequence of human VLCAD with those of the other human ACD enzymes revealed extensive sequence homology. Moreover, human VLCAD and human acyl-CoA oxidase showed...... extensive sequence homology corroborating the notion that these genes are evolutionarily related. Southern blot analysis of genomic DNA from hybrid cell lines was used to localize the VLCAD gene to human chromosome 17p11.2-p11.13105. Using Northern and Western blot analysis to investigate the tissue...

  2. Construction of a genomic library of the food spoilage yeast Zygosaccharomyces bailii and isolation of the beta-isopropylmalate dehydrogenase gene (ZbLEU2).

    Science.gov (United States)

    Rodrigues, F; Zeeman, A M; Alves, C; Sousa, M J; Steensma, H Y; Côrte-Real, M; Leão, C

    2001-04-01

    A genomic library of the yeast Zygosaccharomyces bailii ISA 1307 was constructed in pRS316, a shuttle vector for Saccharomyces cerevisiae and Escherichia coli. The library has an average insert size of 6 kb and covers the genome more than 20 times assuming a genome size similar to that of S. cerevisiae. This new tool has been successfully used, by us and others, to isolate Z. bailii genes. One example is the beta-isopropylmalate dehydrogenase gene (ZbLEU2) of Z. bailii, which was cloned by complementation of a leu2 mutation in S. cerevisiae. An open reading frame encoding a protein with a molecular mass of 38.7 kDa was found. The nucleotide sequence of ZbLEU2 and the deduced amino acid sequence showed a significant degree of identity to those of beta-isopropylmalate dehydrogenases from several other yeast species. The sequence of ZbLEU2 has been deposited in the EMBL data library under accession number AJ292544.

  3. Production of natural fragrance aromatic acids by coexpression of trans-anethole oxygenase and p-anisaldehyde dehydrogenase genes of Pseudomonas putida JYR-1 in Escherichia coli.

    Science.gov (United States)

    Han, Dongfei; Kurusarttra, Somwang; Ryu, Ji-Young; Kanaly, Robert A; Hur, Hor-Gil

    2012-12-05

    A gene encoding p-anisaldehyde dehydrogenase (PAADH), which catalyzes the oxidation of p-anisaldehyde to p-anisic acid, was identified to be clustered with the trans-anethole oxygenase (tao) gene in Pseudomonas putida JYR-1. Heterologously expressed PAADH in Escherichia coli catalyzed the oxidation of vanillin, veratraldehyde, and piperonal to the corresponding aromatic acids vanillic acid, veratric acid, and piperonylic acid, respectively. Coexpression of trans-anethole oxygenase (TAO) and PAADH in E. coli also resulted in the successful transformation of trans-anethole, isoeugenol, O-methyl isoeugenol, and isosafrole to p-anisic acid, vanillic acid, veratric acid, and piperonylic acid, respectively, which are compounds found in plants as secondary metabolites. Because of the relaxed substrate specificity and high transformation rates by coexpressed TAO and PAADH in E. coli , the engineered strain has potential to be applied in the fragrance industry.

  4. Recurrent vomiting and ethylmalonic aciduria associated with rare mutations in the short-chain acyl-CoA dehydrogenase (SCAD) gene

    DEFF Research Database (Denmark)

    Seidel, J.; Streck, S.; Bellstedt, K.

    2003-01-01

    We report identification of short-chain acyl-CoA dehydrogenase (SCAD) deficiency in a 12-year-old boy who suffered from recurrent attacks of vomiting once or twice a year from infancy. Growth and development were normal and there were no muscular symptoms. Metabolic screening was performed during...... blood spots. Neither of the frequent SCAD gene variants 625G>A and 511C>T was present, but direct sequencing of the promoter and coding regions of the SCAD gene revealed that the patient had mutations on both alleles: 417G>C (Trpl15Cys) and 1095G>T (Gln341His). Neither mutation has been described before...... in compound heterozygosity or homozygosity. Enzymatic investigations subsequently confirmed a defect of SCAD in both fibroblasts and muscle extracts. Furthermore, expression studies of both mutations demonstrated impaired enzyme function or structure. To our knowledge, this case is the first description...

  5. 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.

  6. Disease-causing mutations in exon 11 of the medium-chain acyl-CoA dehydrogenase gene

    DEFF Research Database (Denmark)

    Andresen, B S; Jensen, T G; Bross, P

    1994-01-01

    Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most commonly recognized defect of the mitochondrial beta-oxidation in humans. It is a potentially fatal, autosomal recessive inherited defect. Most patients with MCAD deficiency are homozygous for a single disease-causing mutation (G985......), causing a change from lysine to glutamate at position 304 (K304E) in the mature MCAD. Only seven non-G985 mutations, all of which are rare, have been reported. Because the G985 mutation and three of the non-G985 mutations are located in exon 11, it has been suggested that this exon may be a mutational hot...

  7. Gene expression patterns in the hippocampus during the development and aging of Glud1 (Glutamate Dehydrogenase 1) transgenic and wild type mice.

    Science.gov (United States)

    Wang, Xinkun; Patel, Nilam D; Hui, Dongwei; Pal, Ranu; Hafez, Mohamed M; Sayed-Ahmed, Mohamed M; Al-Yahya, Abdulaziz A; Michaelis, Elias K

    2014-03-04

    Extraneuronal levels of the neurotransmitter glutamate in brain rise during aging. This is thought to lead to synaptic dysfunction and neuronal injury or death. To study the effects of glutamate hyperactivity in brain, we created transgenic (Tg) mice in which the gene for glutamate dehydrogenase (Glud1) is over-expressed in neurons and in which such overexpression leads to excess synaptic release of glutamate. In this study, we analyzed whole genome expression in the hippocampus, a region important for learning and memory, of 10 day to 20 month old Glud1 and wild type (wt) mice. During development, maturation and aging, both Tg and wt exhibited decreases in the expression of genes related to neurogenesis, neuronal migration, growth, and process elongation, and increases in genes related to neuro-inflammation, voltage-gated channel activity, and regulation of synaptic transmission. Categories of genes that were differentially expressed in Tg vs. wt during development were: synaptic function, cytoskeleton, protein ubiquitination, and mitochondria; and, those differentially expressed during aging were: synaptic function, vesicle transport, calcium signaling, protein kinase activity, cytoskeleton, neuron projection, mitochondria, and protein ubiquitination. Overall, the effects of Glud1 overexpression on the hippocampus transcriptome were greater in the mature and aged than the young. Glutamate hyperactivity caused gene expression changes in the hippocampus at all ages. Some of these changes may result in premature brain aging. The identification of these genomic expression differences is important in understanding the effects of glutamate dysregulation on neuronal function during aging or in neurodegenerative diseases.

  8. d-Lactate Dehydrogenase Gene (ldhD) Inactivation and Resulting Metabolic Effects in the Lactobacillus johnsonii Strains La1 and N312

    Science.gov (United States)

    Lapierre, Luciane; Germond, Jacques-Edouard; Ott, Andreas; Delley, Michele; Mollet, Beat

    1999-01-01

    Lactobacillus johnsonii La1, a probiotic bacterium with demonstrated health effects, grows in milk, where it ferments lactose to d- and l-lactate in a 60:40% ratio. The d-lactate dehydrogenase (D-LDH) gene (ldhD) of this strain was isolated, and an in vitro-truncated copy of that gene was used to inactivate the genomic copy in two strains, La1 and N312, by gene replacement. For that, an 8-bp deletion was generated within the cloned ldhD gene to inactivate its function. The plasmid containing the altered ldhD was transferred to L. johnsonii via conjugative comobilization with Lactococcus lactis carrying pAMβ1. Crossover integrations of the plasmid at the genomic ldhD site were selected, and appropriate resolution of the cointegrate structures resulted in mutants that had lost the plasmid and in which the original ldhD was replaced by the truncated copy. These mutants completely lacked D-LDH activity. Nevertheless, the lower remaining L-LDH activity of the cells was sufficient to reroute most of the accumulating pyruvate to l-lactate. Only a marginal increase in production of the secondary end products acetaldehyde, diacetyl, and acetoin was observed. It can be concluded that in L. johnsonii D- and L-LDH are present in substantial excess for their role to eliminate pyruvate and regenerate NAD+ and that accumulated pyruvate is therefore not easily redirected in high amounts to secondary metabolic routes. PMID:10473408

  9. 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.

  10. Crystal structure of the bis(cyclohexylammonium succinate succinic acid salt adduct

    Directory of Open Access Journals (Sweden)

    Modou Sarr

    2015-08-01

    Full Text Available The crystal structure of the title salt adduct, 2C6H14N+·C4H4O42−·C4H6O4, consists of two cyclohexylammonium cations, one succcinate dianion and one neutral succinic acid molecule. Succinate dianions and succinic acid molecules are self-assembled head-to-tail through O—H...O hydrogen bonds and adopt a syn–syn configuration, leading to a strand-like arrangement along [101]. The cyclohexylammonium cations have a chair conformation and act as multidentate hydrogen-bond donors linking adjacent strands through intermolecular N—H...O interactions to both the succinate and the succinic acid components. This results in two-dimensional supramolecular layered structures lying parallel to (010.

  11. The frequency of a disease-causing point mutation in the gene coding for medium-chain acyl-CoA dehydrogenase in sudden infant death syndrome

    DEFF Research Database (Denmark)

    Banner, Jytte; Gregersen, N; Kølvraa, S

    1993-01-01

    syndrome is still a matter of controversy. The present study investigated 120 well-defined cases of sudden infant death syndrome in order to detect the frequency of the most common disease-causing point mutation in the gene coding for medium-chain acyl-CoA dehydrogenase (G985) compared with the frequency...... in the general population. A highly specific polymerase chain reaction assay was applied on dried blood spots. No over-representation of homo- or heterozygosity for G985 appears to exist in such a strictly defined population, for which reason it may be more relevant to look at a broader spectrum of clinical...... presentations of inherited metabolic disorders and examine a wider range of sudden death in infancy....

  12. The most common mutation causing medium-chain acyl-CoA dehydrogenase deficiency is strongly associated with a particular haplotype in the region of the gene

    DEFF Research Database (Denmark)

    Kølvraa, S; Gregersen, N; Blakemore, A I

    1991-01-01

    RFLP haplotypes in the region containing the medium-chain acyl-CoA dehydrogenase (MCAD) gene on chromosome 1 have been determined in patients with MCAD deficiency. The RFLPs were detected after digestion of patient DNA with the enzymes BanII. PstI and TaqI and with an MCAD cDNA-clone as a probe....... Of 32 disease-causing alleles studied, 31 possessed the previously published A----G point-mutation at position 985 of the cDNA. This mutation has been shown to result in inactivity of the MCAD enzyme. In at least 30 of the 31 alleles carrying this G985 mutation a specific RFLP haplotype was present...

  13. 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.

  14. 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.

  15. Diverse point mutations in the human glucose-6-phosphate dehydrogenase gene cause enzyme deficiency and mild or severe hemolytic anemia

    Energy Technology Data Exchange (ETDEWEB)

    Vulliamy, T.J.; D' Urso, M.; Battistuzzi, G.; Estrada, M.; Foulkes, N.S.; Martini, G.; Calabro, V.; Poggi, V.; Giordano, R.; Town, M.; Luzzatto, L.; Persico, M.G. (Royal Postgraduate Medical School, London (England))

    1988-07-01

    Glucose-6-phosphate dehydrogenase deficiency is a common genetic abnormality affecting an estimated 400 million people worldwide. Clinical and biochemical analyses have identified many variants exhibiting a range of phenotypes, which have been well characterized from the hematological point of view. However, until now, their precise molecular basis has remained unknown. The authors have cloned and sequenced seven mutant G6PD alleles. In the nondeficient polymorphic African variant G6PD A they have found a single point mutation. The other six mutants investigated were all associated with enzyme deficiency. The mutations observed show a striking predominance of C {yields} T transitions, with CG doublets involved in four of seven cases. Thus, diverse point mutations may account largely for the phenotypic heterogeneity of G6PD deficiency.

  16. Targeted disruption of the housekeeping gene encoding glucose 6-phosphate dehydrogenase (G6PD-null): G6PD is dispensable for pentose synthesis but essential for defense against oxidative stress.

    NARCIS (Netherlands)

    P.P. Pandolfi; F. Sonati; R. Rivi; P. Mason; F.G. Grosveld (Frank); L. Luzzatto

    1995-01-01

    textabstractGlucose 6-phosphate dehydrogenase (G6PD) is a housekeeping enzyme encoded in mammals by an X-linked gene. It has important functions in intermediary metabolism because it catalyzes the first step in the pentose phosphate pathway and provides reductive potential in the form of NADPH. In

  17. Characterization of a dehydrogenase activity responsible for oxidation of 11-cis-retinol in the retinal pigment epithelium of mice with a disrupted RDH5 gene. A model for the human hereditary disease fundus albipunctatus.

    NARCIS (Netherlands)

    Jang, G.F.; Hooser, J.P. van; Kuksa, V.; McBee, J.K.; He, Y.G.; Janssen, J.J.M.; Driessen, C.A.G.G.; Palczewski, K.

    2001-01-01

    In the vertebrate retina, the final step of visual chromophore production is the oxidation of 11-cis-retinol to 11-cis-retinal. This reaction is catalyzed by 11-cis-retinol dehydrogenases (11-cis-RDHs), prior to the chromophore rejoining with the visual pigment apo-proteins. The RDH5 gene encodes a

  18. Selection of reference genes for qRT-PCR analysis of gene expression in sea cucumber Apostichopus japonicus during aestivation

    Science.gov (United States)

    Zhao, Ye; Chen, Muyan; Wang, Tianming; Sun, Lina; Xu, Dongxue; Yang, Hongsheng

    2014-11-01

    Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) is a technique that is widely used for gene expression analysis, and its accuracy depends on the expression stability of the internal reference genes used as normalization factors. However, many applications of qRT-PCR used housekeeping genes as internal controls without validation. In this study, the expression stability of eight candidate reference genes in three tissues (intestine, respiratory tree, and muscle) of the sea cucumber Apostichopus japonicus was assessed during normal growth and aestivation using the geNorm, NormFinder, delta CT, and RefFinder algorithms. The results indicate that the reference genes exhibited significantly different expression patterns among the three tissues during aestivation. In general, the β-tubulin (TUBB) gene was relatively stable in the intestine and respiratory tree tissues. The optimal reference gene combination for intestine was 40S ribosomal protein S18 (RPS18), TUBB, and NADH dehydrogenase (NADH); for respiratory tree, it was β-actin (ACTB), TUBB, and succinate dehydrogenase cytochrome B small subunit (SDHC); and for muscle it was α-tubulin (TUBA) and NADH dehydrogenase [ubiquinone] 1 α subcomplex subunit 13 (NDUFA13). These combinations of internal control genes should be considered for use in further studies of gene expression in A. japonicus during aestivation.

  19. Selection of housekeeping genes for gene expression studies in human reticulocytes using real-time PCR

    Science.gov (United States)

    Silver, Nicholas; Best, Steve; Jiang, Jie; Thein, Swee Lay

    2006-01-01

    Background Control genes, which are often referred to as housekeeping genes, are frequently used to normalise mRNA levels between different samples. However, the expression level of these genes may vary among tissues or cells and may change under certain circumstances. Thus, the selection of housekeeping genes is critical for gene expression studies. To address this issue, 7 candidate housekeeping genes including several commonly used ones were investigated in isolated human reticulocytes. For this, a simple ΔCt approach was employed by comparing relative expression of 'pairs of genes' within each sample. On this basis, stability of the candidate housekeeping genes was ranked according to repeatability of the gene expression differences among 31 samples. Results Initial screening of the expression pattern demonstrated that 1 of the 7 genes was expressed at very low levels in reticulocytes and was excluded from further analysis. The range of expression stability of the other 6 genes was (from most stable to least stable): GAPDH (glyceraldehyde 3-phosphate dehydrogenase), SDHA (succinate dehydrogenase), HPRT1 (hypoxanthine phosphoribosyl transferase 1), HBS1L (HBS1-like protein) and AHSP (alpha haemoglobin stabilising protein), followed by B2M (beta-2-microglobulin). Conclusion Using this simple approach, GAPDH was found to be the most suitable housekeeping gene for expression studies in reticulocytes while the commonly used B2M should be avoided. PMID:17026756

  20. Cloning of D-lactate dehydrogenase genes of Lactobacillus delbrueckii subsp. bulgaricus and their roles in D-lactic acid production.

    Science.gov (United States)

    Huang, Yanna; You, Chunping; Liu, Zhenmin

    2017-07-01

    Lactobacillus delbrueckii subsp. bulgaricus is a heterogenous lactic acid bacterium that converts pyruvate mainly to D-lactic acid using D-lactate dehydrogenases (D-LDHs), whose functional properties remain poorly characterized. Here, the D-LDHs genes (ldb0101, ldb0813, ldb1010, ldb1147 and ldb2021) were cloned and overexpressed in Escherichia coli JM109 from an inducible pUC18 vector, respectively, and the resulting strains were compared in terms of D-lactic acid production. The strain expressing ldb0101 and ldb1010 gene individually produced more D-lactate than other three strains. Further study revealed that Ldb0101 activity was down-regulated by the oxygen and, therefore, achieved a highest titer of D-lactate (1.94 g/L) under anaerobic condition, and introduction of ldb1010 gene enhanced D-lactate formation (0.94 and 0.85 g/L, respectively) both in aerobic and anaerobic conditions due to a relatively stable q d-lactate . Our results suggested that the enzyme Ldb0101 and Ldb1010 played a role of more importance in D-lactate formation. To the best of our knowledge, we demonstrate for the first time the roles of different D-LDH homologs from L. bulgaricus in D-lactic acid production.

  1. Performance of Glutamate Dehydrogenase and Triose Phosphate Isomerase Genes in the Analysis of Genotypic Variability of Isolates of Giardia duodenalis from Livestocks

    Science.gov (United States)

    Fava, Natália M. N.; Soares, Rodrigo M.; Scalia, Luana A. M.; Kalapothakis, Evanguedes; Pena, Isabella F.; Vieira, Carlos U.; Faria, Elaine S. M.; Cunha, Maria J.; Couto, Talles R.; Cury, Márcia Cristina

    2013-01-01

    Giardia duodenalis is a small intestinal protozoan parasite of several terrestrial vertebrates. This work aims to assess the genotypic variability of Giardia duodenalis isolates from cattle, sheep and pigs in the Southeast of Brazil, by comparing the standard characterization between glutamate dehydrogenase (gdh) and triose phosphate isomerase (tpi) primers. Fecal samples from the three groups of animals were analyzed using the zinc sulphate centrifugal flotation technique. Out of 59 positive samples, 30 were from cattle, 26 from sheep and 3 from pigs. Cyst pellets were stored and submitted to PCR and nested-PCR reactions with gdh and tpi primers. Fragment amplification of gdh and tpi genes was observed in 25 (42.4%) and 36 (61.0%) samples, respectively. Regarding the sequencing, 24 sequences were obtained with gdh and 20 with tpi. For both genes, there was a prevalence of E specific species assemblage, although some isolates have been identified as A and B, by the tpi sequencing. This has also shown a larger number of heterogeneous sequences, which have been attribute to mixed infections between assemblages B and E. The largest variability of inter-assemblage associated to the frequency of heterogeneity provided by tpi sequencing reinforces the polymorphic nature of this gene and makes it an excellent target for studies on molecular epidemiology. PMID:24308010

  2. Effect of hypoxia on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells

    Directory of Open Access Journals (Sweden)

    O. H. Minchenko

    2016-06-01

    Full Text Available We have studied the effect of hypoxia on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells under the inhibition of IRE1 (inositol requiring enzyme-1, which controls cell proliferation and tumor growth as a central mediator of endoplasmic reticulum stress. It was shown that hypoxia down-regulated gene expression of malate dehydrogenase 2 (MDH2, malic enzyme 2 (ME2, mitochondrial aspartate aminotransferase (GOT2, and subunit B of succinate dehydrogenase (SDHB in control (transfected by empty vector glioma cells in a gene specific manner. At the same time, the expression level of mitochondrial NADP+-dependent isocitrate dehydrogenase 2 (IDH2 and subunit D of succinate dehydrogenase (SDHD genes in these cells does not significantly change in hypoxic conditions. It was also shown that the inhibition of ІRE1 signaling enzyme function in U87 glioma cells decreases the effect of hypoxia on the expression of ME2, GOT2, and SDHB genes and introduces the sensitivity of IDH2 gene to hypoxia. Furthermore, the expression of all studied genes depends on IRE1-mediated endoplasmic reticulum stress signaling in gene specific manner, because ІRE1 knockdown significantly decreases their expression in normoxic conditions, except for IDH2 gene, which expression level is strongly up-regulated. Therefore, changes in the expression level of nuclear genes encoding ME2, MDH2, IDH2, SDHB, SDHD, and GOT2 proteins possibly reflect metabolic reprogramming of mitochondria by hypoxia and IRE1-mediated endoplasmic reticulum stress signaling and correlate with suppression of glioma cell proliferation under inhibition of the IRE1 enzyme function.

  3. Regulation Mechanism of the ald Gene Encoding Alanine Dehydrogenase in Mycobacterium smegmatis and Mycobacterium tuberculosis by the Lrp/AsnC Family Regulator AldR.

    Science.gov (United States)

    Jeong, Ji-A; Hyun, Jaekyung; Oh, Jeong-Il

    2015-10-01

    In the presence of alanine, AldR, which belongs to the Lrp/AsnC family of transcriptional regulators and regulates ald encoding alanine dehydrogenase in Mycobacterium smegmatis, changes its quaternary structure from a homodimer to an octamer with an open-ring conformation. Four AldR-binding sites (O2, O1, O4, and O3) with a consensus sequence of GA/T-N2-NWW/WWN-N2-A/TC were identified upstream of the M. smegmatis ald gene by means of DNase I footprinting analysis. O2, O1, and O4 are required for the induction of ald expression by alanine, while O3 is directly involved in the repression of ald expression. In addition to O3, both O1 and O4 are also necessary for full repression of ald expression in the absence of alanine, due to cooperative binding of AldR dimers to O1, O4, and O3. Binding of a molecule of the AldR octamer to the ald control region was demonstrated to require two AldR-binding sites separated by three helical turns between their centers and one additional binding site that is in phase with the two AldR-binding sites. The cooperative binding of AldR dimers to DNA requires three AldR-binding sites that are aligned with a periodicity of three helical turns. The aldR gene is negatively autoregulated independently of alanine. Comparative analysis of ald expression of M. smegmatis and Mycobacterium tuberculosis in conjunction with sequence analysis of both ald control regions led us to suggest that the expression of the ald genes in both mycobacterial species is regulated by the same mechanism. In mycobacteria, alanine dehydrogenase (Ald) is the enzyme required both to utilize alanine as a nitrogen source and to grow under hypoxic conditions by maintaining the redox state of the NADH/NAD(+) pool. Expression of the ald gene was reported to be regulated by the AldR regulator that belongs to the Lrp/AsnC (feast/famine) family, but the underlying mechanism was unknown. This study revealed the regulation mechanism of ald in Mycobacterium smegmatis and

  4. Gene structure and mutations of glutaryl-coenzyme A dehydrogenase: impaired association of enzyme subunits that is due to an A421V substitution causes glutaric acidemia type I in the Amish.

    OpenAIRE

    Biery, B. J.; Stein, D. E.; Morton, D. H.; Goodman, S. I.

    1996-01-01

    The structure of the human glutaryl coenzyme A dehydrogenase (GCD) gene was determined to contain 11 exons and to span approximately 7 kb. Fibroblast DNA from 64 unrelated glutaric acidemia type I (GA1) patients was screened for mutations by PCR amplification and analysis of SSCP. Fragments with altered electrophoretic mobility were subcloned and sequenced to detect mutations that caused GA1. This report describes the structure of the GCD gene, as well as point mutations and polymorphisms fou...

  5. Isolation and characterization of poly(butylene succinate-co-butylene adipate)-degrading microorganism.

    Science.gov (United States)

    Hayase, Nobuki; Yano, Hidetoshi; Kudoh, Emi; Tsutsumi, Chikara; Ushio, Kazutoshi; Miyahara, Yasushi; Tanaka, Suminori; Nakagawa, Katsuhiko

    2004-01-01

    Poly(butylene succinate-co-butylene adipate) (PBSA)-degrading bacterium, strain 1-A, was isolated from soil. Strain 1-A was identified as Bacillus pumilus on the basis of its physiological properties and partial 16S rRNA gene sequence. Strain 1-A also degraded poly(butylene succinate) (PBS) and poly(epsilon-caprolactone). On the other hand, poly(butylene adipate terephthalate) and poly(lactic acid) were minimally degraded by strain 1-A. The NMR spectra of degradation products from PBSA indicated that the adipate units were more rapidly degraded than 1,4-butanediol and succinate units. This seems to be one of the reasons why strain 1-A degraded PBSA faster than PBS.

  6. Construction of a recombinant plasmid harbouring the glyceraldenyde-3-phosphate dehydrogenase gene of periodic Brugia malayi and observation on DNA immunity.

    Science.gov (United States)

    Fang, Z; Tong, H; Zhang, S; Fang, H; Lu, S; Xu, B

    2012-01-01

    Controlling and eliminating lymphatic filariasis will require further research of preventative measures and implementation. Parasite is dependent on glycolysis for ATP production. The glycolytic enzyme glyceraldenyde-3-phosphate dehydrogenase (GAPDH) plays an important role in glycolysis and therefore is either a potential target for anti-parasite drug development or a vaccine candidate. Therefore, we tried to investigate the DNA vaccine-elicited immune responses in BALB/c mice. We cloned a gene encoding the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from periodic Brugia malayi into vector pcDNA3.1. Mice were injected at a dosage of 100 μg recombinant plasmid DNA with CpG intramuscular injection and immunized three times at 2-week intervals. pcDNA3.1 and normal saline were used as control. The tissue of muscles at the 4 weeks after the third injection was collected and target genes were detected using RT-PCR. The humoral responses elicited in mice by inoculation with the recombinant plasmid pcDNA3.1-BmGAPDH were detected using a standard ELISA. Two weeks after the third immunization, stimulation index (SI) was measured using the MTT method and the level of secreted IL-4 and INF-g were detected using ELISA. Specific gene fragment coding GAPDH was amplified and the recombinant plasmid pcDNA3.1-BmGAPDH was constructed. Post-challenge sera from the mice immunized with the DNA vaccine had specific antibody titres of 1:1600 to 1:6400, and the highest titre was observed in the mice that were inoculated by pcDNA3.1-BmGAPDH/CpG at 6 weeks. At 4 weeks after immunization, the spleens of the mice were obviously enlarged. The proliferation of spleen T lymphocytes seen on the MTT assay was higher in the pcDNA3.1-BmGAPDH group than in the control group (P value immunized mice were significantly higher than that of the control (P value immune responses in mice.

  7. 21 CFR 172.275 - Synthetic paraffin and succinic derivatives.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Synthetic paraffin and succinic derivatives. 172... FOOD FOR HUMAN CONSUMPTION Coatings, Films and Related Substances § 172.275 Synthetic paraffin and succinic derivatives. Synthetic paraffin and succinic derivatives identified in this section may be safely...

  8. Overexpression of the NADP+-specific isocitrate dehydrogenase gene (icdA) in citric acid-producing Aspergillus niger WU-2223L.

    Science.gov (United States)

    Kobayashi, Keiichi; Hattori, Takasumi; Hayashi, Rie; Kirimura, Kohtaro

    2014-01-01

    In the tricarboxylic acid (TCA) cycle, NADP(+)-specific isocitrate dehydrogenase (NADP(+)-ICDH) catalyzes oxidative decarboxylation of isocitric acid to form α-ketoglutaric acid with NADP(+) as a cofactor. We constructed an NADP(+)-ICDH gene (icdA)-overexpressing strain (OPI-1) using Aspergillus niger WU-2223L as a host and examined the effects of increase in NADP(+)-ICDH activity on citric acid production. Under citric acid-producing conditions with glucose as the carbon source, the amounts of citric acid produced and glucose consumed by OPI-1 for the 12-d cultivation period decreased by 18.7 and 10.5%, respectively, compared with those by WU-2223L. These results indicate that the amount of citric acid produced by A. niger can be altered with the NADP(+)-ICDH activity. Therefore, NADP(+)-ICDH is an important regulator of citric acid production in the TCA cycle of A. niger. Thus, we propose that the icdA gene is a potentially valuable tool for modulating citric acid production by metabolic engineering.

  9. Cloning and molecular characterization of the glyceraldehyde-3-phosphate dehydrogenase-encoding gene and cDNA from the plant pathogenic fungus Glomerella cingulata.

    Science.gov (United States)

    Templeton, M D; Rikkerink, E H; Solon, S L; Crowhurst, R N

    1992-12-01

    The glyceraldehyde-3-phosphate dehydrogenase gene (gpdA) has been identified from a genomic DNA library prepared from the plant pathogenic fungus Glomerella cingulata. Nucleotide sequence data revealed that this gene codes for a putative 338-amino-acid protein encoded by two exons of 129 and 885 bp, separated by an intron 216 bp long. The 5' leader sequence is also spliced by an intron of 156 bp. A cDNA clone was prepared using the polymerase chain reaction, the sequence of which was used to confirm the presence of the intron in the coding sequence and the splicing of the 5' leader sequence. The transcriptional start point (tsp) was mapped at -253 nt from the site of the initiation of translation by primer extension and is adjacent to a 42-bp pyrimidine-rich region. The general structure of the 5' flanking region shows similarities to gpdA from Aspergillus nidulans. The putative protein product is 71-86% identical at the aa level to GPDs from Aspergillus nidulans, Cryphonectria parasitica, Curvularia lunata, Podospora anserina and Ustilago maydis.

  10. 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.

  11. Caenorhabditis elegans expressing the Saccharomyces cerevisiae NADH alternative dehydrogenase Ndi1p, as a tool to identify new genes involved in complex I related diseases

    Directory of Open Access Journals (Sweden)

    Raynald eCossard

    2015-06-01

    Full Text Available Isolated complex I deficiencies are one of the most commonly observed biochemical features in patients suffering from mitochondrial disorders. In the majority of these clinical cases the molecular bases of the diseases remain unknown suggesting the involvement of unidentified factors that are critical for complex I function.The Saccharomyces cerevisiae NDI1 gene, encoding the mitochondrial internal NADH dehydrogenase was previously shown to complement a complex I deficient strain in Caenorhabitis elegans with notable improvements in reproduction, whole organism respiration. These features indicate that Ndi1p can functionally integrate the respiratory chain, allowing complex I deficiency complementation. Taking into account the Ndi1p ability to bypass complex I, we evaluate the possibility to extend the range of defects/mutations causing complex I deficiencies that can be alleviated by NDI1 expression.We report here that NDI1 expressing animals unexpectedly exhibit a slightly shortened lifespan, a reduction in the progeny and a depletion of the mitochondrial genome. However, Ndi1p is expressed and targeted to the mitochondria as a functional protein that confers rotenone resistance to those animals and without affecting their respiration rate and ATP content.We show that the severe embryonic lethality level caused by the RNAi knockdowns of complex I structural subunit encoding genes (e.g. NDUFV1, NDUFS1, NDUFS6, NDUFS8 or GRIM-19 human orthologs in wild type animals is significantly reduced in the Ndi1p expressing worm.All together these results open up the perspective to identify new genes involved in complex I function, assembly or regulation by screening an RNAi library of genes leading to embryonic lethality that should be rescued by NDI1 expression.

  12. From gene to structure: Lactobacillus bulgaricus D-lactate dehydrogenase from yogurt as an integrated curriculum model for undergraduate molecular biology and biochemistry laboratory courses.

    Science.gov (United States)

    Lawton, Jeffrey A; Prescott, Noelle A; Lawton, Ping X

    2018-03-07

    We have developed an integrated, project-oriented curriculum for undergraduate molecular biology and biochemistry laboratory courses spanning two semesters that is organized around the ldhA gene from the yogurt-fermenting bacterium Lactobacillus bulgaricus, which encodes the enzyme d-lactate dehydrogenase. The molecular biology module, which consists of nine experiments carried out over eleven sessions, begins with the isolation of genomic DNA from L. bulgaricus in yogurt and guides students through the process of cloning the ldhA gene into a prokaryotic expression vector, followed by mRNA isolation and characterization of recombinant gene expression levels using RT-PCR. The biochemistry module, which consists of nine experiments carried out over eight sessions, begins with overexpression of the cloned ldhA gene and guides students through the process of affinity purification, biochemical characterization of the purified LdhA protein, and analysis of enzyme kinetics using various substrates and an inhibitor, concluding with a guided inquiry investigation of structure-function relationships in the three-dimensional structure of LdhA using molecular visualization software. Students conclude by writing a paper describing their work on the project, formatted as a manuscript to be submitted for publication in a scientific journal. Overall, this curriculum, with its emphasis on experiential learning, provides hands-on training with a variety of common laboratory techniques in molecular biology and biochemistry and builds experience with the process of scientific reasoning, along with reinforcement of essential transferrable skills such as critical thinking, information literacy, and written communication, all within the framework of an extended project having the look and feel of a research experience. © 2018 by The International Union of Biochemistry and Molecular Biology, 2018. © 2018 The International Union of Biochemistry and Molecular Biology.

  13. 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.

  14. The effect of alpha-tocopheryl succinate on succinate respiration in rat liver mitochondria

    Czech Academy of Sciences Publication Activity Database

    Sobotka, O.; Drahota, Zdeněk; Kučera, O.; Endlicher, R.; Rauchová, Hana; Červinková, Z.

    2015-01-01

    Roč. 64, Suppl.5 (2015), S609-S615 ISSN 0862-8408 Institutional support: RVO:67985823 Keywords : tocopheryl succinate * Complex II * liver * mitochondria * homogenate * hepatocytes Subject RIV: ED - Physiology Impact factor: 1.643, year: 2015

  15. Gold nanoparticles/water-soluble carbon nanotubes/aromatic diamine polymer composite films for highly sensitive detection of cellobiose dehydrogenase gene

    Energy Technology Data Exchange (ETDEWEB)

    Zeng Guangming, E-mail: zgming@hnu.cn [College of Environmental Science and Engineering, Hunan University, Changsha 410082 (China); Key Laboratory of Environmental Biology and Pollution Control (Hunan University), Ministry of Education, Changsha 410082 (China); Li Zhen, E-mail: happylizhen@yeah.ne [College of Environmental Science and Engineering, Hunan University, Changsha 410082 (China); Key Laboratory of Environmental Biology and Pollution Control (Hunan University), Ministry of Education, Changsha 410082 (China); Tang Lin; Wu Mengshi; Lei Xiaoxia; Liu Yuanyuan; Liu Can; Pang Ya; Zhang Yi [College of Environmental Science and Engineering, Hunan University, Changsha 410082 (China); Key Laboratory of Environmental Biology and Pollution Control (Hunan University), Ministry of Education, Changsha 410082 (China)

    2011-05-01

    Highlights: > Gold nanoparticles/multiwalled carbon nanotubes/poly (1,5-naphthalenediamine) modified electrode was fabricated. > The sensor was applied for the detection of cellobiose dehydrogenase genes. > An effective method to distribute MWCNTs and attach to the electrode was proposed. > The composite films greatly improved the sensitivity and enhanced the DNA immobilization. > The DNA biosensor exhibited fairly high sensitivity and quite low detection limit. - Abstract: An electrochemical sensor based on gold nanoparticles (GNPs)/multiwalled carbon nanotubes (MWCNTs)/poly (1,5-naphthalenediamine) films modified glassy carbon electrode (GCE) was fabricated. The effectiveness of the sensor was confirmed by sensitive detection of cellobiose dehydrogenase (CDH) gene which was extracted from Phanerochaete chrysosporium using polymerase chain reaction (PCR). The monomer of 1,5-naphthalenediamine was electropolymerized on the GCE surface with abundant free amino groups which enhanced the stability of MWCNTs modified electrode. Congo red (CR)-functionalized MWCNTs possess excellent conductivity as well as high solubility in water which enabled to form the uniform and stable network nanostructures easily and created a large number of binding sites for electrodeposition of GNPs. The continuous GNPs together with MWCNTs greatly increased the surface area, conductivity and electrocatalytic activity. This electrode structure significantly improved the sensitivity of sensor and enhanced the DNA immobilization and hybridization. The thiol modified capture probes were immobilized onto the composite films-modified GCE by a direct formation of thiol-Au bond and horseradish peroxidase-streptavidin (HRP-SA) conjugates were labeled to the biotinylated detection probes through biotin-streptavidin bond. Scanning electron microscopy (SEM), electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were used to investigate the film assembly and DNA hybridization processes

  16. 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.

  17. The enhancement of tolerance to salt and cold stresses by modifying the redox state and salicylic acid content via the cytosolic malate dehydrogenase gene in transgenic apple plants.

    Science.gov (United States)

    Wang, Qing-Jie; Sun, Hong; Dong, Qing-Long; Sun, Tian-Yu; Jin, Zhong-Xin; Hao, Yu-Jin; Yao, Yu-Xin

    2016-10-01

    In this study, we characterized the role of an apple cytosolic malate dehydrogenase gene (MdcyMDH) in the tolerance to salt and cold stresses and investigated its regulation mechanism in stress tolerance. The MdcyMDH transcript was induced by mild cold and salt treatments, and MdcyMDH-overexpressing apple plants possessed improved cold and salt tolerance compared to wild-type (WT) plants. A digital gene expression tag profiling analysis revealed that MdcyMDH overexpression largely altered some biological processes, including hormone signal transduction, photosynthesis, citrate cycle and oxidation-reduction. Further experiments verified that MdcyMDH overexpression modified the mitochondrial and chloroplast metabolisms and elevated the level of reducing power, primarily caused by increased ascorbate and glutathione, as well as the increased ratios of ascorbate/dehydroascorbate and glutathione/glutathione disulphide, under normal and especially stress conditions. Concurrently, the transgenic plants produced a high H2 O2 content, but a low O2·- production rate was observed compared to the WT plants. On the other hand, the transgenic plants accumulated more free and total salicylic acid (SA) than the WT plants under normal and stress conditions. Taken together, MdcyMDH conferred the transgenic apple plants a higher stress tolerance by producing more reductive redox states and increasing the SA level; MdcyMDH could serve as a target gene to genetically engineer salt- and cold-tolerant trees. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  18. Genes for selenium dependent and independent formate dehydrogenase in the gut microbial communities of three lower, wood-feeding termites and a wood-feeding roach.

    Science.gov (United States)

    Zhang, Xinning; Matson, Eric G; Leadbetter, Jared R

    2011-02-01

    The bacterial Wood-Ljungdahl pathway for CO(2)-reductive acetogenesis is important for the nutritional mutualism occurring between wood-feeding insects and their hindgut microbiota. A key step in this pathway is the reduction of CO(2) to formate, catalysed by the enzyme formate dehydrogenase (FDH). Putative selenocysteine- (Sec) and cysteine- (Cys) containing paralogues of hydrogenase-linked FDH (FDH(H)) have been identified in the termite gut acetogenic spirochete, Treponema primitia, but knowledge of their relevance in the termite gut environment remains limited. In this study, we designed degenerate PCR primers for FDH(H) genes (fdhF) and assessed fdhF diversity in insect gut bacterial isolates and the gut microbial communities of termites and cockroaches. The insects examined herein represent three wood-feeding termite families, Termopsidae, Kalotermitidae and Rhinotermitidae (phylogenetically 'lower' termite taxa); the wood-feeding roach family Cryptocercidae (the sister taxon to termites); and the omnivorous roach family Blattidae. Sec and Cys FDH(H) variants were identified in every wood-feeding insect but not the omnivorous roach. Of 68 novel alleles obtained from inventories, 66 affiliated phylogenetically with enzymes from T. primitia. These formed two subclades (37 and 29 phylotypes) almost completely comprised of Sec-containing and Cys-containing enzymes respectively. A gut cDNA inventory showed transcription of both variants in the termite Zootermopsis nevadensis (family Termopsidae). The gene patterns suggest that FDH(H) enzymes are important for the CO(2)-reductive metabolism of uncultured acetogenic treponemes and imply that the availability of selenium, a trace element, shaped microbial gene content in the last common ancestor of dictyopteran, wood-feeding insects, and continues to shape it to this day.

  19. 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 ...

  20. Long-term correction of very long-chain acyl-coA dehydrogenase deficiency in mice using AAV9 gene therapy.

    Science.gov (United States)

    Keeler, Allison M; Conlon, Thomas; Walter, Glenn; Zeng, Huadong; Shaffer, Scott A; Dungtao, Fu; Erger, Kirsten; Cossette, Travis; Tang, Qiushi; Mueller, Christian; Flotte, Terence R

    2012-06-01

    Very long-chain acyl-coA dehydrogenase (VLCAD) is the rate-limiting step in mitochondrial fatty acid oxidation. VLCAD-deficient mice and patients clinical symptoms stem from not only an energy deficiency but also long-chain metabolite accumulations. VLCAD-deficient mice were treated systemically with 1 × 10(12) vector genomes of recombinant adeno-associated virus 9 (rAAV9)-VLCAD. Biochemical correction was observed in vector-treated mice beginning 2 weeks postinjection, as characterized by a significant drop in long-chain fatty acyl accumulates in whole blood after an overnight fast. Changes persisted through the termination point around 20 weeks postinjection. Magnetic resonance spectroscopy (MRS) and tandem mass spectrometry (MS/MS) revealed normalization of intramuscular lipids in treated animals. Correction was not observed in liver tissue extracts, but cardiac muscle extracts showed significant reduction of long-chain metabolites. Disease-specific phenotypes were characterized, including thermoregulation and maintenance of euglycemia after a fasting cold challenge. Internal body temperatures of untreated VLCAD(-/-) mice dropped below 20 °C and the mice became lethargic, requiring euthanasia. In contrast, all rAAV9-treated VLCAD(-/-) mice and the wild-type controls maintained body temperatures. rAAV9-treated VLCAD(-/-) mice maintained euglycemia, whereas untreated VLCAD(-/-) mice suffered hypoglycemia following a fasting cold challenge. These promising results suggest rAAV9 gene therapy as a potential treatment for VLCAD deficiency in humans.

  1. Over-expression of Arabidopsis thaliana SFD1/GLY1, the gene encoding plastid localized glycerol-3-phosphate dehydrogenase, increases plastidic lipid content in transgenic rice plants.

    Science.gov (United States)

    Singh, Vijayata; Singh, Praveen Kumar; Siddiqui, Adnan; Singh, Subaran; Banday, Zeeshan Zahoor; Nandi, Ashis Kumar

    2016-03-01

    Lipids are the major constituents of all membranous structures in plants. Plants possess two pathways for lipid biosynthesis: the prokaryotic pathway (i.e., plastidic pathway) and the eukaryotic pathway (i.e., endoplasmic-reticulum (ER) pathway). Whereas some plants synthesize galactolipids from diacylglycerol assembled in the plastid, others, including rice, derive their galactolipids from diacylglycerols assembled by the eukaryotic pathway. Arabidopsis thaliana glycerol-3-phosphate dehydrogenase (G3pDH), coded by SUPPRESSOR OF FATTY ACID DESATURASE 1 (SFD1; alias GLY1) gene, catalyzes the formation of glycerol 3-phosphate (G3p), the backbone of many membrane lipids. Here SFD1 was introduced to rice as a transgene. Arabidopsis SFD1 localizes in rice plastids and its over-expression increases plastidic membrane lipid content in transgenic rice plants without any major impact on ER lipids. The results suggest that over-expression of plastidic G3pDH enhances biosynthesis of plastid-localized lipids in rice. Lipid composition in the transgenic plants is consistent with increased phosphatidylglycerol synthesis in the plastid and increased galactolipid synthesis from diacylglycerol produced via the ER pathway. The transgenic plants show a higher photosynthetic assimilation rate, suggesting a possible application of this finding in crop improvement.

  2. Purification and Characterization of Cytoplasmic NADP+-Isocitrate Dehydrogenase, and Amplification of the Nadp+-IDH Gene from the Wing-Dimorphic Sand Field Cricket, Gryllus firmus

    Science.gov (United States)

    Zera, Anthony J.; Newman, Susan; Berkheim, David; Black, Christine; Klug, Lindsay; Crone, Erica

    2011-01-01

    Cytoplasmic NADP+-isocitrate dehydrogenase (NADP+-IDH) has been purified and characterized, and its gene sequenced in many animal, plant, and yeast species. However, much less information is available on this enzyme-gene in insects. As a first step in investigating the biochemical and molecular mechanisms by which NADP+-IDH contributes to adaptations for flight vs. reproduction in insects, the enzyme was purified to homogeneity in the wing-dimorphic cricket, Gryllus firmus, characterized, and its corresponding gene sequenced. Using a combination of polyethylene glycol precipitation, Cibacron-Blue affinity chromatography, and hydrophobic interaction chromatography the enzyme was purified 291-fold (7% yield; specific activity = 15.8 µmol NADPH/min/mg protein). The purified enzyme exhibited a single band on SDS PAGE (46.3 kD), but consisted of two N-terminal amino acid sequences that differed in the first two amino acids. Purified enzyme exhibited standard Michaelis-Menten kinetics at pH 8.0 and 28° C (KM(NADP+) = 2.3 ± 0.4 µM; KM(Na+-Isocitrate) = 14.7 + 1.8 µM). Subunit molecular mass and KMS were similar to published values for NADP+-IDHs from a variety of vertebrate and two insect species. PCR amplification of an internal sequence using genomic DNA followed by 3′ and 5′ RACE yielded a nucleotide sequence of the mature protein and translated amino-acid sequences that exhibited high similarity (40–50% and 70–80%, respectively) to sequences from insect and vertebrate NADP+-IDHs. Two potential ATG start codons were identified. Both Nterminal amino-acid sequences matched the nucleotide sequence, consistent with both enzyme forms being transcribed from the same gene, although these variants could also be encoded by different genes. Bioinformatic analyses and differential centrifugation indicated that the majority, if not all, of the enzyme is cytoplasmic. The enzyme exhibited high specific activity in fat body, head and gut, and a single band on native PAGE

  3. The gdhB gene of Pseudomonas aeruginosa encodes an arginine-inducible NAD(+)-dependent glutamate dehydrogenase which is subject to allosteric regulation.

    Science.gov (United States)

    Lu, C D; Abdelal, A T

    2001-01-01

    The NAD(+)-dependent glutamate dehydrogenase (NAD-GDH) from Pseudomonas aeruginosa PAO1 was purified, and its amino-terminal amino acid sequence was determined. This sequence information was used in identifying and cloning the encoding gdhB gene and its flanking regions. The molecular mass predicted from the derived sequence for the encoded NAD-GDH was 182.6 kDa, in close agreement with that determined from sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified enzyme (180 kDa). Cross-linking studies established that the native NAD-GDH is a tetramer of equal subunits. Comparison of the derived amino acid sequence of NAD-GDH from P. aeruginosa with the GenBank database showed the highest homology with hypothetical polypeptides from Pseudomonas putida, Mycobacterium tuberculosis, Rickettsia prowazakii, Legionella pneumophila, Vibrio cholerae, Shewanella putrefaciens, Sinorhizobium meliloti, and Caulobacter crescentus. A moderate degree of homology, primarily in the central domain, was observed with the smaller tetrameric NAD-GDH (protomeric mass of 110 kDa) from Saccharomyces cerevisiae or Neurospora crassa. Comparison with the yet smaller hexameric GDH (protomeric mass of 48 to 55 kDa) of other prokaryotes yielded a low degree of homology that was limited to residues important for binding of substrates and for catalytic function. NAD-GDH was induced 27-fold by exogenous arginine and only 3-fold by exogenous glutamate. Primer extension experiments established that transcription of gdhB is initiated from an arginine-inducible promoter and that this induction is dependent on the arginine regulatory protein, ArgR, a member of the AraC/XyIS family of regulatory proteins. NAD-GDH was purified to homogeneity from a recombinant strain of P. aeruginosa and characterized. The glutamate saturation curve was sigmoid, indicating positive cooperativity in the binding of glutamate. NAD-GDH activity was subject to allosteric control by arginine and citrate, which

  4. Ontogeny of glucocorticoid receptor and 11beta-hydroxysteroid dehydrogenase type-1 gene expression identifies potential critical periods of glucocorticoid susceptibility during development.

    Science.gov (United States)

    Speirs, H J L; Seckl, J R; Brown, R W

    2004-04-01

    Glucocorticoids play important roles in organ development and 'fetal programming'. Fetal exposure to excess glucocorticoids reduces birth weight and causes later hypertension. To investigate these processes further we have determined the detailed ontogeny in the mouse of the glucocorticoid receptor (GR) and 11beta-hydroxysteroid dehydrogenase type-1 (11beta-HSD1), which amplifies glucocorticoid levels locally; the ontogeny was determined using in situ hybridisation from embryonic day 9.5 (E9.5, term=E19) until after birth. At E9.5 fetal GR mRNA levels are very low, except in fetal placenta. GR gene expression rises during gestation with striking tissue-specific differences in timing and extent. Before E13.5, an increase is clear in gastrointestinal (GI) and upper respiratory tracts, discrete central nervous system (CNS) regions, precartilage and especially in the liver (E10.5-E12). Later, further increases occur in lung, GI and upper respiratory tracts, muscle, pituitary and thymus. In a few tissues such increases are temporary, e.g. ureteric ducts (E13.5-E16.5) and pancreas (E14.5-E16.5, expression later falling sharply). Fetal 11beta-HSD1 mRNA expression is first clearly observed at E14.5-E15, initially in the fetal placenta then in the umbilical cord. Later, 11beta-HSD1 expression is seen as follows: (i) from E15 in lung and liver, rising strongly; (ii) thymus, from E15 (lower level); (iii) at low levels in a few brain regions, including the hippocampus (E16.5+); and (iv) in muscle group fascial planes and tendon insertions. This is the first detailed study of the ontogeny of these two genes and, in combination with previous work on the ontogeny of 11beta-HSD2 and the mineralocorticoid receptor, suggests potential critical periods of glucocorticoid sensitivity during development for several organ systems.

  5. Over-expression of a glutamate dehydrogenase gene, MgGDH, from Magnaporthe grisea confers tolerance to dehydration stress in transgenic rice.

    Science.gov (United States)

    Zhou, Yanbiao; Zhang, Caisheng; Lin, Jianzhong; Yang, Yuanzhu; Peng, Yuchong; Tang, Dongying; Zhao, Xiaoying; Zhu, Yonghua; Liu, Xuanming

    2015-03-01

    Heterologous expression of a fungal NADP(H)-GDH gene ( MgGDH ) from Magnaporthe grisea can improve dehydration stress tolerance in rice by preventing toxic accumulation of ammonium. Glutamate dehydrogenase (GDH; EC 1.4.1.2 and EC 1.4.1.4) may act as a stress-responsive enzyme in detoxification of high intracellular ammonia and production of glutamate for proline synthesis under stress conditions. In present study, a fungal NADP(H)-GDH gene (MgGDH) from Magnaporthe grisea was over-expressed in rice (Oryza sativa L. cv. 'kitaake'), and the transgenic plants showed the improvement of tolerance to dehydration stress. The kinetic analysis showed that His-TF-MgGDH preferentially utilizes ammonium to produce L-glutamate. Moreover, the affinity of His-TF-MgGDH for ammonium was dramatically higher than that of His-TF-OsGDH for ammonium. Over-expressing MgGDH transgenic rice plants showed lower water-loss rate and higher completely close stomata than the wild-type plants under dehydration stress conditions. In transgenic plants, the NADP(H)-GDH activities were markedly higher than those in wild-type plants and the amination activity was significantly higher than the deamination activity. Compared with wild-type plants, the transgenic plants accumulated much less NH4 (+) but higher amounts of glutamate, proline and soluble sugar under dehydration stress conditions. These results indicate that heterologous expression of MgGDH can prevent toxic accumulation of ammonium and in return improve dehydration stress tolerance in rice.

  6. Fluxomic evidence for impaired contribution of short-chain acyl-CoA dehydrogenase to mitochondrial palmitate β-oxidation in symptomatic patients with ACADS gene susceptibility variants.

    Science.gov (United States)

    Dessein, Anne-Frédérique; Fontaine, Monique; Joncquel-Chevalier Curt, Marie; Briand, Gilbert; Sechter, Claire; Mention-Mulliez, Karine; Dobbelaere, Dries; Douillard, Claire; Lacour, Arnaud; Redonnet-Vernhet, Isabelle; Lamireau, Delphine; Barth, Magalie; Minot-Myhié, Marie-Christine; Kuster, Alice; de Lonlay, Pascale; Gregersen, Niels; Acquaviva, Cécile; Vianey-Saban, Christine; Vamecq, Joseph

    2017-08-01

    Despite ACADS (acyl-CoA dehydrogenase, short-chain) gene susceptibility variants (c.511C>T and c.625G>A) are considered to be non-pathogenic, encoded proteins are known to exhibit altered kinetics. Whether or not, they might affect overall fatty acid β-oxidation still remains, however, unclear. De novo biosynthesis of acylcarnitines by whole blood samples incubated with deuterated palmitate (16- 2 H 3 ,15- 2 H 2 -palmitate) is suitable as a fluxomic exploration to distinguish between normal and disrupted β-oxidation, abnormal profiles and ratios of acylcarnitines with different chain-lengths being indicative of the site for enzymatic blockade. Determinations in 301 control subjects of ratios between deuterated butyrylcarnitine and sum of deuterated C2 to C14 acylcarnitines served here as reference values to state specifically functional SCAD impairment in patients addressed for clinical and/or biological suspicion of a β-oxidation disorder. Functional SCAD impairment was found in 39 patients. The 27 patients accepting subsequent gene studies were all positive for ACADS mutations. Twenty-six of 27 patients were positive for c.625G>A variant. Twenty-three of 27 patients harbored susceptibility variants as sole ACADS alterations (18 homozygous and 3 heterozygous for c.625G>A, 2 compound heterozygous for c.625G>A/c.511C>T). Our present fluxomic assessment of SCAD suggests a link between ACADS susceptibility variants and abnormal β-oxidation consistent with known altered kinetics of these variants. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. RNAi knock-downs support roles for the mucin-like (AeIMUC1) gene and short-chain dehydrogenase/reductase (SDR) gene in Aedes aegypti susceptibility to Plasmodium gallinaceum.

    Science.gov (United States)

    Berois, M; Romero-Severson, J; Severson, D W

    2012-03-01

    The mosquito midgut represents the first barrier encountered by the Plasmodium parasite (Haemosporida: Plasmodiidae) when it is ingested in blood from an infected vertebrate. Previous studies identified the Aedes aegypti (L.) (Diptera: Culicidae) mucin-like (AeIMUC1) and short-chain dehydrogenase/reductase (SDR) genes as midgut-expressed candidate genes influencing susceptibility to infection by Plasmodium gallinaceum (Brumpt). We used RNA inference (RNAi) by double-stranded RNA (dsRNA) injections to examine ookinete survival to the oocyst stage following individual gene knock-downs. Double-stranded RNA gene knock-downs were performed 3 days prior to P. gallinaceum infection and oocyst development was evaluated at 7 days post-infection. Mean numbers of parasites developing to the oocyst stage were significantly reduced by 52.3% in dsAeIMUC1-injected females and by 36.5% in dsSDR-injected females compared with females injected with a dsβ-gal control. The prevalence of infection was significantly reduced in dsAeIMUC1- and dsSDR-injected females compared with females injected with dsβ-gal; these reductions resulted in a two- and three-fold increase in the number of uninfected individuals, respectively. Overall, these results suggest that both AeIMUC1 and SDR play a role in Ae. aegypti vector competence to P. gallinaceum. © 2011 The Authors. Medical and Veterinary Entomology © 2011 The Royal Entomological Society.

  8. Mutagencity Testing of WR238605 Succinate

    Science.gov (United States)

    1996-05-03

    the study. Diane B. Madsen QUALITY ASSURANCE i-urtc* DATE L5178Y/TK+’ MOUSE LYMPHOMA MUTAGENESIS ASSAY FINAL REPORT Sponsor: Toxicology...FINAL REPORT Study Title CHROMOSOME ABERRATIONS IN CHINESE HAMSTER OVARY (CHO) CELLS Test Article WR238605 Succinate Authors Patrick T. Cuny ...IkuJdfflafiiAA l-5-fo Diane B. Madsen QUALITY ASSURANCE DATE CHO/HGPRT MUTATION ASSAY FINAL REPORT Sponsor: Toxicology Research Laboratory University

  9. Deletion of meso-2,3-butanediol dehydrogenase gene budC for enhanced D-2,3-butanediol production in Bacillus licheniformis

    Science.gov (United States)

    2014-01-01

    Background D-2,3-butanediol has many industrial applications such as chiral reagents, solvents, anti-freeze agents, and low freezing point fuels. Traditional D-2,3-butanediol producing microorganisms, such as Klebsiella pneumonia and K. xoytoca, are pathogenic and not capable of producing D-2,3-butanediol at high optical purity. Bacillus licheniformis is a potential 2,3-butanediol producer but the wild type strain (WX-02) produces a mix of D- and meso-type isomers. BudC in B. licheniformis is annotated as 2,3-butanediol dehydrogenase or acetoin reductase, but no pervious experiment was performed to verify this hypothesis. Results We developed a genetically modified strain of B. licheniformis (WX-02 ΔbudC) as a D-2,3-butanediol producer with high optimal purity. A marker-less gene deletion protocol based on a temperature sensitive knock-out plasmid T2-Ori was used to knock out the budC gene in B. licheniformis WX-02. The budC knock-out strain successfully abolished meso-2,3-butanediol production with enhanced D-2,3-butanediol production. No meso-BDH activity was detectable in cells of this strain. On the other hand, the complementary strain restored the characteristics of wild strain, and produced meso-2,3-butanediol and possessed meso-BDH activity. All of these data suggested that budC encoded the major meso-BDH catalyzing the reversible reaction from acetoin to meso-2,3-butanediol in B. licheniformis. The budC knock-out strain produced D-2,3-butanediol isomer only with a high yield of 30.76 g/L and a productivity of 1.28 g/L-h. Conclusions We confirmed the hypothesis that budC gene is responsible to reversibly transfer acetoin to meso-2,3-butanediol in B. licheniformis. A mutant strain of B. licheniformis with depleted budC gene was successfully developed and produced high level of the D-2,3-butanediol with high optimal purity. PMID:24475980

  10. BER-Myriant Succinic Acid Biorefinery

    Energy Technology Data Exchange (ETDEWEB)

    Shmorhun, Mark [Myriant Lake Providence, Inc., Lake Providence, LA (United States)

    2015-12-31

    Myriant Corporation (Myriant) has successfully produced the bioproduct succinic acid by the fermentation of glucose at a commercial scale operation in Lake Providence, Louisiana. The MySAB facility (Myriant Succinic Acid Biorefinery) came on stream in May 2013 and has been producing product since then. The MySAB facility is a demonstration-scale plant, capable of utilizing sorghum grits and commercially available dextrose, to ferment glucose into succinic acid. A downstream processing train has demonstrated the ability to produce an industrial, a standard and a polymer grade product. It consists of cell separation, membrane filtration, continuous chromatography, polishing to remove ionic and color bodies impurities, and final evaporation and crystallization. A by-product of the process is ammonium sulfate which is sold as a liquid fertilizer product. Since 2007 when development work began in the Woburn, Massachusetts R&D laboratories, the succinic acid bio-process has evolved through: Process development (microbiology, fermentation, and downstream) – R&D development laboratories; Piloting efforts at Fermic S.A. de C.V., Mexico City, Mexico – upstream and downstream processes; Design, construction, commissioning, and commercial production operations at the MySAB facility Additionally, Myriant became a wholly-owned subsidiary of the PTT Global Chemical Plc., Thailand, in late 2015, their investment into and support of Myriant goes back to 2011. The support of PTT Global Chemical Plc. helped to improve the upstream and downstream processes, and produce significant metric ton quantities of high quality bio-based succinic acid. The product has gone into a number of commercial markets worldwide for customer applications, development and production. The experience base gained via operations at the MySAB facility since May 2013, along with continued R&D development efforts involving Microbiology, Fermentation, and Downstream processes, at both the Woburn, Massachusetts

  11. Comparison of Spectrophotometry, Chromate Inhibition, and Cytofluorometry Versus Gene Sequencing for Detection of Heterozygously Glucose-6-Phosphate Dehydrogenase-Deficient Females.

    Science.gov (United States)

    Peters, Anna L; Veldthuis, Martijn; van Leeuwen, Karin; Bossuyt, Patrick M M; Vlaar, Alexander P J; van Bruggen, Robin; de Korte, Dirk; Van Noorden, Cornelis J F; van Zwieten, Rob

    2017-11-01

    Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme deficiency worldwide. Detection of heterozygously deficient females can be difficult as residual activity in G6PD-sufficient red blood cells (RBCs) can mask deficiency. In this study, we compared accuracy of 4 methods for detection of G6PD deficiency in females. Blood samples from females more than 3 months of age were used for spectrophotometric measurement of G6PD activity and for determination of the percentage G6PD-negative RBCs by cytofluorometry. An additional sample from females suspected to have G6PD deficiency based on the spectrophotometric G6PD activity was used for measuring chromate inhibition and sequencing of the G6PD gene. Of 165 included females, 114 were suspected to have heterozygous deficiency. From 75 females, an extra sample was obtained. In this group, mutation analysis detected 27 heterozygously deficient females. The sensitivity of spectrophotometry, cytofluorometry, and chromate inhibition was calculated to be 0.52 (confidence interval [CI]: 0.32-0.71), 0.85 (CI: 0.66-0.96), and 0.96 (CI: 0.71-1.00, respectively, and the specificity was 1.00 (CI: 0.93-1.00), 0.88 (CI: 0.75-0.95), and 0.98 (CI: 0.89-1.00), respectively. Heterozygously G6PD-deficient females with a larger percentage of G6PD-sufficient RBCs are missed by routine methods measuring total G6PD activity. However, the majority of these females can be detected with both chromate inhibition and cytofluorometry.

  12. Microbiota-Produced Succinate Improves Glucose Homeostasis via Intestinal Gluconeogenesis

    DEFF Research Database (Denmark)

    De Vadder, Filipe; Kovatcheva-Datchary, Petia; Zitoun, Carine

    2016-01-01

    Beneficial effects of dietary fiber on glucose and energy homeostasis have long been described, focusing mostly on the production of short-chain fatty acids by the gut commensal bacteria. However, bacterial fermentation of dietary fiber also produces large amounts of succinate and, to date...... homeostasis. Accordingly, dietary succinate improved glucose and insulin tolerance in wild-type mice, but those effects were absent in mice deficient in IGN. Conventional mice colonized with the succinate producer Prevotella copri exhibited metabolic benefits, which could be related to succinate-activated IGN...

  13. Redox engineering by ectopic expression of glutamate dehydrogenase genes links NADPH availability and NADH oxidation with cold growth in Saccharomyces cerevisiae.

    Science.gov (United States)

    Ballester-Tomás, Lidia; Randez-Gil, Francisca; Pérez-Torrado, Roberto; Prieto, Jose Antonio

    2015-07-09

    Cold stress reduces microbial growth and metabolism being relevant in industrial processes like wine making and brewing. Knowledge on the cold transcriptional response of Saccharomyces cerevisiae suggests the need of a proper redox balance. Nevertheless, there are no direct evidence of the links between NAD(P) levels and cold growth and how engineering of enzymatic reactions requiring NAD(P) may be used to modify the performance of industrial strains at low temperature. Recombinant strains of S. cerevisiae modified for increased NADPH- and NADH-dependent Gdh1 and Gdh2 activity were tested for growth at low temperature. A high-copy number of the GDH2-encoded glutamate dehydrogenase gene stimulated growth at 15°C, while overexpression of GDH1 had detrimental effects, a difference likely caused by cofactor preferences. Indeed, neither the Trp(-) character of the tested strains, which could affect the synthesis of NAD(P), nor changes in oxidative stress susceptibility by overexpression of GDH1 and GDH2 account for the observed phenotypes. However, increased or reduced NADPH availability by knock-out or overexpression of GRE3, the NADPH-dependent aldose reductase gene, eliminated or exacerbated the cold-growth defect observed in YEpGDH1 cells. We also demonstrated that decreased capacity of glycerol production impairs growth at 15 but not at 30°C and that 15°C-grown baker's yeast cells display higher fermentative capacity than those cultivated at 30°C. Thus, increasing NADH oxidation by overexpression of GDH2 would help to avoid perturbations in the redox metabolism induced by a higher fermentative/oxidative balance at low temperature. Finally, it is shown that overexpression of GDH2 increases notably the cold growth in the wine yeast strain QA23 in both standard growth medium and synthetic grape must. Redox constraints limit the growth of S. cerevisiae at temperatures below the optimal. An adequate supply of NAD(P) precursors as well as a proper level of reducing

  14. [Clinical features and ACADVL gene mutation spectrum analysis of 11 Chinese patients with very long chain acyl-CoA dehydrogenase deficiency].

    Science.gov (United States)

    Jinjun, Cao; Wenjuan, Qiu; Ruinan, Zhang; Jun, Ye; Lianshu, Han; Huiwen, Zhang; Qigang, Zhang; Xuefan, Gu

    2015-04-01

    To investigate the clinical and laboratory features of very long chain acyl-CoA dehydrogenase deficiency ( VLCADD ) and the correlations between its genotype and phenotype. Eleven patients diagnosed as VLCADD of Shanghai Jiaotong University School of Medicine seen from September 2006 to May 2014 were included. There were 9 boys and 2 girls, whose age was 2 d-17 years. Analysis was performed on clinical features, routine laboratory examination, and tandem mass spectrometry (MS-MS) , gas chromatography mass spectrometry (GC-MS) and genetic analysis were conducted. All cases had elevated levels of blood tetradecanoylcarnitine (C14:1) recognized as the characteristic biomarker for VLCADD. The eleven patients were classified into three groups: six cases in neonatal onset group, three in infancy onset group form patients and two in late onset group. Neonatal onset patients were characterized by hypoactivity, hypoglycemia shortly after birth. Infancy onset patients presented hepatomegaly and hypoglycemia in infancy. The two adolescent patients showed initial manifestations of exercise intolerance or rhabdomyolysis. Six of the eleven patients died at the age of 2-8 months, including four neonatal onset and two infant onset patients, with one or two null mutations. The other two neonatal onset patients were diagnosed since early birth through neonatal screening and their clinical manifestation are almost normal after treatments. Among 11 patients, seventeen different mutations in the ACADVL gene were identified, with a total mutation detection rate of 95.45% (21/22 alleles), including eleven reported mutations ( p. S22X, p. G43D, p. R511Q, p. W427X, p. A213T, p. C215R, p. G222R, p. R450H, p. R456H, c. 296-297delCA, c. 1605 + 1G > T) and six novel mutations (p. S72F, p. Q100X, p. M437T, p. D466Y, c. 1315delG insAC, IVS7 + 4 A > G). The p. R450H was the most frequent mutation identified in three alleles (13.63%, 3/22 alleles), followed by p. S22X and p. D466Y mutations which

  15. 78 FR 76567 - Tall Oil, Polymer With Polyethylene Glycol and Succinic Anhydride Monopolyisobutylene Derivs...

    Science.gov (United States)

    2013-12-18

    ... AGENCY 40 CFR Part 180 Tall Oil, Polymer With Polyethylene Glycol and Succinic Anhydride... oil, polymer with polyethylene glycol and succinic anhydride monopolyisobutylene derivs. (CAS Reg. No... residues of tall oil, polymer with polyethylene glycol and succinic anhydride monopolyisobutylene derivs...

  16. Gene-environment interactions on the risk of esophageal cancer among Asian populations with the G48A polymorphism in the alcohol dehydrogenase-2 gene: a meta-analysis.

    Science.gov (United States)

    Zhang, Long; Jiang, Yingjiu; Wu, Qingcheng; Li, Qiang; Chen, Dan; Xu, Ling; Zhang, Cheng; Zhang, Min; Ye, Ling

    2014-05-01

    The aim of this study is to investigate the gene-environment interactions between the G48A polymorphism in the alcohol dehydrogenase-2 (ADH2) gene and environmental factors in determining the risk of esophageal cancer (EC). A literature search was conducted in the PubMed, Embase, Web of Science, Cochrane Library, and Google Scholar databases to indentify eligible studies published before November 1, 2013. We performed a meta-analysis of 18 case-control studies with a total of 8,906 EC patients and 13,712 controls. The overall analysis suggested that individuals with the GG genotype were associated with a 2.77-fold increased risk of EC, compared with carriers of the GA and AA genotypes. In a stratified analysis by ethnic group, Japanese, Mainland Chinese, and Taiwan Chinese with the GG genotype had a significantly higher risk of EC, compared with Thai and Iranian populations, indicating ethnic variance in EC susceptibility. An analysis of combined effect indicated that GG genotype of ADH2 G48A was associated with the highest risk of EC in heavy drinkers and smokers. A striking difference was found to exist between males and females, showing gender variance for the association between ADH2 G48A and EC risk. This meta-analysis shows that the GG genotype of ADH2 G48A may be associated with an increased risk of EC in Asian populations. In addition, significant gene-environment interactions were found. Heavy drinkers, smokers, and males with the GG genotype may have a higher EC risk. Thus, our results shed new light on the complex gene-environment interactions that exist between environmental factors and ADH2 G48A polymorphism in EC risk.

  17. Spurious cooperativity in alkylated succinic acids

    Science.gov (United States)

    Ben-Naim, A.

    1998-03-01

    The proton-proton correlation, as measured by the ratio between the second and the first dissociation constants of dibasic acid, is sometimes very large and far beyond what could be explained by electrostatic theories. We propose a novel interpretation of this phenomenon based on the idea of spurious cooperativity. The general theoretical framework underlying the onset of spurious cooperativity is developed first. The basic result is that whenever a binding (or dissociating) two-site (or more) system splits into a mixture of noninterconverting isomers the binding isotherm (or the titration curve) behaves as if it is more negatively cooperative compared with the genuine cooperativities of the individual isomer. The theory is applied to a specific system of α-α' dialkyl succinic acid. It is known that the Meso form of these alkylated derivatives show a normal correlation of the same order of magnitude as in succinic acid. On the other hand, the Racemic form of these alkylated derivatives shows anomalous strong negative correlations when the alkyl groups become large (e.g., isopropyl and tert butyl). It is shown that the theory of spurious cooperativity can explain the different behavior of the Racemic and the Meso forms, as well as the onset of anomalous strong negative correlations when the alkyl groups become large.

  18. TREATMENT OF CHRONIC HEART FAILURE: FOCUS ON METOPROLOL SUCCINATE

    Directory of Open Access Journals (Sweden)

    O. D. Ostroumova

    2015-12-01

    Full Text Available Advantages of metoprolol succinate in patients with chronic heart failure (CHF are covered. Results of MERIT-HF study are taken as the main evidences. Patterns of the metoprolol succinate use in the treatment of different categories of patients with CHF (women, the elderly , severe CHF forms, CHF with concomitant hypertension or diabetes are considered.

  19. Selective hydrodeoxygenation of tartaric acid to succinic acid

    Energy Technology Data Exchange (ETDEWEB)

    Fu, Jiayi [Department of Chemical and Biomolecular Engineering; University of Delaware; Newark; USA; Catalysis Center for Energy Innovation; Vasiliadou, Efterpi S. [Catalysis Center for Energy Innovation; University of Delaware; Newark; USA; Goulas, Konstantinos A. [Catalysis Center for Energy Innovation; University of Delaware; Newark; USA; Saha, Basudeb [Catalysis Center for Energy Innovation; University of Delaware; Newark; USA; Vlachos, Dionisios G. [Department of Chemical and Biomolecular Engineering; University of Delaware; Newark; USA; Catalysis Center for Energy Innovation

    2017-01-01

    A novel one-step process for the selective production of succinic acid from tartaric acid is developed. High succinic yield is achieved in an efficient catalytic system comprised of MoOx/BC, HBr and acetic acid under hydrogen atmosphere.

  20. Microbiota-Produced Succinate Improves Glucose Homeostasis via Intestinal Gluconeogenesis.

    Science.gov (United States)

    De Vadder, Filipe; Kovatcheva-Datchary, Petia; Zitoun, Carine; Duchampt, Adeline; Bäckhed, Fredrik; Mithieux, Gilles

    2016-07-12

    Beneficial effects of dietary fiber on glucose and energy homeostasis have long been described, focusing mostly on the production of short-chain fatty acids by the gut commensal bacteria. However, bacterial fermentation of dietary fiber also produces large amounts of succinate and, to date, no study has focused on the role of succinate on host metabolism. Here, we fed mice a fiber-rich diet and found that succinate was the most abundant carboxylic acid in the cecum. Dietary succinate was identified as a substrate for intestinal gluconeogenesis (IGN), a process that improves glucose homeostasis. Accordingly, dietary succinate improved glucose and insulin tolerance in wild-type mice, but those effects were absent in mice deficient in IGN. Conventional mice colonized with the succinate producer Prevotella copri exhibited metabolic benefits, which could be related to succinate-activated IGN. Thus, microbiota-produced succinate is a previously unsuspected bacterial metabolite improving glycemic control through activation of IGN. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Structural and magnetic studies on copper succinate dihydrate ...

    Indian Academy of Sciences (India)

    M P BINITHA

    2017-08-21

    Aug 21, 2017 ... Abstract. Single crystals of copper succinate dihydrate were grown in silica gel by slow diffusion of copper chloride to sodium metasilicate gel impregnated with succinic acid. The grown crystal was subjected to single crystal X-ray diffraction studies. In its structure each copper atom is penta co-ordinated to ...

  2. Reduction of experimental myocardial infarct size by combined treatment with methylprednisolone sodium succinate and betahistine hydrochloride.

    Science.gov (United States)

    Joyce, L D; Smith, J M; Mauer, H G; Lillehei, R C

    1978-01-01

    This laboratory and others have shown that methylprednisolone sodium succinate and betahistine hydrochloride can each reduce the size of experimental myocardial infarct in dogs at six hours. In light of the fact that these two agents probably act via different mechanisms, a study was carried out to determine if there would be cumulative effects of using these two agents together. Using a left anterior descending coronary artery ligation model to create an experimental myocardial infarction and an intracellular lactic dehydrogenase (LDH) stain to measure the infarct size, 66 dogs were studied. Nineteen dogs served as controls with no therapy; 20 received a continuous intravenous infusion of betahistine hydrochloride (0.24 mg/kg/min) for six hours following ligation; 10 dogs received methylprednisolone sodium succinate intravenously 30 mg/kg at one hour postligation; and 17 other dogs received betahistine hydrochloride intravenously (0.24 mg/kg/min) over six hours following ligation, plus methylprednisolone sodium succinate intravenously (30 mg/kg) at one hour postligation. At six hours, the combined treatment demonstrated no significant improvement over betahistine HCl alone (control, 16.0%; betahistine HCl, 11.4%; combined, 11.2% P less than 0.05). At 24 hours, only the combined treatment group demonstrated a significant infarct size reduction (control, 15.5%; methylprednisolone, 13.1%; betahistine--HCl, 14.2%; combined, 9.7%; P less than 0.0025). Other parameters that were evaluated and analyzed include mean aortic pressure, left atrial pressure, cardiac index, total peripheral resistance, arterial and coronary sinus pH, pCO2, pO2, hematocrit, O2 consumption, O2 content difference, and coronary sinus lactate and creatine phosphokinase (CPK). These results suggest a significant cumulative effect in reducing infarct size over that achieved with one agent alone; however, further studies are needed to determine the appropriate dosage and temporal factors.

  3. Comparison of Spectrophotometry, Chromate Inhibition, and Cytofluorometry Versus Gene Sequencing for Detection of Heterozygously Glucose-6-Phosphate Dehydrogenase-Deficient Females

    NARCIS (Netherlands)

    Peters, Anna L.; Veldthuis, Martijn; van Leeuwen, Karin; Bossuyt, Patrick M. M.; Vlaar, Alexander P. J.; van Bruggen, Robin; de Korte, Dirk; van Noorden, Cornelis J. F.; van Zwieten, Rob

    2017-01-01

    Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme deficiency worldwide. Detection of heterozygously deficient females can be difficult as residual activity in G6PD-sufficient red blood cells (RBCs) can mask deficiency. In this study, we compared accuracy of 4 methods for

  4. Role of Hfq in iron-dependent and -independent gene regulation in Neisseria meningitidis.

    Science.gov (United States)

    Mellin, J R; McClure, Ryan; Lopez, Delia; Green, Olivia; Reinhard, Bjorn; Genco, Caroline

    2010-08-01

    In Neisseria meningitidis, iron-responsive gene regulation is mediated primarily by the ferric uptake regulator (Fur) protein. When complexed with iron, Fur represses gene expression by preventing transcription initiation. Fur can also indirectly activate gene expression via the repression of regulatory small RNAs (sRNA). One such Fur- and iron-regulated sRNA, NrrF, was previously identified in N. meningitidis and shown to repress expression of the sdhA and sdhC genes encoding subunits of the succinate dehydrogenase complex. In the majority of Gram-negative bacteria, sRNA-mediated regulation requires a cofactor RNA-binding protein (Hfq) for proper gene regulation and stabilization. In this study, we examined the role of Hfq in NrrF-mediated regulation of the succinate dehydrogenase genes in N. meningitidis and the effect of an hfq mutation on iron-responsive gene regulation more broadly. We first demonstrated that the stability of NrrF, as well as the regulation of sdhC and sdhA in vivo, was unaltered in the hfq mutant. Secondly, we established that iron-responsive gene regulation of the Fur-regulated sodB gene was dependent on Hfq. Finally, we demonstrated that in N. meningitidis, Hfq functions in a global manner to control expression of many ORFs and intergenic regions via iron-independent mechanisms. Collectively these studies demonstrate that in N. meningitidis, iron- and NrrF-mediated regulation of sdhC and sdhA can occur independently of Hfq, although Hfq functions more globally to control regulation of other N. meningitidis genes primarily by iron-independent mechanisms.

  5. Enhanced succinic acid production in Aspergillus saccharolyticus by heterologous expression of fumarate reductase from Trypanosoma brucei

    DEFF Research Database (Denmark)

    Yang, Lei; Lübeck, Mette; Ahring, Birgitte K.

    2015-01-01

    of the enzymes, pyruvate carboxylase (PYC), malate dehydrogenase (MDH), and fumarase (FUM), involved in the reductive tricarboxylic acid (rTCA) branch, were examined and compared in cells harvested from the acid production medium and a complete medium. The results showed that ambient pH had a significant impact...... on the pattern and the amount of organic acids produced by A. saccharolyticus. The wild-type strain produced higher amount of malic acid and succinic acid in the pH buffered condition (pH 6.5) compared with the pH non-buffered condition. The enzyme assays showed that the rTCA branch was active in the acid...

  6. Biodegradable polyesters based on succinic acid

    Directory of Open Access Journals (Sweden)

    Nikolić Marija S.

    2003-01-01

    Full Text Available Two series of aliphatic polyesters based on succinic acid were synthesized by copolymerization with adipic acid for the first series of saturated polyesters, and with fumaric acid for the second series. Polyesters were prepared starting from the corresponding dimethyl esters and 1,4-butanediol by melt transesterification in the presence of a highly effective catalyst tetra-n-butyl-titanate, Ti(0Bu4. The molecular structure and composition of the copolyesters was determined by 1H NMR spectroscopy. The effect of copolymer composition on the physical and thermal properties of these random polyesters were investigated using differential scanning calorimetry. The degree of crystallinity was determined by DSC and wide angle X-ray. The degrees of crystallinity of the saturated and unsaturated copolyesters were generally reduced with respect to poly(butylene succinate, PBS. The melting temperatures of the saturated polyesters were lower, while the melting temperatures of the unsaturated copolyesters were higher than the melting temperature of PBS. The biodegradability of the polyesters was investigated by enzymatic degradation tests. The enzymatic degradation tests were performed in a buffer solution with Candida cylindracea lipase and for the unsaturated polyesters with Rhizopus arrhizus lipase. The extent of biodegradation was quantified as the weight loss of polyester films. Also the surface of the polyester films after degradation was observed using optical microscopy. It could be concluded that the biodegradability depended strongly on the degree of crystallinity, but also on the flexibility of the chain backbone. The highest biodegradation was observed for copolyesters containing 50 mol.% of adipic acid units, and in the series of unsaturated polyesters for copolyesters containing 5 and 10 mol.% of fumarate units. Although the degree of crystallinity of the unsaturated polyesters decreased slightly with increasing unsaturation, the biodegradation

  7. E1 of α-ketoglutarate dehydrogenase defends Mycobacterium tuberculosis against glutamate anaplerosis and nitroxidative stress.

    Science.gov (United States)

    Maksymiuk, Christina; Balakrishnan, Anand; Bryk, Ruslana; Rhee, Kyu Y; Nathan, Carl F

    2015-10-27

    Enzymes of central carbon metabolism (CCM) in Mycobacterium tuberculosis (Mtb) make an important contribution to the pathogen's virulence. Evidence is emerging that some of these enzymes are not simply playing the metabolic roles for which they are annotated, but can protect the pathogen via additional functions. Here, we found that deficiency of 2-hydroxy-3-oxoadipate synthase (HOAS), the E1 component of the α-ketoglutarate (α-KG) dehydrogenase complex (KDHC), did not lead to general metabolic perturbation or growth impairment of Mtb, but only to the specific inability to cope with glutamate anaplerosis and nitroxidative stress. In the former role, HOAS acts to prevent accumulation of aldehydes, including growth-inhibitory succinate semialdehyde (SSA). In the latter role, HOAS can participate in an alternative four-component peroxidase system, HOAS/dihydrolipoyl acetyl transferase (DlaT)/alkylhydroperoxide reductase colorless subunit gene (ahpC)-neighboring subunit (AhpD)/AhpC, using α-KG as a previously undescribed source of electrons for reductase action. Thus, instead of a canonical role in CCM, the E1 component of Mtb's KDHC serves key roles in situational defense that contribute to its requirement for virulence in the host. We also show that pyruvate decarboxylase (AceE), the E1 component of pyruvate dehydrogenase (PDHC), can participate in AceE/DlaT/AhpD/AhpC, using pyruvate as a source of electrons for reductase action. Identification of these systems leads us to suggest that Mtb can recruit components of its CCM for reactive nitrogen defense using central carbon metabolites.

  8. Succinic Semialdehyde Promotes Prosurvival Capability of Agrobacterium tumefaciens.

    Science.gov (United States)

    Wang, Chao; Tang, Desong; Gao, Yong-Gui; Zhang, Lian-Hui

    2016-01-11

    Succinic semialdehyde (SSA), an important metabolite of γ-aminobutyric acid (GABA), is a ligand of the repressor AttJ regulating the expression of the attJ-attKLM gene cluster in the plant pathogen Agrobacterium tumefaciens. While the response of A. tumefaciens to GABA and the function of attKLM have been extensively studied, genetic and physiological responses of A. tumefaciens to SSA remain unknown. In combination with microarray and genetic approaches, this study sets out to explore new roles of the SSA-AttJKLM regulatory mechanism during bacterial infection. The results showed that SSA plays a key role in regulation of several bacterial activities, including C4-dicarboxylate utilization, nitrate assimilation, and resistance to oxidative stress. Interestingly, while the SSA relies heavily on the functional AttKLM in mediating nitrate assimilation and oxidative stress resistance, the compound could regulate utilization of C4-dicarboxylates independent of AttJKLM. We further provide evidence that SSA controls C4-dicarboxylate utilization through induction of an SSA importer and that disruption of attKLM attenuates the tumorigenicity of A. tumefaciens. Taken together, these findings indicate that SSA could be a potent plant signal which, together with AttKLM, plays a vital role in promoting the bacterial prosurvival abilities during infection. Agrobacterium tumefaciens is a plant pathogen causing crown gall diseases and has been well known as a powerful tool for plant genetic engineering. During the long history of microbe-host interaction, A. tumefaciens has evolved the capabilities of recognition and response to plant-derived chemical metabolites. Succinic semialdehyde (SSA) is one such metabolite. Previous results have demonstrated that SSA functions to activate a quorum-quenching mechanism and thus to decrease the level of quorum-sensing signals, thereby avoiding the elicitation of a plant defense. Here, we studied the effect of SSA on gene expression at a

  9. Molecular cloning of three pyranose dehydrogenase-encoding genes from Agaricus meleagris and analysis of their expression by real-time RT-PCR

    Czech Academy of Sciences Publication Activity Database

    Kittl, R.; Sygmund, Ch.; Halada, Petr; Volc, Jindřich; Divne, Ch.; Haltrich, D.; Peterbauer, C.

    2008-01-01

    Roč. 53, č. 2 (2008), s. 117-127 ISSN 0172-8083 R&D Projects: GA MŠk LC545 Grant - others:GA MŠk(CZ) Kontakt 6-06-4 Institutional research plan: CEZ:AV0Z50200510 Keywords : pyranose dehydrogenase * lignocellulose degradation * agarices spp Subject RIV: EE - Microbiology, Virology Impact factor: 2.323, year: 2008

  10. Genes for selenium dependent and independent formate dehydrogenase in the gut microbial communities of three lower, wood-feeding termites and a wood-feeding roach

    OpenAIRE

    Zhang, Xinning; Matson, Eric G.; Leadbetter, Jared R.

    2011-01-01

    The bacterial Wood-Ljungdahl pathway for CO_2-reductive acetogenesis is important for the nutritional mutualism occurring between wood-feeding insects and their hindgut microbiota. A key step in this pathway is the reduction of CO_2 to formate, catalysed by the enzyme formate dehydrogenase (FDH). Putative selenocysteine- (Sec) and cysteine- (Cys) containing paralogues of hydrogenase-linked FDH (FDH_H) have been identified in the termite gut acetogenic spirochet...

  11. Evaluation of housekeeping genes for quantitative gene expression analysis in the equine kidney.

    Science.gov (United States)

    Azarpeykan, Sara; Dittmer, Keren E

    2016-01-01

    Housekeeping genes (HKGs) are used as internal controls for normalising and calculating the relative expression of target genes in RT-qPCR experiments. There is no unique universal HKG and HKGs vary among organisms and tissues, so this study aimed to determine the most stably expressed HKGs in the equine kidney. The evaluated HKGs included 18S ribosomal RNA (18S), 28S ribosomal RNA (28S), ribosomal protein L32 (RPL32), β-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), succinate dehydrogenase complex (SDHA), zeta polypeptide (YWHAZ), and hypoxanthine phosphoribosyltransferase 1 (HPRT1). The HKGs expression stability data were analysed with two software packages, geNorm and NormFinder. The lowest stability values for geNorm suggests that YWHAZ and HPRT1 would be most optimal (M=0.31 and 0.32, respectively). Further, these two genes had the best pairwise stability value using NormFinder (geNorm V=0.085). Therefore, these two genes were considered the most useful for RT-qPCR studies in equine kidney.

  12. Malaria Protection In Glucose-6-Phosphate Dehydrogenase ...

    African Journals Online (AJOL)

    The high frequency of glucose-6-phosphate dehydrogenase (G6PD) deficiency gene in malaria endemic regions is believed to be due to the enzyme deficiency advantage against fatal malaria. However, the mechanism of this protection is not well understood and therefore was investigated by comparing differences in ...

  13. Gene structure and mutations of glutaryl-coenzyme A dehydrogenase: impaired association of enzyme subunits that is due to an A421V substitution causes glutaric acidemia type I in the Amish.

    Science.gov (United States)

    Biery, B J; Stein, D E; Morton, D H; Goodman, S I

    1996-11-01

    The structure of the human glutaryl coenzyme A dehydrogenase (GCD) gene was determined to contain 11 exons and to span approximately 7 kb. Fibroblast DNA from 64 unrelated glutaric acidemia type I (GA1) patients was screened for mutations by PCR amplification and analysis of SSCP. Fragments with altered electrophoretic mobility were subcloned and sequenced to detect mutations that caused GA1. This report describes the structure of the GCD gene, as well as point mutations and polymorphisms found in 7 of its 11 exons. Several mutations were found in more than one patient, but no one prevalent mutation was detected in the general population. As expected from pedigree analysis, a single mutant allele causes GA1 in the Old Order Amish of Lancaster County, Pennsylvania. Several mutations have been expressed in Escherichia coli, and all produce diminished enzyme activity. Reduced activity in GCD encoded by the A421V mutation in the Amish may be due to impaired association of enzyme subunits.

  14. The ACADS gene variation spectrum in 114 patients with short-chain acyl-CoA dehydrogenase (SCAD) deficiency is dominated by missense variations leading to protein misfolding at the cellular level

    DEFF Research Database (Denmark)

    Pedersen, Christina Bak; Kølvrå, Steen; Kølvraa, Agnete

    2008-01-01

    Short-chain acyl-CoA dehydrogenase (SCAD) deficiency is an inherited disorder of mitochondrial fatty acid oxidation associated with variations in the ACADS gene and variable clinical symptoms. In addition to rare ACADS inactivating variations, two common variations, c.511C > T (p.Arg171Trp) and c.......625G > A (p.Gly209Ser), have been identified in patients, but these are also present in up to 14% of normal populations leading to questions of their clinical relevance. The common variant alleles encode proteins with nearly normal enzymatic activity at physiological conditions in vitro. SCAD enzyme...... function, however, is impaired at increased temperature and the tendency to misfold increases under conditions of cellular stress. The present study examines misfolding of variant SCAD proteins identified in patients with SCAD deficiency. Analysis of the ACADS gene in 114 patients revealed 29 variations...

  15. An immunohistochemical procedure to detect patients with paraganglioma and phaeochromocytoma with germline SDHB, SDHC, or SDHD gene mutations: a retrospective and prospective analysis

    NARCIS (Netherlands)

    F.H. van Nederveen (Francien); J. Gaal (José); J. Favier (Judith); E. Korpershoek (Esther); R.A. Oldenburg (Rogier); E.M. de Bruyn (Elly); H.F. Sleddens (Hein); P. Derkx (Pieter); J. Rivière (Julie); H. Dannenberg (Hilde); B.J. Petri; P. Komminoth (Paul); K. Pacak (Karel); W.C.J. Hop (Wim); P.J. Pollard (Patrick); M. Mannelli (Massimo); J.P. Bayley; A. Perren (Aurel); S. Niemann (Stephan); A.A. Verhofstad (Albert); A.P. de Bruïne (Adriaan); E.R. Maher (Eamonn); F. Tissier (Frédérique); T. Méatchi (Tchao); C. Badoual (Cécile); J. Bertherat (Jerome); L. Amar (Laurence); D. Alataki (Despoina); E.A. van Marck (Eric); F. Ferrau (Francesco); J.F. François (Joseph); W.W. de Herder (Wouter); M.P.F.V. Peeters; A. van Linge (Anne); J.W. Lenders (Jacques); A.P. Gimenez-Roqueplo; R.R. de Krijger (Ronald); W.N.M. Dinjens (Winand)

    2009-01-01

    textabstractBackground: Phaeochromocytomas and paragangliomas are neuro-endocrine tumours that occur sporadically and in several hereditary tumour syndromes, including the phaeochromocytoma-paraganglioma syndrome. This syndrome is caused by germline mutations in succinate dehydrogenase B (SDHB), C

  16. Homozygous mutation, p.Pro304His, in IDH3A, encoding isocitrate dehydrogenase subunit is associated with severe encephalopathy in infancy.

    Science.gov (United States)

    Fattal-Valevski, Aviva; Eliyahu, Hila; Fraenkel, NItai D; Elmaliach, Ganit; Hausman-Kedem, Moran; Shaag, Avraham; Mandel, Dror; Pines, Ophry; Elpeleg, Orly

    2017-01-01

    Mitochondrial encephalopathies are a heterogeneous group of disorders which generally carries a grave prognosis. Using exome sequencing, we identified a homozygous mutation, Pro-304-His in the IDH3A gene, in a patient suffering from infantile encephalopathy with peripheral and autonomic nervous system involvement. Mammalian isocitrate dehydrogenase (IDH) 3 is a heterotetramer of 2alfa, 1beta, and 1gamma subunits, and IDH3A encodes the alfa subunit of the mitochondrial NAD + -dependent IDH. Here we show that in contrast to wild-type human IDH3A, the human IDH3A which harbor the p.Pro304His mutation does not complement the yeast Δidh1/Δidh2 growth defect on ethanol-acetate. We therefore propose that homozygosity for the p.Pro304His mutation is deleterious for mitochondrial NAD + -specific IDH3A activity in human. IDH3A now joins the list of TCA cycle-related proteins, which includes ACO2, DLD, SLC25A19, FH, and succinate dehydrogenase subunits, all associated with neurological disorders.

  17. Hydroxypropyl Methylcellulose Acetate Succinate: Potential Drug–Excipient Incompatibility

    OpenAIRE

    Dong, Zedong; Choi, Duk Soon

    2008-01-01

    The stability of hydroxypropyl methylcellulose acetate succinate (HPMC-AS) and its potential incompatibility with active pharmaceutical ingredients (API) carrying hydroxyl group(s) were investigated in this research. HPMC-AS may undergo hydrolysis under harsh processing conditions with the generation of succinic acid and acetic acid, which can form ester bond(s) with the hydroxyl group(s) in API. In this case, the hot-melt extrusion (HME) product prepared from HPMC-AS and our model compound (...

  18. Pyruvate dehydrogenase complex regulator (PdhR) gene deletion boosts glucose metabolism in Escherichia coli under oxygen-limited culture conditions.

    Science.gov (United States)

    Maeda, Soya; Shimizu, Kumiko; Kihira, Chie; Iwabu, Yuki; Kato, Ryuichi; Sugimoto, Makoto; Fukiya, Satoru; Wada, Masaru; Yokota, Atsushi

    2017-04-01

    Pyruvate dehydrogenase complex regulator (PdhR) is a transcriptional regulator that negatively regulates formation of pyruvate dehydrogenase complex (PDHc), NADH dehydrogenase (NDH)-2, and cytochrome bo 3 oxidase in Escherichia coli. To investigate the effects of a PdhR defect on glucose metabolism, a pdhR deletion mutant was derived from the wild-type E. coli W1485 strain by λ Red-mediated recombination. While no difference in the fermentation profiles was observed between the two strains under oxygen-sufficient conditions, under oxygen-limited conditions, the growth level of the wild-type strain was significantly decreased with retarded glucose consumption accompanied by by-production of substantial amounts of pyruvic acid and acetic acid. In contrast, the mutant grew and consumed glucose more efficiently than did the wild-type strain with enhanced respiration, little by-production of pyruvic acid, less production yield and rates of acetic acid, thus displaying robust metabolic activity. As expected, increased activities of PDHc and NDH-2 were observed in the mutant. The increased activity of PDHc may explain the loss of pyruvic acid by-production, probably leading to decreased acetic acid formation, and the increased activity of NDH-2 may explain the enhanced respiration. Measurement of the intracellular NAD + /NADH ratio in the mutant revealed more oxidative or more reductive intracellular environments than those in the wild-type strain under oxygen-sufficient and -limited conditions, respectively, suggesting another role of PdhR: maintaining redox balance in E. coli. The overall results demonstrate the biotechnological advantages of pdhR deletion in boosting glucose metabolism and also improve our understanding of the role of PdhR in bacterial physiology. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  19. 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...

  20. Differential levels of glutamate dehydrogenase 1 (GLUD1) in Balb/c and C57BL/6 mice and the effects of overexpression of the Glud1 gene on glutamate release in striatum.

    Science.gov (United States)

    Hascup, Kevin N; Bao, Xiaodong; Hascup, Erin R; Hui, Dongwei; Xu, Wenhao; Pomerleau, Francois; Huettl, Peter; Michaelis, Mary L; Michaelis, Elias K; Gerhardt, Greg A

    2011-04-21

    We have previously shown that overexpression of the Glud1 (glutamate dehydrogenase 1) gene in neurons of C57BL/6 mice results in increased depolarization-induced glutamate release that eventually leads to selective neuronal injury and cell loss by 12 months of age. However, it is known that isogenic lines of Tg (transgenic) mice produced through back-crossing with one strain may differ in their phenotypic characteristics from those produced using another inbred mouse strain. Therefore, we decided to introduce the Glud1 transgene into the Balb/c strain that has endogenously lower levels of GLUD1 (glutamate dehydrogenase 1) enzyme activity in the brain as compared with C57BL/6. Using an enzyme-based MEA (microelectrode array) that is selective for measuring glutamate in vivo, we measured depolarization-induced glutamate release. Within a discrete layer of the striatum, glutamate release was significantly increased in Balb/c Tg mice compared with wt (wild-type) littermates. Furthermore, Balb/c mice released approx. 50-60% of the amount of glutamate compared with C57BL/6 mice. This is similar to the lower levels of endogenous GLUD1 protein in Balb/c compared with C57BL/6 mice. The development of these Glud1-overexpressing mice may allow for the exploration of key molecular events produced by chronic exposure of neurons to moderate, transient increases in glutamate release, a process hypothesized to occur in neurodegenerative disorders.

  1. Simultaneous Expression of NAD-Dependent Isocitrate Dehydrogenase and Other Krebs Cycle Genes after Nitrate Resupply to Short-Term Nitrogen-Starved Tobacco

    Science.gov (United States)

    Lancien, Muriel; Ferrario-Méry, Sylvie; Roux, Yvette; Bismuth, Evelyne; Masclaux, Céline; Hirel, Bertrand; Gadal, Pierre; Hodges, Michael

    1999-01-01

    Mitochondrial NAD-dependent (IDH) and cytosolic NADP-dependent isocitrate dehydrogenases have been considered as candidates for the production of 2-oxoglutarate required by the glutamine synthetase/glutamate synthase cycle. The increase in IDH transcripts in leaf and root tissues, induced by nitrate or NH4+ resupply to short-term N-starved tobacco (Nicotiana tabacum) plants, suggested that this enzyme could play such a role. The leaf and root steady-state mRNA levels of citrate synthase, acotinase, IDH, and glutamine synthetase were found to respond similarly to nitrate, whereas those for cytosolic NADP-dependent isocitrate dehydrogenase and fumarase responded differently. This apparent coordination occurred only at the mRNA level, since activity and protein levels of certain corresponding enzymes were not altered. Roots and leaves were not affected to the same extent either by N starvation or nitrate addition, the roots showing smaller changes in N metabolite levels. After nitrate resupply, these organs showed different response kinetics with respect to mRNA and N metabolite levels, suggesting that under such conditions nitrate assimilation was preferentially carried out in the roots. The differential effects appeared to reflect the C/N status after N starvation, the response kinetics being associated with the nitrate assimilatory capacity of each organ, signaled either by nitrate status or by metabolite(s) associated with its metabolism. PMID:10398706

  2. Reference gene selection for qRT-PCR analysis in the sweetpotato whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae).

    Science.gov (United States)

    Li, Rumei; Xie, Wen; Wang, Shaoli; Wu, Qingjun; Yang, Nina; Yang, Xin; Pan, Huipeng; Zhou, Xiaomao; Bai, Lianyang; Xu, Baoyun; Zhou, Xuguo; Zhang, Youjun

    2013-01-01

    Accurate evaluation of gene expression requires normalization relative to the expression of reliable reference genes. Expression levels of "classical" reference genes can differ, however, across experimental conditions. Although quantitative real-time PCR (qRT-PCR) has been used extensively to decipher gene function in the sweetpotato whitefly Bemisia tabaci, a world-wide pest in many agricultural systems, the stability of its reference genes has rarely been validated. In this study, 15 candidate reference genes from B. tabaci were evaluated using two Excel-based algorithms geNorm and Normfinder under a diverse set of biotic and abiotic conditions. At least two reference genes were selected to normalize gene expressions in B. tabaci under experimental conditions. Specifically, for biotic conditions including host plant, acquisition of a plant virus, developmental stage, tissue (body region of the adult), and whitefly biotype, ribosomal protein L29 was the most stable reference gene. In contrast, the expression of elongation factor 1 alpha, peptidylprolyl isomerase A, NADH dehydrogenase, succinate dehydrogenase complex subunit A and heat shock protein 40 were consistently stable across various abiotic conditions including photoperiod, temperature, and insecticide susceptibility. Our finding is the first step toward establishing a standardized quantitative real-time PCR procedure following the MIQE (Minimum Information for publication of Quantitative real time PCR Experiments) guideline in an agriculturally important insect pest, and provides a solid foundation for future RNA interference based functional study in B. tabaci.

  3. Reference gene selection for qRT-PCR analysis in the sweetpotato whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae.

    Directory of Open Access Journals (Sweden)

    Rumei Li

    Full Text Available Accurate evaluation of gene expression requires normalization relative to the expression of reliable reference genes. Expression levels of "classical" reference genes can differ, however, across experimental conditions. Although quantitative real-time PCR (qRT-PCR has been used extensively to decipher gene function in the sweetpotato whitefly Bemisia tabaci, a world-wide pest in many agricultural systems, the stability of its reference genes has rarely been validated.In this study, 15 candidate reference genes from B. tabaci were evaluated using two Excel-based algorithms geNorm and Normfinder under a diverse set of biotic and abiotic conditions. At least two reference genes were selected to normalize gene expressions in B. tabaci under experimental conditions. Specifically, for biotic conditions including host plant, acquisition of a plant virus, developmental stage, tissue (body region of the adult, and whitefly biotype, ribosomal protein L29 was the most stable reference gene. In contrast, the expression of elongation factor 1 alpha, peptidylprolyl isomerase A, NADH dehydrogenase, succinate dehydrogenase complex subunit A and heat shock protein 40 were consistently stable across various abiotic conditions including photoperiod, temperature, and insecticide susceptibility.Our finding is the first step toward establishing a standardized quantitative real-time PCR procedure following the MIQE (Minimum Information for publication of Quantitative real time PCR Experiments guideline in an agriculturally important insect pest, and provides a solid foundation for future RNA interference based functional study in B. tabaci.

  4. Identification and characterization of genes related to the production of organic acids in yeast.

    Science.gov (United States)

    Yoshida, Satoshi; Yokoyama, Aki

    2012-05-01

    Organic acids contribute to the flavor of many foods and drinks including alcoholic beverages. To study the cellular processes affecting organic acid production, here we screened collections of Saccharomyces cerevisiae deletion mutants and identified 36 yeast mutants forming a yellow halo on YPD plates containing bromocresol purple, indicating that the pH of the medium had been lowered. The disrupted genes encoded TCA cycle enzymes, transcription factors, signal transducers, and ubiquitin-related proteins. Acetate, pyruvate, and succinate are produced by yeast fermentation in rich medium, and their production was affected by mutations of the genes GTR1, GTR2, LIP5, LSM1, PHO85, PLM2, RTG1, RTG2 and UBP3, and also succinate dehydrogenase-related genes including EMI5, SDH1, SDH2, SDH4, TCM62 and YDR379C-A. Among the genes identified, overexpression of only LIP5 affected the production of acetate in S. cerevisiae. However, overexpression of EMI5, LIP5, RTG2 and UBP3 had a significant effect on the production of acetate, citrate, lactate, and succinate in the bottom-fermenting yeast Saccharomyces pastorianus. Furthermore, phenotypic analysis of the S. cerevisiae disruptants involved in organic acid production showed that azaserine, citrate, ethionine, and sulfite are useful compounds by which mutants with altered organic acid production might be selected. Taken together, these results suggest that the regulation of many organic acids might be simultaneously achieved by activation or inactivation of a single gene. Copyright © 2012 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  5. Effect of Hydroxyl Monomers on the Enzymatic Degradation of Poly(ethylene succinate, Poly(butylene succinate, and Poly(hexylene succinate

    Directory of Open Access Journals (Sweden)

    Zhenhui Bai

    2018-01-01

    Full Text Available Poly(ethylene succinate (PES, poly(butylene succinate (PBS, and poly(hexylene succinate (PHS, were synthesized using succinic acid and different dihydric alcohols as materials. Enzymatic degradability by cutinase of the three kinds of polyesters was studied, as well as their solid-state properties. The biodegradation behavior relied heavily on the distance between ester groups, crystallinity, and the hydrophilicity-hydrophobicity balance of polyester surfaces. The weight loss through degradation of the three kinds of polyesters with different hydroxyl monomers took place in the order PHS > PBS > PES. The degradation behavior of the polyesters before and after degradation was analyzed by scanning electron microscopy, differential scanning calorimetry, powder X-ray diffraction, Fourier transform infrared spectroscopy, gel permeation chromatography, and thermogravimetric analysis. The decrease in relative intensity at 1800–1650 estedpolyesters were degraded simultaneously. The frequencies of the crystalline and amorphous bands were almost identical before and after degradation. Thus, enzymatic degradation did not change the crystalline structure but destroyed it, and the degree of crystallinity markedly decreased. The molecular weight and polydispersity index only changed slightly. The thermal stability of the three kinds of polyesters decreased during enzymatic degradation.

  6. Recombinant adeno-associated virus-mediated gene delivery of long chain acyl coenzyme A dehydrogenase (LCAD) into LCAD-deficient mice.

    Science.gov (United States)

    Beattie, Stuart G; Goetzman, Eric; Tang, Qiuishi; Conlon, Thomas; Campbell-Thompson, Martha; Matern, Dietrich; Vockley, Jerry; Flotte, Terence R

    2008-10-01

    Very long chain acyl coenzyme A (CoA) dehydrogenase (VLCAD) deficiency is a relatively common mitochondrial beta-oxidation disorder. The most severe form of VLCAD deficiency presents with neonatal cardiomyopathy and hepatic failure and is generally fatal within the first year of life. Mice deficient for long chain acyl CoA dehydrogenase (LCAD) closely resemble the clinical syndrome observed in VLCAD-deficient humans. Recombinant adeno-associated viral (rAAV) vectors with pseudotype capsids were investigated for their potential towards correcting the phenotype observed in mice heterozygous (+/-) for LCAD (i.e. liver and muscle steatosis). rAAV containing the mouse LCAD cDNA (mLCAD) under the transcriptional control of the CMV/chicken beta-actin hybrid promoter were injected intramuscularly into the tibialis anterior (TA) muscle of LCAD(+/-) mice or injected into the portal vein to transduce hepatocytes. Ten weeks post-injection of rAAV1-mLCAD into the TA muscle, significantly increased levels of mLCAD within mitochondria were demonstrated by immunostaining of TA sections, immunoblotting of mitochondrial isolates and by the electron transfer flavoprotein (ETF) fluorescence reduction enzyme activity assay. Magnetic resonance spectroscopy of vector-injected TA muscle demonstrated a reduction in the lipid content compared to phosphate-buffered saline-injected mice, whereas a systemic effect was observed as a reduction in liver macrosteatosis. Eight weeks after portal vein injection of rAAV8-mLCAD into LCAD(+/-) mice, increased levels of mLCAD within hepatocyte mitochondria were demonstrated by immunostaining and also by the ETF assay. Scoring of the hepatosteatosis observed in partially deficient LCAD mice indicated a reduction in the lipid content within livers of vector-treated mice. These studies show that rAAV-mediated delivery of mLCAD was efficient and led to an amelioration of local and systemic pathologies observed in partially deficient LCAD mice. Copyright (c

  7. 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

  8. A putrescine-inducible pathway comprising PuuE-YneI in which gamma-aminobutyrate is degraded into succinate in Escherichia coli K-12.

    Science.gov (United States)

    Kurihara, Shin; Kato, Kenji; Asada, Kei; Kumagai, Hidehiko; Suzuki, Hideyuki

    2010-09-01

    Gamma-aminobutyrate (GABA) is metabolized to succinic semialdehyde by GABA aminotransferase (GABA-AT), and the succinic semialdehyde is subsequently oxidized to succinate by succinic semialdehyde dehydrogenase (SSADH). In Escherichia coli, there are duplicate GABA-ATs (GabT and PuuE) and duplicate SSADHs (GabD and YneI). While GabT and GabD have been well studied previously, the characterization and expression analysis of PuuE and YneI are yet to be investigated. By analyzing the amino acid profiles in cells of DeltapuuE and/or DeltagabT mutants, this study demonstrated that PuuE plays an important role in GABA metabolism in E. coli cells. The similarity of the amino acid sequences of PuuE and GabT is 67.4%, and it was biochemically demonstrated that the catalytic center of GabT is conserved as an amino acid residue important for the enzymatic activity in PuuE as Lys-247. However, the regulation of expression of PuuE is significantly different from that of GabT. PuuE is induced by the addition of putrescine to the medium and is repressed by succinate and low aeration conditions; in contrast, GabT is almost constitutive. Similarly, YneI is induced by putrescine, while GabD is not. For E. coli, PuuE is important for utilization of putrescine as a sole nitrogen source and both PuuE and YneI are important for utilization of putrescine as a sole carbon source. The results demonstrate that the PuuE-YneI pathway was a putrescine-inducible GABA degradation pathway for utilizing putrescine as a nutrient source.

  9. A Putrescine-Inducible Pathway Comprising PuuE-YneI in Which γ-Aminobutyrate Is Degraded into Succinate in Escherichia coli K-12▿

    Science.gov (United States)

    Kurihara, Shin; Kato, Kenji; Asada, Kei; Kumagai, Hidehiko; Suzuki, Hideyuki

    2010-01-01

    γ-Aminobutyrate (GABA) is metabolized to succinic semialdehyde by GABA aminotransferase (GABA-AT), and the succinic semialdehyde is subsequently oxidized to succinate by succinic semialdehyde dehydrogenase (SSADH). In Escherichia coli, there are duplicate GABA-ATs (GabT and PuuE) and duplicate SSADHs (GabD and YneI). While GabT and GabD have been well studied previously, the characterization and expression analysis of PuuE and YneI are yet to be investigated. By analyzing the amino acid profiles in cells of ΔpuuE and/or ΔgabT mutants, this study demonstrated that PuuE plays an important role in GABA metabolism in E. coli cells. The similarity of the amino acid sequences of PuuE and GabT is 67.4%, and it was biochemically demonstrated that the catalytic center of GabT is conserved as an amino acid residue important for the enzymatic activity in PuuE as Lys-247. However, the regulation of expression of PuuE is significantly different from that of GabT. PuuE is induced by the addition of putrescine to the medium and is repressed by succinate and low aeration conditions; in contrast, GabT is almost constitutive. Similarly, YneI is induced by putrescine, while GabD is not. For E. coli, PuuE is important for utilization of putrescine as a sole nitrogen source and both PuuE and YneI are important for utilization of putrescine as a sole carbon source. The results demonstrate that the PuuE-YneI pathway was a putrescine-inducible GABA degradation pathway for utilizing putrescine as a nutrient source. PMID:20639325

  10. Production of ultrafine sumatriptan succinate particles for pulmonary delivery.

    Science.gov (United States)

    Yang, Zong-Yang; Le, Yuan; Hu, Ting-Ting; Shen, Zhigang; Chen, Jian-Feng; Yun, Jimmy

    2008-09-01

    Drug particle physical properties are critical for the efficiency of a drug delivered to the lung. The purpose of this study was to produce ultrafine sumatriptan succinate particles for inhalation. Sumatriptan succinate particles were produced via reactive precipitation without any surfactants. Several low toxic organic solvents such as acetone, isopropanol, and tetrahydrofuran were investigated as the reaction medium. And the dry powder was obtained via spray drying. FT-IR, HPLC, SEM and XRD were exploited to characterize the physicochemical properties of the ultrafine sumatriptan succinate dry powder. The aerosol performance of the powder was evaluated using an Aeroliser connected to a multi stage liquid impinger operating at 60 l/min. The mean particle size of the ultrafine sumatriptan succinate particles obtained under optimum conditions was in the range of 630-679 nm and consequently they were in the respirable range. The spray-dried powder whose fine particle fraction was increased up to 50.6 +/- 8.2% showed good aerosol performance whereas the vacuum-dried powder was approximate 18.2 +/- 3.0%. Good aerosol performance ultrafine sumatriptan succinate particles could be produced by reactive precipitation without any additives followed by spray drying at the optimum parameters.

  11. Metabolic capabilities of Actinobacillus succinogenes for succinic acid production

    Directory of Open Access Journals (Sweden)

    R. Rafieenia

    2014-12-01

    Full Text Available Attention has been focused on microbial succinic acid production as an alternative for conventional chemical synthesis that is associated with environmental pollution. A metabolic model for Actinobacillus succinogenes 130Z was developed with a mixture of glucose and xylose as substrate. The metabolic fluxes during succinicate production were determined using flux balance analysis by linear programming optimization in the MATLAB environment. Different glucose ratios (0.3, 0.4 and 0.7 mol.mol-1substrate were used as model assumptions to calculate optimal fluxes, maximum growth and succinate production. The model revealed that higher growth rates and product yields were correlated with higher glucose content in the substrate mixture. When glucose constituted 0.5 mol.mol-1 substrate, a lower succinate yield (0.64 mol.mol-1 substrate was obtained, compared to 0.73 mol.mol-1 substrate when glucose was used individually. Deletion of different unessential reactions in the model showed that a knockout of the acetate formation pathway would increase the succinate yield by 21% when glucose and xylose were used in equal molar ratios.

  12. Succinic Acid Synthesis by Ethanol-Grown Yeasts

    Directory of Open Access Journals (Sweden)

    Svetlana V. Kamzolova

    2009-01-01

    Full Text Available The synthesis of succinic acid in ethanol-containing media has been tested in 32 yeasts of different genera (Debaryomyces, Candida, Pichia, Saccharomyces, Torulopsis. The capability of succinic acid synthesis was revealed in 29 strains, from which two most effective producers were selected. When grown in a fermentor under high aeration in mineral medium with pulsed addition of ethanol, the strain Candida catenulata VKM Y-5 produced succinic acid up to 5.2 g/L with mass yield of 32.6 % and energy yield of 14.8 %; the other strain, Candida zeylanoides VKM Y-2324, excreted 9.4 g/L of succinic acid with mass and energy yields of 39 and 17.8 %, respectively. It was indicated that succinic acid formation in the yeasts was accompanied by the synthesis of considerable amounts of malic acid, which was apparently due to a high activity of the glyoxylate cycle. Growth characteristics of both strains were studied in dependence on the concentrations of ethanol, zinc ions and nitrogen in the medium.

  13. 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...

  14. Genome-Wide Identification and Expression Profiling of Cytokinin Oxidase/Dehydrogenase (CKX) Genes Reveal Likely Roles in Pod Development and Stress Responses in Oilseed Rape (Brassica napus L.).

    Science.gov (United States)

    Liu, Pu; Zhang, Chao; Ma, Jin-Qi; Zhang, Li-Yuan; Yang, Bo; Tang, Xin-Yu; Huang, Ling; Zhou, Xin-Tong; Lu, Kun; Li, Jia-Na

    2018-03-16

    Cytokinin oxidase/dehydrogenases (CKXs) play a critical role in the irreversible degradation of cytokinins, thereby regulating plant growth and development. Brassica napus is one of the most widely cultivated oilseed crops worldwide. With the completion of whole-genome sequencing of B. napus , genome-wide identification and expression analysis of the BnCKX gene family has become technically feasible. In this study, we identified 23 BnCKX genes and analyzed their phylogenetic relationships, gene structures, conserved motifs, protein subcellular localizations, and other properties. We also analyzed the expression of the 23 BnCKX genes in the B. napus cultivar Zhong Shuang 11 ('ZS11') by quantitative reverse-transcription polymerase chain reaction (qRT-PCR), revealing their diverse expression patterns. We selected four BnCKX genes based on the results of RNA-sequencing and qRT-PCR and compared their expression in cultivated varieties with extremely long versus short siliques. The expression levels of BnCKX5-1 , 5-2 , 6-1 , and 7-1 significantly differed between the two lines and changed during pod development, suggesting they might play roles in determining silique length and in pod development. Finally, we investigated the effects of treatment with the synthetic cytokinin 6-benzylaminopurine (6-BA) and the auxin indole-3-acetic acid (IAA) on the expression of the four selected BnCKX genes. Our results suggest that regulating BnCKX expression is a promising way to enhance the harvest index and stress resistance in plants.

  15. Stably expressed housekeeping genes across developmental stages in the two-spotted spider mite, Tetranychus urticae.

    Science.gov (United States)

    Yang, Chunxiao; Pan, Huipeng; Liu, Yong; Zhou, Xuguo

    2015-01-01

    Quantitative real-time PCR (qRT-PCR) is a reliable and reproducible technique for measuring mRNA expression. To facilitate gene expression studies and obtain more accurate qRT-PCR analysis, normalization relative to stable housekeeping genes is mandatory. In this study, ten housekeeping genes, including beta-actin (Actin) , elongation factor 1 α (EF1A) , glyceralde hyde-3-phosphate dehydrogenase (GAPDH) , ribosomal protein L13 (RPL13) , ribosomal protein 49 (RP49) , α-tubulin (Tubulin) , vacuolar-type H+-ATPase (v-ATPase) , succinate dehydrogenase subunit A (SDHA) , 28S ribosomal RNA (28S) , and 18S ribosomal RNA (18S) from the two-spotted spider mite, Tetranychus urticae, were selected as the candidate reference genes. Four algorithms, geNorm, Normfinder, BestKeeper, and the ΔCt method, were used to evaluate the performance of these candidates as endogenous controls across different developmental stages. In addition, RefFinder, which integrates the above-mentioned software tools, provided the overall ranking of the stability/suitability of these candidate reference genes. Among them, PRL13 and v-ATPase were the two most stable housekeeping genes across different developmental stages. This work is the first step toward establishing a standardized qRT-PCR analysis in T. urticae following the MIQE guideline. With the recent release of the T. urticae genome, results from this study provide a critical piece for the subsequent genomics and functional genomics research in this emerging model system.

  16. Warburg tumours and the mechanisms of mitochondrial tumour suppressor genes. Barking up the right tree?

    Science.gov (United States)

    Bayley, Jean-Pierre; Devilee, Peter

    2010-06-01

    The past decade has seen a revival of interest in the metabolic adaptations of tumours, named for their original discoverer, Otto Warburg. Warburg reported a high rate of glycolysis in tumours, and a concurrent defect in mitochondrial respiration. The rediscovery of Warburg's hypothesis coincided with the discovery of mitochondrial tumours suppressor genes that may conform to Warburg's hypothesis. Succinate dehydrogenase and fumarate hydratase are mitochondrial proteins of the TCA cycle and the respiratory chain and when mutated lead to tumours of the nervous system known as paragangliomas and pheochromocytomas, and in the case of fumarate hydratase, cutaneous and uterine leiomyomas and renal cell cancer. Recently a novel mitochondrial protein, SDHAF2 (SDH5), was also shown to be a paraganglioma-related tumour suppressor gene. Another mitochondrial and TCA cycle-related protein, isocitrate dehydrogenase 2 is, together with IDH1, frequently mutated in the brain tumour glioblastoma. There are currently many competing hypotheses on the role of these genes in tumourigenesis, but frequent themes are the stabilization of hypoxia inducible factor 1 and upregulation of genes involved in angiogenesis, glucose transport and glycolysis. Other postulated mechanisms include the inhibition of developmental apoptosis, altered gene expression due to histone deregulation and the acquisition of novel catalytic properties. Here we discuss these diverse hypotheses and highlight very recent findings on the possible effects of IDH gene mutations.

  17. [Agrobacterium-mediated sunflower transformation (Helianthus annuus L.) in vitro and in Planta using strain of LBA4404 harboring binary vector pBi2E with dsRNA-suppressor proline dehydrogenase gene].

    Science.gov (United States)

    Tishchenko, E N; Komisarenko, A G; Mikhal'skaia, S I; Sergeeva, L E; Adamenko, N I; Morgun, B V; Kochetov, A V

    2014-01-01

    To estimate the efficiency of proline dehydrogenase gene suppression towards increasing of sunflower (Helianthus annuus L.) tolerance level to water deficit and salinity, we employed strain LBA4404 harboring pBi2E with double-stranded RNA-suppressor, which were prepared on basis arabidopsis ProDH1 gene. The techniques of Agrobacterium-mediated transformation in vitro and in planta during fertilization sunflower have been proposed. There was shown the genotype-depended integration of T-DNA in sunflower genome. PCR-analysis showed that ProDH1 presents in genome of inbred lines transformed in planta, as well as in T1- and T2-generations. In trans-genic regenerants the essential accumulation of free L-proline during early stages of in vitro cultivation under normal conditions was shown. There was established the essential accumulation of free proline in transgenic regenerants during cultivation under lethal stress pressure (0.4 M mannitol and 2.0% sea water salts) and its decline upon the recovery period. These data are declared about effectiveness of suppression of sunflower ProDH and gene participation in processes connected with osmotolerance.

  18. MODIFIKASI ASAM SUKSINAT - GELOMBANG PENDEK UNTUK\tPRODUKSI TAPIOKA SUKSINAT Succinic Acid-Microwave Modification to Produce Succinic Tapioca

    Directory of Open Access Journals (Sweden)

    Heny Herawati

    2012-05-01

    Full Text Available Indonesia as tropical country has great cassava potency. The great chance of cassava product development could be increased its added value through modified tapioca processing. One of modified starch that could be implemented as a food additive is succinic starch. The tapioca succinilation processed through combination process of reacting tapioca with succinic acid and microwave treatment. The research method was conducted by factorial design with 3 factors: substrate concentration (30 %, 40 %, succinic acid concentration (1 %, 3 %, 5 %, and drying method (oven and microwave. Succinic tapioca was analyzed both physical and chemical characteristics, while optimal product was fur- ther analyzed for nutrition contents and surface microstructure using SEM. Succinic acid and microwave modification influenced to the physical and chemical succinic tapioca, except ash content. The highest substitution degree value was 0.929 which was obtained by combination of substrate concentration 40 %, succinic acid added 5 % and microwave processed. The change of granule size was not significant, just the distribution among granule correlated with the tapi- oca modification. The succinic tapioca granule size ranged 5.35 µm until 17.20 µm with average 11.15 µm. Succinic tapioca characteristic hopefully could be advanced food implementation. ABSTRAK Indonesia merupakan negara tropis yang memiliki potensi produksi ubi kayu yang cukup besar. Peluang pengem- bangan produk berbasis ubi kayu di Indonesia masih cukup besar diantaranya yaitu peningkatan nilai tambah ubi kayu melalui proses modifikasi tapioka. Salah satu potensi pati termodifikasi yang dapat dipergunakan untuk bahan tambahan makanan yaitu pati suksinat. Pada penelitian ini proses suksinilasi tapioka dilakukan dengan cara mereak- sikan asam suksinat yang dikombinasikan dengan mempergunakan microwave. Metodologi penelitian yang dilakukan menggunakan rancangan faktorial dengan 3 faktor, yaitu konsentrasi

  19. A new compound heterozygous frameshift mutation in the type II 3{beta}-hydroxysteroid dehydrogenase 3{beta}-HSD gene causes salt-wasting 3{beta}-HSD deficiency congenital adrenal hyperplasia

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, L.; Sakkal-Alkaddour, S.; Chang, Ying T.; Yang, Xiaojiang; Songya Pang [Univ. of Illinois, Chicago, IL (United States)

    1996-01-01

    We report a new compound heterozygous frameshift mutation in the type II 3{Beta}-hydroxysteroid dehydrogenase (3{beta}-HSD) gene in a Pakistanian female child with the salt-wasting form of 3{Beta}-HSD deficiency congenital adrenal hyperplasia. The etiology for her congenital adrenal hyperplasia was not defined. Although the family history suggested possible 3{beta}-HSd deficiency disorder, suppressed adrenal function caused by excess glucocorticoid therapy in this child at 7 yr of age did not allow hormonal diagnosis. To confirm 3{beta}-HSD deficiency, we sequenced the type II 3{beta}-HSD gene in the patient, her family, and the parents of her deceased paternal cousins. The type II 3{beta}-HSD gene region of a putative promotor, exons I, II, III, and IV, and exon-intron boundaries were amplified by PCR and sequenced in all subjects. The DNA sequence of the child revealed a single nucleotide deletion at codon 318 [ACA(Thr){r_arrow}AA] in exon IV in one allele, and two nucleotide deletions at codon 273 [AAA(Lys){r_arrow}A] in exon IV in the other allele. The remaining gene sequences were normal. The codon 318 mutation was found in one allele from the father, brother, and parents of the deceased paternal cousins. The codon 273 mutation was found in one allele of the mother and a sister. These findings confirmed inherited 3{beta}-HSD deficiency in the child caused by the compound heterozygous type II 3{beta}-HSD gene mutation. Both codons at codons 279 and 367, respectively, are predicted to result in an altered and truncated type II 3{beta}-HSD protein, thereby causing salt-wasting 3{beta}-HSD deficiency in the patient. 21 refs., 2 figs., 1 tab.

  20. SMALL SCALE PREPARATION OF C14 LABELED SUCCINIC, MALIC, FUMARICAND TARTARIC ACIDS

    Energy Technology Data Exchange (ETDEWEB)

    Jorgensen, E.C.; Bassham, J.A.; Calvin, M.; Tolbert, B.M.

    1951-06-12

    The directions for preparing the following acids from labeled acetic acid on a 1 to 20 millimole scale is presented: Succinic acid, malic acid, fumaric acid and tartaric acid. Two methods for preparing the succinic acid are detailed.

  1. Medium- and short-chain dehydrogenase/reductase gene and protein families : the SDR superfamily: functional and structural diversity within a family of metabolic and regulatory enzymes.

    Science.gov (United States)

    Kavanagh, K L; Jörnvall, H; Persson, B; Oppermann, U

    2008-12-01

    Short-chain dehydrogenases/reductases (SDRs) constitute a large family of NAD(P)(H)-dependent oxidoreductases, sharing sequence motifs and displaying similar mechanisms. SDR enzymes have critical roles in lipid, amino acid, carbohydrate, cofactor, hormone and xenobiotic metabolism as well as in redox sensor mechanisms. Sequence identities are low, and the most conserved feature is an alpha/beta folding pattern with a central beta sheet flanked by 2 - 3 alpha-helices from each side, thus a classical Rossmannfold motif for nucleotide binding. The conservation of this element and an active site, often with an Asn-Ser-Tyr-Lys tetrad, provides a platform for enzymatic activities encompassing several EC classes, including oxidoreductases, epimerases and lyases. The common mechanism is an underlying hydride and proton transfer involving the nicotinamide and typically an active site tyrosine residue, whereas substrate specificity is determined by a variable C-terminal segment. Relationships exist with bacterial haloalcohol dehalogenases, which lack cofactor binding but have the active site architecture, emphasizing the versatility of the basic fold in also generating hydride transfer-independent lyases. The conserved fold and nucleotide binding emphasize the role of SDRs as scaffolds for an NAD(P)(H) redox sensor system, of importance to control metabolic routes, transcription and signalling.

  2. Efficient poly(3-hydroxypropionate) production from glycerol using Lactobacillus reuteri and recombinant Escherichia coli harboring L. reuteri propionaldehyde dehydrogenase and Chromobacterium sp. PHA synthase genes.

    Science.gov (United States)

    Linares-Pastén, Javier A; Sabet-Azad, Ramin; Pessina, Laura; Sardari, Roya R R; Ibrahim, Mohammad H A; Hatti-Kaul, Rajni

    2015-03-01

    Poly(3-hydroxypropionate), P(3HP), is a polymer combining good biodegradability with favorable material properties. In the present study, a production system for P(3HP) was designed, comprising conversion of glycerol to 3-hydroxypropionaldehyde (3HPA) as equilibrium mixture with 3HPA-hydrate and -dimer in aqueous system (reuterin) using resting cells of native Lactobacillus reuteri in a first stage followed by transformation of the 3HPA to P(3HP) using recombinant Escherichia coli strain co-expressing highly active coenzyme A-acylating propionaldehyde dehydrogenase (PduP) from L. reuteri and polyhydroxyalkanoate synthase (PhaCcs) from Chromobacterium sp. P(3HP) content of up to 40% (w/w) cell dry weight was reached, and the yield with respect to the reuterin consumed by the cells was 78%. Short biotransformation period (4.5h), lack of additives or expensive cofactors, and use of a cheap medium for cultivation of the recombinant strain, provides a new efficient and potentially economical system for P(3HP) production. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Gene structure and mutations of glutaryl-coenzyme A dehydrogenase: Impaired association of enzyme subunits that is due to an A421V substitution causes glutaric acidemia type I in the Amish

    Energy Technology Data Exchange (ETDEWEB)

    Biery, B.J.; Stein, D.E.; Goodman, S.I. [Univ. of Colorado School of Medicine, Denver, CO (United States)] [and others

    1996-11-01

    The structure of the human glutaryl coenzyme A dehydrogenase (GCD) gene was determined to contain 11 exons and to span {approximately}7 kb. Fibroblast DNA from 64 unrelated glutaric academia type I (GA1) patients was screened for mutations by PCR amplification and analysis of SSCP. Fragments with altered electrophoretic mobility were subcloned and sequenced to detect mutations that caused GA1. This report describes the structure of the GCD gene, as well as point mutations and polymorphisms found in 7 of its 11 exons. Several mutations were found in more than one patient, but no one prevalent mutation was detected in the general population. As expected from pedigree analysis, a single mutant allele causes GA1 in the Old Order Amish of Lancaster County, Pennsylvania. Several mutations have been expressed in Escherichia coli, and all produce diminished enzyme activity. Reduced activity in GCD encoded by the A421V mutation in the Amish may be due to impaired association of enzyme subunits. 13 refs., 5 figs., 3 tabs.

  4. 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.

  5. [A novel homozygous mutation p.E25X in the HSD3B2 gene causing salt wasting 3β-hydroxysteroid dehydrogenases deficiency in a Chinese pubertal girl: a delayed diagnosis until recurrent ovary cysts].

    Science.gov (United States)

    Huang, Yonglan; Zheng, Jipeng; Xie, Ting; Xiao, Qing; Lu, Shaomei; Li, Xiuzhen; Cheng, Jing; Chen, Lihe; Liu, Li

    2014-12-01

    3β- hydroxysteroid dehydrogenase deficiency (3βHSD), a rare form of congenital adrenal hyperplasia (CAH) resulted from mutations in the HSD3B2 gene that impair steroidogenesis in both adrenals and gonads. We report clinical features and the results of HSD3B2 gene analysis of a Chinese pubertal girl with salt wasting 3βHSD deficiency. We retrospectively reviewed clinical presentations and steroid profiles of the patient diagnosed in Guangzhou Women and Children's Medical Center in 2013. PCR and direct sequencing were used to identify any mutation in the HSD3B2 gene. A 13-year-old girl was diagnosed as CAH after birth because of salt-wasting with mild clitorimegaly and then was treated with glucocorticoid replacement. Breast and pubic hair development were normal, and menarche occurred at 12 yr, followed by menstrual bleeding about every 45 days. In the last one year laparoscopic operation and ovariocentesis were performed one after another for recurrent ovary cysts. Under corticoid acetate therapy, ACTH 17.10 pmol/L (normal 0-10.12), testosterone 1.31 nmol/L (normal T (p.E25X) was identified in HSD3B2 gene. The girl was homozygous and her mother was heterozygous, while her father was not identified with this mutation. A classic 3βHSD deficiency is characterized by salt wasting and mild virilization in female. Ovary cysts may be the one of features of gonad phenotype indicating ovary 3βHSD deficiency. A novel homozygous mutation c.73G >T(p.E25X) was related to the classical phenotype.

  6. Involvement of 17β-hydroxysteroid dehydrogenase type gene 1 937 A>G polymorphism in infertility in Polish Caucasian women with endometriosis.

    Science.gov (United States)

    Osiński, Maciej; Mostowska, Adrianna; Wirstlein, Przemyslaw; Skrzypczak, Jana; Jagodziński, Paweł Piotr; Szczepańska, Malgorzata

    2017-06-01

    Endometriosis is considered to be an estrogen-related chronic inflammatory disease. The 17β-hydroxysteroid dehydrogenase 1 (HSD17B1) converts estrone to 17β estradiol. The role of HSD17B1 937 A>G (rs605059) single nucleotide polymorphism (SNP) in development of endometriosis is still disputable. This study evaluated the association of the HSD17B1 937 A>G (rs605059) SNP with infertile women affected by endometriosis from Polish Caucasian population. The genotyping of cases (n = 290) and fertile women (n = 410) was conducted by high-resolution melting curve analysis. Statistical analysis demonstrated that the HSD17B1 937 A>G SNP is associated with endometriosis in stages I and II. The p trend and p allelic values calculated for the HSD17B1 937 A>G polymorphism were statistically significant and were equal to 0.001 and 0.0009, respectively. There was a significant association for the dominant model: (AG + GG vs AA) OR = 1.973 (95% CI = 1.178-3.304), p = 0.009, and for the recessive model: (GG vs AG + AA) OR = 1.806 (95% CI = 1.178-2.770), p = 0.006. However, we did not find statistical association of HSD17B1 937 A>G polymorphism with all infertile women with endometriosis or infertile women with endometriosis in stages III and IV. Our genetic study demonstrated HSD17B1 937 G variant as a risk factor for infertility in women with stage I and II endometriosis in Polish Caucasian patients.

  7. Population study of 1311 C/T polymorphism of Glucose 6 Phosphate Dehydrogenase gene in Pakistan – an analysis of 715 X-chromosomes

    Directory of Open Access Journals (Sweden)

    Naqvi Zulfiqar

    2009-07-01

    Full Text Available Abstract Background Nucleotide 1311 polymorphism at exon 11 of G6PD gene is widely prevalent in various populations of the world. The aim of the study was to evaluate 1311 polymorphism in subjects carrying G6PD Mediterranean gene and in general population living in Pakistan. Results Patients already known to be G6PD deficient were tested for 563C-T (G6PD Mediterranean and 1311 C-T mutation through RFLP based PCR and gene sequencing. A control group not known to be G6PD deficient was tested for 1311C/T only. C-T transition at nt 1311 was detected in 60/234 X-chromosomes with 563 C-T mutation (gene frequency of 0.26 while in 130 of normal 402 X-chromosomes (gene frequency of 0.32. Conclusion We conclude that 1311 T is a frequent polymorphism both in general populations and in subjects with G6PD Mediterranean gene in Pakistan. The prevalence is higher compared to most of the populations of the world. The present study will help in understanding genetic basis of G6PD deficiency in Pakistani population and in developing ancestral links of its various ethnic groups.

  8. Effect of a novel insulinotropic agent, succinic acid monoethyl ester ...

    Indian Academy of Sciences (India)

    Madhu

    In the present study, the effect of succinic acid monoethyl ester (EMS) on the pattern of lipids and lipoproteins in streptozotocin-nicotinamide induced type 2 diabetes was investigated. Type 2 diabetes was induced in male Wistar rats by single intraperitoneal injection (i.p.) of 45 mg/kg streptozotocin, 15 min after the i.p ...

  9. Effect of a novel insulinotropic agent, succinic acid monoethyl ester ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Biosciences; Volume 31; Issue 5. Effect of a novel insulinotropic agent, succinic acid monoethyl ester, on lipids and lipoproteins levels in rats with streptozotocin-nicotinamideinduced type 2 diabetes. Ramalingam Saravanan Leelavinothan Pari. Articles Volume 31 Issue 5 December 2006 pp 581- ...

  10. 21 CFR 522.1884 - Prednisolone sodium succinate injection.

    Science.gov (United States)

    2010-04-01

    ...-1, 4-diene-3, 20-dione 21-succinate sodium salt. (b) Specifications. Each milliliter of prednisolone... horses, dogs, and cats as provided in paragraphs (d)(1), (2) (i), (ii), and (iii) of this section. (d) Conditions of use. (1) The drug is intended for the treatment of horses, dogs, and cats.1 1 These conditions...

  11. Structural and magnetic studies on copper succinate dihydrate

    Indian Academy of Sciences (India)

    The four bis-bidendate succinate anions form syn–syn bridges among two copper atomsto form a polymeric two-dimensional chain. From room temperature vibrating sample magnetometer (VSM) studies themagnetic moment of the material is calculated as 1.35 Bohr magneton (BM), indicating antiferromagnetic interaction ...

  12. Evaluation of reference genes for real-time PCR studies of Brazilian Somalis sheep infected by gastrointestinal nematodes

    Directory of Open Access Journals (Sweden)

    Lilian Giotto Zaros

    2010-01-01

    Full Text Available Precise normalization with reference genes is necessary, in order to obtain reliable relative expression data in response to gastrointestinal nematode infection. By using sheep from temperate regions as models, three reference genes, viz., ribosomal protein LO (RPLO, glyceraldehyde 3-phosphate dehydrogenase (GAPDH and succinate dehydrogenase complex subunit A (SDHA, were investigated in the abomasum, abomasal lymph nodes and small intestine of Brazilian Somalis sheep, either resistant or susceptible to gastrointestinal nematodes infections. Real time PCR was carried out by using SYBR Green I dye, and gene stability was tested by geNorm. RPLO was an ideal reference gene, since its expression was constant across treatments, presented lower variation, and was ranked as the most stable in abomasum and lymph node tissues. On the other hand, SDHA was the most stable in the small intestine followed by RPLO and GAPDH. These findings demonstrate the importance of correctly choosing reference genes prior to relative quantification. In addition, we determined that reference genes used in sheep from temperate regions, when properly tested, can be applied in animals from tropical regions such as the Brazilian Somalis sheep.

  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. Cloning and in silico analysis of a cinnamyl alcohol dehydrogenase ...

    Indian Academy of Sciences (India)

    2014-04-16

    Apr 16, 2014 ... 1992; Kim et al. 2004), the relationship between CAD genes and their functions was of great impor- tance. Since the important role in regulation of lignin con- tent and composition, more and more CAD genes and their. Keywords. lignin biosynthesis; cinnamyl alcohol dehydrogenase; clone; in silico analysis ...

  15. Method for construction of bacterial strains with increased succinic acid production

    Science.gov (United States)

    Donnelly, Mark I.; Sanville-Millard, Cynthia; Chatterjee, Ranjini

    2000-01-01

    A fermentation process for producing succinic acid is provided comprising selecting a bacterial strain that does not produce succinic acid in high yield, disrupting the normal regulation of sugar metabolism of said bacterial strain, and combining the mutant bacterial strain and selected sugar in anaerobic conditions to facilitate production of succinic acid. Also provided is a method for changing low yield succinic acid producing bacteria to high yield succinic acid producing bacteria comprising selecting a bacterial strain having a phosphotransferase system and altering the phosphotransferase system so as to allow the bacterial strain to simultaneously metabolize different sugars.

  16. ROS generation and multiple forms of mammalian mitochondrial glycerol-3-phosphate dehydrogenase.

    Science.gov (United States)

    Mráček, Tomáš; Holzerová, Eliška; Drahota, Zdeněk; Kovářová, Nikola; Vrbacký, Marek; Ješina, Pavel; Houštěk, Josef

    2014-01-01

    Overproduction of reactive oxygen species (ROS) has been implicated in a range of pathologies. Mitochondrial flavin dehydrogenases glycerol-3-phosphate dehydrogenase (mGPDH) and succinate dehydrogenase (SDH) represent important ROS source, but the mechanism of electron leak is still poorly understood. To investigate the ROS production by the isolated dehydrogenases, we used brown adipose tissue mitochondria solubilized by digitonin as a model. Enzyme activity measurements and hydrogen peroxide production studies by Amplex Red fluorescence, and luminol luminescence in combination with oxygraphy revealed flavin as the most likely source of electron leak in SDH under in vivo conditions, while we propose coenzyme Q as the site of ROS production in the case of mGPDH. Distinct mechanism of ROS production by the two dehydrogenases is also apparent from induction of ROS generation by ferricyanide which is unique for mGPDH. Furthermore, using native electrophoretic systems, we demonstrated that mGPDH associates into homooligomers as well as high molecular weight supercomplexes, which represent native forms of mGPDH in the membrane. By this approach, we also directly demonstrated that isolated mGPDH itself as well as its supramolecular assemblies are all capable of ROS production. © 2013.

  17. [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.

  18. [Informatics analysis of malate dehydrogenase from Taenia saginata asiatica].

    Science.gov (United States)

    Huang, Jiang; Hu, Xu-Chu; Huang, Yan; Yu, Xin-Bing; Bao, Huai-En; Lang, Shu-Yuan; Liao, Xing-Jiang

    2008-06-30

    Tools from bioinformatics websites such as NCBI, ExPaSy were used for the analysis. The malate dehydrogenase full-length gene from Taenia saginata asiatica was 1 212 bp in length, with a coding region of 30-1 028 bp and coding 332 amino acids. It was a complete and full-length gene compared with the homologues in GenBank. The protein showed no transmembrane region, with stable physical-chemical characteristics. Three major linear epitopes located aa95-aa100, aa322-aa327 and aa117-aa122, with certain distance from each other on the surface of spatial structure of malate dehydrogenase (MDH). The last one was the linear epitope of Taenia. This cytosolic malate dehydrogenase gene is a potential antigen for diagnosis.

  19. Molecular Differentiation of Fasciola Species and Characterization of Genetic Diversity of F. gigantica Using NADH Dehydrogenase I (ND1 Gene in the Endemic Areas of Iran.

    Directory of Open Access Journals (Sweden)

    Shabnam Tadayon

    2015-03-01

    Full Text Available Fasciola hepatica and F. gigantica are the causative agents of fasciolosis in domestic animals and humans. Based on the morphometric criteria, differential diagnosis between them is problematic. In addition, intermediate forms of Fasciola have been found in Iran, which makes the differentiation more difficult. The aim of the present study was to provide molecular evidence for the existence of F. gigantica in Iran using sequencing analysis of ND1 and PCR-RFLP analysis of ITS2 regions and to study the intraspecies variations of F. gigantica based on mitochondrial ND1 gene polymorphism.Forty Fasciola spp. samples collected from four distinct provinces (Fars, Khuzestan, Gilan, Khorasan Razavi in Iran were collected for morphological and molecular characterization. In molecular method, PCR-RFLP analysis of ITS2 using pagI restriction enzyme was used as a screening approach for F. gigantica differentiation. Then mitochondrial DNA sequence variations in the ND1 gene were used for phylogenetic analysis.Based on the morphometric criteria and RFLP analysis, 14 parasitic samples were initially identified to be F. gigantica. Phylogenetic results showed that there are at least 10 different genotypes of F. gigantica in Iran, which are different from those existing in the GenBank. Twenty-six points out of 410 base pairs of sequenced ND1 gene in 10 varieties of F. gigantica were diagnosed to be polymorphic. From 26 points of polymorphism, only eight resulted in the post-translational amino acid changes in ND1 gene product structure.Data revealed noticeable genetic diversity (up to 4.63% between different varieties of F. gigantica in Iran.

  20. 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 ...

  1. Identification, Cloning, and Characterization of l-Phenylserine Dehydrogenase from Pseudomonas syringae NK-15

    Directory of Open Access Journals (Sweden)

    Sakuko Ueshima

    2010-01-01

    Full Text Available The gene encoding d-phenylserine dehydrogenase from Pseudomonas syringae NK-15 was identified, and a 9,246-bp nucleotide sequence containing the gene was sequenced. Six ORFs were confirmed in the sequenced region, four of which were predicted to form an operon. A homology search of each ORF predicted that orf3 encoded l-phenylserine dehydrogenase. Hence, orf3 was cloned and overexpressed in Escherichia coli cells and recombinant ORF3 was purified to homogeneity and characterized. The purified ORF3 enzyme showed l-phenylserine dehydrogenase activity. The enzymological properties and primary structure of l-phenylserine dehydrogenase (ORF3 were quite different from those of d-phenylserine dehydrogenase previously reported. l-Phenylserine dehydrogenase catalyzed the NAD+-dependent oxidation of the β-hydroxyl group of l-β-phenylserine. l-Phenylserine and l-threo-(2-thienylserine were good substrates for l-phenylserine dehydrogenase. The genes encoding l-phenylserine dehydrogenase and d-phenylserine dehydrogenase, which is induced by phenylserine, are located in a single operon. The reaction products of both enzymatic reactions were 2-aminoacetophenone and CO2.

  2. GlnR negatively regulates the transcription of the alanine dehydrogenase encoding gene ald in Amycolatopsis mediterranei U32 under nitrogen limited conditions via specific binding to its major transcription initiation site.

    Directory of Open Access Journals (Sweden)

    Ying Wang

    Full Text Available Ammonium assimilation is catalyzed by two enzymatic pathways, i.e., glutamine synthetase/glutamate synthase (GS/GOGAT and alanine dehydrogenase (AlaDH in Amycolatopsis mediterranei U32. Under nitrogen-rich conditions, the AlaDH pathway is the major route for ammonium assimilation, while the GS/GOGAT pathway takes over when the extracellular nitrogen supply is limited. The global nitrogen regulator GlnR was previously characterized to activate the transcription of the GS encoding gene glnA in response to nitrogen limitation and is demonstrated in this study as a repressor for the transcription of the AlaDH encoding gene ald, whose regulation is consistent with the switch of the ammonium assimilation pathways from AlaDH to GS/GOGAT responding to nitrogen limitation. Three transcription initiation sites (TISs of ald were determined with primer extension assay, among which transcription from aldP2 contributed the major transcripts under nitrogen-rich conditions but was repressed to an undetectable level in response to nitrogen limitation. Through DNase I footprinting assay, two separate regions were found to be protected by GlnR within ald promoter, within which three GlnR binding sites (a1, b1 sites in region I and a2 site in region II were defined. Interestingly, the major TIS aldP2 is located in the middle of a2 site within region II. Therefore, one may easily conclude that GlnR represses the transcription of ald via specific binding to the GlnR binding sites, which obviously blocks the transcription initiation from aldP2 and therefore reduces ald transcripts.

  3. Aldehyde dehydrogenase protein superfamily in maize.

    Science.gov (United States)

    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.

  4. Biophysical properties of phenyl succinic acid derivatised hyaluronic acid

    DEFF Research Database (Denmark)

    Neves-Petersen, Maria Teresa; Klitgaard, Søren; Skovsen, Esben

    2010-01-01

    Modification of hyaluronic acid (HA) with aryl succinic anhydrides results in new biomedical properties of HA as compared to non-modified HA, such as more efficient skin penetration, stronger binding to the skin, and the ability to blend with hydrophobic materials. In the present study, hyaluronic...... acid has been derivatised with the anhydride form of phenyl succinic acid (PheSA). The fluorescence of PheSA was efficiently quenched by the HA matrix. HA also acted as a singlet oxygen scavenger. Fluorescence lifetime(s) of PheSA in solution and when attached to the HA matrix has been monitored...... with ps resolved streak camera technology. Structural and fluorescence properties changes induced on HA-PheSA due to the presence of singlet oxygen were monitored using static light scattering (SLS), steady state fluorescence and ps time resolved fluorescence studies. SLS studies provided insight...

  5. Structural and magnetic studies on copper succinate dihydrate ...

    Indian Academy of Sciences (India)

    M P BINITHA

    2017-08-21

    Aug 21, 2017 ... DOI 10.1007/s12034-017-1459-0. Structural and magnetic studies on copper succinate dihydrate single crystals. M P BINITHA1,∗ and P P PRADYUMNAN2 ..... Chem. doi:10.1021/ic402194c. [5] Pilkington M, Gross M, Franz P, Biner M, Decurtins S,. Stoeckli-Evans H et al 2001 J. Solid State Chem. 159 262.

  6. Integration of succinic acid and ethanol production with potential application in a corn or barley biorefinery.

    Science.gov (United States)

    Nghiem, Nhuan P; Hicks, Kevin B; Johnston, David B

    2010-11-01

    Production of succinic acid from glucose by Escherichia coli strain AFP184 was studied in a batch fermentor. The bases used for pH control included NaOH, KOH, NH(4)OH, and Na(2)CO(3). The yield of succinic acid without and with carbon dioxide supplied by an adjacent ethanol fermentor using either corn or barley as feedstock was examined. The carbon dioxide gas from the ethanol fermentor was sparged directly into the liquid media in the succinic acid fermentor without any pretreatment. Without the CO(2) supplement, the highest succinic acid yield was observed with Na(2)CO(3), followed by NH(4)OH, and lowest with the other two bases. When the CO(2) produced in the ethanol fermentation was sparged into the media in the succinic acid fermentor, no improvement of succinic acid yield was observed with Na(2)CO(3). However, several-fold increases in succinic acid yield were observed with the other bases, with NH(4)OH giving the highest yield increase. The yield of succinic acid with CO(2) supplement from the ethanol fermentor when NH(4)OH was used for pH control was equal to that obtained when Na(2)CO(3) was used, with or without CO(2) supplementation. The benefit of sparging CO(2) from ethanol fermentation on the yield of succinic acid demonstrated the feasibility of integration of succinic acid fermentation with ethanol fermentation in a biorefinery for production of fuels and industrial chemicals.

  7. Synthesis of allitol from D-psicose using ribitol dehydrogenase and ...

    African Journals Online (AJOL)

    Purpose: To synthesize allitol from D-psicose by a combination of novel ribitol dehydrogenase (RDH) and formate dehydrogenase (FDH) under optimised production conditions. Methods: RDH and FDH genes were cloned and introduced into pET-22b(+) vectors for expression in Escherichia coli to produce the ...

  8. Measurement of DPD and TS transcripts aimed to predict clinical benefit from fluoropyrimidines: confirmation of the trend in Russian colorectal cancer series and caution regarding the gene referees.

    Science.gov (United States)

    Iyevleva, Aglaya G; Buslov, Konstantin G; Togo, Alexandr V; Matsko, Dmitry E; Filimonenko, Vasily P; Moiseyenko, Vladimir M; Imyanitov, Evgeny N

    2007-06-01

    Measurement of intratumoral expression of dihydropyrimidine dehydrogenase (DPD) and thymidylate synthase (TS) may have some value in predicting the response to fluoropyrimidine-containing therapy. We attempted to validate this association in a series of Russian metastatic colorectal cancer cases. While replicating already published protocols, we unexpectedly found that the use of commonly utilized gene referees, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-actin, may lead to artifacts due to pseudogene-driven amplification from the genomic DNA template. We have developed a real-time PCR protocol which amplifies short PCR fragments, thus allowing efficient analysis of archival formalin-fixed paraffin-embedded tumor samples, and relies on succinate dehydrogenase (SDHA) as a gene referee, therefore avoiding amplification from genomic DNA. Low content of DPD transcripts was observed in 13/20 (65%) patients with disease control (tumor response or disease stabilization) as compared to only 3/9 (33%) subjects with progressive disease (p = 0.11). Despite the low number of patients, this association reached the level of statistical significance when similar analysis was done for TS expression (11/20 (55%) vs. 1/9 (11%); p = 0.03). Our data confirm that low DPD and TS expressors have higher chances of success of fluoropyrimidine-containing regimens.

  9. Unsuspected task for an old team: succinate, fumarate and other Krebs cycle acids in metabolic remodeling.

    Science.gov (United States)

    Bénit, Paule; Letouzé, Eric; Rak, Malgorzata; Aubry, Laetitia; Burnichon, Nelly; Favier, Judith; Gimenez-Roqueplo, Anne-Paule; Rustin, Pierre

    2014-08-01

    Seventy years from the formalization of the Krebs cycle as the central metabolic turntable sustaining the cell respiratory process, key functions of several of its intermediates, especially succinate and fumarate, have been recently uncovered. The presumably immutable organization of the cycle has been challenged by a number of observations, and the variable subcellular location of a number of its constitutive protein components is now well recognized, although yet unexplained. Nonetheless, the most striking observations have been made in the recent period while investigating human diseases, especially a set of specific cancers, revealing the crucial role of Krebs cycle intermediates as factors affecting genes methylation and thus cell remodeling. We review here the recent advances and persisting incognita about the role of Krebs cycle acids in diverse aspects of cellular life and human pathology. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Adipocyte protein modification by Krebs cycle intermediates and fumarate ester-derived succination.

    Science.gov (United States)

    Manuel, Allison M; Frizzell, Norma

    2013-11-01

    Protein succination, the non-enzymatic modification of cysteine residues by fumarate, is distinguishable from succinylation, an enzymatic reaction forming an amide bond between lysine residues and succinyl-CoA. Treatment of adipocytes with 30 mM glucose significantly increases protein succination with only a small change in succinylation. Protein succination may be significantly increased intracellularly after treatment with fumaric acid esters, however, the ester must be removed by saponification to permit 2SC-antibody detection of the fumarate adduct.

  11. Progress of succinic acid production from renewable resources: Metabolic and fermentative strategies.

    Science.gov (United States)

    Jiang, Min; Ma, Jiangfeng; Wu, Mingke; Liu, Rongming; Liang, Liya; Xin, Fengxue; Zhang, Wenming; Jia, Honghua; Dong, Weiliang

    2017-12-01

    Succinic acid is a four-carbon dicarboxylic acid, which has attracted much interest due to its abroad usage as a precursor of many industrially important chemicals in the food, chemicals, and pharmaceutical industries. Facing the shortage of crude oil supply and demand of sustainable development, biological production of succinic acid from renewable resources has become a topic of worldwide interest. In recent decades, robust producing strain selection, metabolic engineering of model strains, and process optimization for succinic acid production have been developed. This review provides an overview of succinic acid producers and cultivation technology, highlight some of the successful metabolic engineering approaches. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Identification of glucose 6 phosphate dehydrogenase mutations by ...

    African Journals Online (AJOL)

    Identification of glucose 6 phosphate dehydrogenase mutations by single strand conformation polymorphism and gene sequencing analysis. ... Subject: Six DNA samples from Turkish males confirmed to have G-6-PD deficiency where available for the study. Results: One subject was found to have an abnormal mobility shift ...

  13. [Cytologic response of the irradiated vagina to estriol succinate].

    Science.gov (United States)

    Cabrera, J; Canahuate, E; del Campo, S L; Poloni, O; Ramírez, S I

    1993-01-01

    The vaginal response to estriol succinate was studied in 40 oncogynecological patients, 20 with irradiation (study group) and 20 without irradiation (control group). Both groups had artificial established menopause at the beginning of the treatment. The previous clinical and cytological patterns to estrogen therapy were more severe in the irradiated group; they presented the highest degree of vaginal atrophy which was related to the type and therapeutic dose used. Response of the study group to a continuous 4 mg of estriol as well as the control group, both has a positive effect in vaginal cytology and alleviates postmenopausal symptoms and improve the quality of life.

  14. A Toolbox for Quantitative Gene Expression in Varroa destructor: RNA Degradation in Field Samples and Systematic Analysis of Reference Gene Stability.

    Directory of Open Access Journals (Sweden)

    Ewan M Campbell

    Full Text Available Varroa destructor is the major pest of Apis mellifera and contributes to the global honey bee health crisis threatening food security. Developing new control strategies to combat Varroa will require the application of molecular biology, including gene expression studies by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR. Both high quality RNA samples and suitable stable internal reference genes are required for accurate gene expression studies. In this study, ten candidate genes (succinate dehydrogenase (SDHA, NADH dehydrogenase (NADH, large ribsosmal subunit, TATA-binding protein, glyceraldehyde-3-phosphate dehydrogenase, 18S rRNA (18S, heat-shock protein 90 (HSP90, cyclophilin, α-tubulin, actin, were evaluated for their suitability as normalization genes using the geNorm, Normfinder, BestKeeper, and comparative ΔCq algorithims. Our study proposes the use of no more than two of the four most stable reference genes (NADH, 18S, SDHA and HSP90 in Varroa gene expression studies. These four genes remain stable in phoretic and reproductive stage Varroa and are unaffected by Deformed wing virus load. When used for determining changes in vitellogenin gene expression, the signal-to-noise ratio (SNR for the relatively unstable genes actin and α-tubulin was much lower than for the stable gene combinations (NADH + HSP90 +18S; NADH + HSP90; or NADH. Using both electropherograms and RT-qPCR for short and long amplicons as quality controls, we demonstrate that high quality RNA can be recovered from Varroa up to 10 days later stored at ambient temperature if collected into RNAlater and provided the body is pierced. This protocol allows the exchange of Varroa samples between international collaborators and field sample collectors without requiring frozen collection or shipping. Our results make important contributions to gene expression studies in Varroa by proposing a validated sampling protocol to obtain high quality Varroa

  15. A Toolbox for Quantitative Gene Expression in Varroa destructor: RNA Degradation in Field Samples and Systematic Analysis of Reference Gene Stability.

    Science.gov (United States)

    Campbell, Ewan M; McIntosh, Catriona H; Bowman, Alan S

    2016-01-01

    Varroa destructor is the major pest of Apis mellifera and contributes to the global honey bee health crisis threatening food security. Developing new control strategies to combat Varroa will require the application of molecular biology, including gene expression studies by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Both high quality RNA samples and suitable stable internal reference genes are required for accurate gene expression studies. In this study, ten candidate genes (succinate dehydrogenase (SDHA), NADH dehydrogenase (NADH), large ribsosmal subunit, TATA-binding protein, glyceraldehyde-3-phosphate dehydrogenase, 18S rRNA (18S), heat-shock protein 90 (HSP90), cyclophilin, α-tubulin, actin), were evaluated for their suitability as normalization genes using the geNorm, Normfinder, BestKeeper, and comparative ΔCq algorithims. Our study proposes the use of no more than two of the four most stable reference genes (NADH, 18S, SDHA and HSP90) in Varroa gene expression studies. These four genes remain stable in phoretic and reproductive stage Varroa and are unaffected by Deformed wing virus load. When used for determining changes in vitellogenin gene expression, the signal-to-noise ratio (SNR) for the relatively unstable genes actin and α-tubulin was much lower than for the stable gene combinations (NADH + HSP90 +18S; NADH + HSP90; or NADH). Using both electropherograms and RT-qPCR for short and long amplicons as quality controls, we demonstrate that high quality RNA can be recovered from Varroa up to 10 days later stored at ambient temperature if collected into RNAlater and provided the body is pierced. This protocol allows the exchange of Varroa samples between international collaborators and field sample collectors without requiring frozen collection or shipping. Our results make important contributions to gene expression studies in Varroa by proposing a validated sampling protocol to obtain high quality Varroa RNA and the

  16. Validation of putative reference genes for gene expression studies in human hepatocellular carcinoma using real-time quantitative RT-PCR

    International Nuclear Information System (INIS)

    Cicinnati, Vito R; Shen, Qingli; Sotiropoulos, Georgios C; Radtke, Arnold; Gerken, Guido; Beckebaum, Susanne

    2008-01-01

    Reference genes, which are often referred to as housekeeping genes are frequently used to normalize mRNA levels between different samples in quantitative reverse transcription polymerase chain reaction (qRT-PCR). The selection of reference genes is critical for gene expression studies because the expression of these genes may vary among tissues or cells and may change under certain circumstances. Here, a systematic evaluation of six putative reference genes for gene expression studies in human hepatocellular carcinoma (HCC) is presented. Six genes, beta-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hydroxymethyl-bilane synthase (HMBS), hypoxanthine phosphoribosyl-transferase 1 (HPRT1), succinate dehydrogenase complex, subunit A (SDHA) and ubiquitin C (UBC), with distinct functional characteristics and expression patterns were evaluated by qRT-PCR. Inhibitory substances in RNA samples were quantitatively assessed and controlled using an external RNA control. The stability of selected reference genes was analyzed using both geNorm and NormFinder software. HMBS and GAPDH were identified as the optimal reference genes for normalizing gene expression data between paired tumoral and adjacent non-tumoral tissues derived from patients with HCC. HMBS, GAPDH and UBC were identified to be suitable for the normalization of gene expression data among tumor tissues; whereas the combination of HMBS, B2M, SDHA and GAPDH was suitable for normalizing gene expression data among five liver cancer cell lines, namely Hep3B, HepG2, HuH7, SK-HEP-1 and SNU-182. The determined gene stability was increased after exclusion of RNA samples containing relatively higher inhibitory substances. Of six genes studied, HMBS was found to be the single best reference gene for gene expression studies in HCC. The appropriate choice of combination of more than one reference gene to improve qRT-PCR accuracy depends on the kind of liver tissues or cells under investigation

  17. The life-extending gene Indy encodes an exchanger for Krebs-cycle intermediates.

    Science.gov (United States)

    Knauf, Felix; Mohebbi, Nilufar; Teichert, Carsten; Herold, Diana; Rogina, Blanka; Helfand, Stephen; Gollasch, Maik; Luft, Friedrich C; Aronson, Peter S

    2006-07-01

    A longevity gene called Indy (for 'I'm not dead yet'), with similarity to mammalian genes encoding sodium-dicarboxylate cotransporters, was identified in Drosophila melanogaster. Functional studies in Xenopus oocytes showed that INDY mediates the flux of dicarboxylates and citrate across the plasma membrane, but the specific transport mechanism mediated by INDY was not identified. To test whether INDY functions as an anion exchanger, we examined whether substrate efflux is stimulated by transportable substrates added to the external medium. Efflux of [14C]citrate from INDY-expressing oocytes was greatly accelerated by the addition of succinate to the external medium, indicating citrate-succinate exchange. The succinate-stimulated [14C]citrate efflux was sensitive to inhibition by DIDS (4,4'-di-isothiocyano-2,2'-disulphonic stilbene), as demonstrated previously for INDY-mediated succinate uptake. INDY-mediated efflux of [14C]citrate was also stimulated by external citrate and oxaloacetate, indicating citrate-citrate and citrate-oxaloacetate exchange. Similarly, efflux of [14C]succinate from INDY-expressing oocytes was stimulated by external citrate, alpha-oxoglutarate and fumarate, indicating succinate-citrate, succinate-alpha-oxoglutarate and succinate-fumarate exchange respectively. Conversely, when INDY-expressing Xenopus oocytes were loaded with succinate and citrate, [14C]succinate uptake was markedly stimulated, confirming succinate-succinate and succinate-citrate exchange. Exchange of internal anion for external citrate was markedly pH(o)-dependent, consistent with the concept that citrate is co-transported with a proton. Anion exchange was sodium-independent. We conclude that INDY functions as an exchanger of dicarboxylate and tricarboxylate Krebs-cycle intermediates. The effect of decreasing INDY activity, as in the long-lived Indy mutants, may be to alter energy metabolism in a manner that favours lifespan extension.

  18. Pyruvate dehydrogenase complex and lactate dehydrogenase as targets for therapy of acute liver failure.

    Science.gov (United States)

    Ferriero, Rosa; Nusco, Edoardo; De Cegli, Rossella; Carissimo, Annamaria; Manco, Giuseppe; Brunetti-Pierri, Nicola

    2018-03-23

    Acute liver failure is a rapidly progressive deterioration of hepatic function resulting in high mortality and morbidity. Metabolic enzymes can translocate in the nucleus to regulate histone acetylation and gene expression. Levels and activities of pyruvate dehydrogenase complex (PDHC) and lactate dehydrogenase (LDH) were evaluated in nuclear fractions of livers of mice exposed to various hepatotoxins including CD95-Ab, α-amanitin, and acetaminophen. Whole-genome gene expression profiling by RNA-seq was performed in livers of mice with acute liver failure and analyzed by Gene Ontology Enrichment Analysis. Efficacy of histone acetyltransferase inhibitor garcinol and LDH inhibitor galloflavin at reducing liver damage was evaluated in mice with induced hepatotoxicity. Levels and activities of PDHC and LDH were increased in cytoplasmatic and nuclear fractions of livers of mice with acute liver failure. The increase of nuclear PDHC and LDH was associated with increased concentrations of acetyl-coA and lactate in nuclear fractions, and histone H3 hyper-acetylation. Gene expression in livers of mice with acute liver failure suggested that increased histone H3 acetylation induces the expression of genes related to response to damage. Reduced histone acetylation by the histone acetyltransferase inhibitor garcinol decreased liver damage and improved survival in mice with acute liver failure. Knock-down of PDHC or LDH improved viability in cells exposed to a pro-apoptotic stimulus. Treatment with the LDH inhibitor galloflavin that was also found to inhibit PDHC, reduced hepatic necrosis, apoptosis, and expression of pro-inflammatory cytokines in mice with acute liver failure. Mice treated with galloflavin also showed a dose-response increase in survival. PDHC and LDH translocate to the nucleus and are targets for therapy of acute liver failure. Acute liver failure is a rapidly progressive and life-threatening deterioration of liver function resulting in high mortality and

  19. Mitochondrial and Nuclear Genes of Mitochondrial Components in Cancer

    Science.gov (United States)

    Kirches, E

    2009-01-01

    Although the observation of aerobic glycolysis of tumor cells by Otto v. Warburg had demonstrated abnormalities of mitochondrial energy metabolism in cancer decades ago, there was no clear evidence for a functional role of mutant mitochondrial proteins in cancer development until the early years of the 21st century. In the year 2000, a major breakthrough was achieved by the observation, that several genes coding for subunits of the respiratory chain (ETC) complex II, succinate dehydrogenase (SDH) are tumor suppressor genes in heritable paragangliomas, fulfilling Knudson’s classical two-hit hypothesis. A functional inactivation of both alleles by germline mutations and chromosomal losses in the tumor tissue was found in the patients. Later, SDH mutations were also identified in sporadic paragangliomas and pheochromocytomas. Genes of the mitochondrial ATP-synthase and of mitochondrial iron homeostasis have been implicated in cancer development at the level of cell culture and mouse experiments. In contrast to the well established role of some nuclear SDH genes, a functional impact of the mitochondrial genome itself (mtDNA) in cancer development remains unclear. Nevertheless, the extremely high frequency of mtDNA mutations in solid tumors raises the question, whether this small circular genome might be applicable to early cancer detection. This is a meaningful approach, especially in cancers, which tend to spread tumor cells early into bodily fluids or faeces, which can be screened by non-invasive methods. PMID:19949549

  20. 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

  1. Extraction chromatography of quadrivalent titanium and zirconium from succinate solution

    International Nuclear Information System (INIS)

    Shete, S.D.; Shinde, V.M.

    1982-01-01

    Liquid ion exchangers such as tri-n-octyl amine (TOA), tri-iso-octylamine (TIOA) and Aliquat 336 have been used in this laboratory for analytical separation of vanadium(V) and niobium(V), indium(III), thalium(III), thorium(IV), cerium(IV) from succinate solutions. Extension of this study has revealed that benzene solutions of TOA, TIOA and Aliquat 336 could be used for anion exchange extraction of quadrivalent titanium and zirconium from succinate solution. The metal ions from the organic phase are stripped and determined spectrophotometrically. Solvent extraction methods for titanium(IV) and zirconium(IV) have been reviewed by De et al. and Korkisch in their monographs. Diethylamine, tri-ethylamine and tributylamine have been used for extraction of titanium from thiocyanate and citrate solution. Similarly high molecular weight amines such as TOA and TIOA have been used for the extraction of zirconium from mineral acids, but systematic solvent extraction separation of titanium(IV) and zirconium(IV) is lacking. In this communication we propose a new method for selective extraction and separation of titaium(IV) and zirconium(IV) from metal ions such as Mn, Cr, Mo, V, U, La, Ta, Th and Hf. The proposed method is comparatively free from drawbacks such as pre-equilibration of phases, multiple scrubbing, multiple extraction and coextraction of a large number of cations and anions

  2. Deliquescence of malonic, succinic, glutaric, and adipic acid particles

    Science.gov (United States)

    Parsons, Matthew T.; Mak, Jackson; Lipetz, Sarah R.; Bertram, Allan K.

    2004-03-01

    In order to understand and predict the role of organic particles in the atmosphere their deliquescence behavior must be understood. Using an optical microscope coupled to a flow cell, we investigated the deliquescence of malonic, succinic, glutaric, and adipic acid particles with sizes ranging from 2 to 40 μm. Deliquescence relative humidities were determined for temperatures ranging from 293 to 243 K. Over this temperature range both succinic acid and adipic acid deliquesced at approximately 100% relative humidity, whereas malonic acid and glutaric acid deliquesced at significantly lower relative humidities. These results are generally in good agreement with previous studies and are within 3% of calculations based on the UNIQUAC (universal quasi-chemical) Functional Group Activity Coefficients (UNIFAC) model and recently published interaction parameters. Our studies also include measurements at temperatures below the eutectic temperatures. At these temperatures, ice did not nucleate; rather the particles underwent deliquescence to form metastable solution droplets. This indicates that solid dicarboxylic acids are not good ice nuclei above 243 K and hence will probably not play a role in ice cloud formation at these temperatures.

  3. Vitamin E Succinate as an Adjuvant for Dendritic Cell Based Vaccines

    National Research Council Canada - National Science Library

    Ramanathapuram, Lalitha V

    2006-01-01

    .... Vitamin E succinate or alpha tocopheryl succinate ( -TOS) is a non-toxic, esterified analogue of Vitamin E that has been shown to be selectively toxic to tumor cell lines in vitro as well as inhibit the growth of tumors in animal models in vivo...

  4. Vitamin E Succinate as an Adjuvant for Dendritic Cell Based Vaccines

    National Research Council Canada - National Science Library

    Ramanathapuram, Lalitha

    2004-01-01

    .... In this study we have employed Vitamin E succinate also known as alpha-tocopheryl succinate (alpha-TOS), a non-toxic esterified analogue of Vitamin E, as an adjuvant to enhance the effectiveness of DC vaccines in treating established murine mammary...

  5. Vitamin E Succinate as an Adjuvant for Dendritic Cell-Based Vaccines

    National Research Council Canada - National Science Library

    Ramanathapuram, Lalitha V; Akporiaye, Emmanuel T

    2005-01-01

    .... Vitamin E succinate or alpha tocopheryl succinate (a-TOS) is a non-toxic, esterified analogue of Vitamin E that has been shown to be selectively toxic to tumor cell lines in vitro as well as inhibit the growth of tumors in animal models in vivo...

  6. Evaporation of methyl- and dimethyl-substituted malonic, succinic, glutaric and adipic acid particles at ambient temperatures

    DEFF Research Database (Denmark)

    Mønster, Jacob Garbrecht; Rosenørn, Thomas; Svenningsson, Birgitta

    2004-01-01

    Evaporation; organic aerosols; vapor pressure; dicarboxylic acid; maonic acid; succinic acid; glutaric acid; adipic acid......Evaporation; organic aerosols; vapor pressure; dicarboxylic acid; maonic acid; succinic acid; glutaric acid; adipic acid...

  7. Molecular characterization and detection of mutations associated with resistance to succinate dehydrogenase inhibiting (SDHI) fungicides in Alternaria solani

    Science.gov (United States)

    Early blight, caused by Alternaria solani, is an economically important foliar disease of potato in several production areas of the United States. Few potato cultivars possess resistance to early blight, therefore, the application of fungicides is a primary means of achieving disease control. Previo...

  8. Structures of Tris(2-hydroxyethyl)ammonium Salts of Succinic Acid

    Science.gov (United States)

    Loginov, S. V.; Dain, I. A.; Rybakov, V. B.; Ofitserov, E. N.; Gordeev, D. A.; Storozhenko, P. A.

    2018-01-01

    Succinic acid salts-tris(2-hydroxyethyl)ammonium succinate (C6H16NO3) 2 + C4H4O 4 2- (monoclinic crystals, sp. gr. P21/ c, Z = 4) and tris(2-hydroxyethyl)ammonium hydrogen succinate (C6H16NO3)+C4H5O 4 - (monoclinic crystals, sp. gr. P21/ c, Z = 4)—were synthesized and structurally characterized. The specific features of the three-dimensional structures of tris(2-hydroxyethyl)ammonium salts of succinic acid are considered. The role of interionic electrostatic interactions in the structure stabilization and the formation of products of composition 1: 1 and 1: 2 derived from succinic acid is discussed.

  9. Thermochemical pretreatments for enhancing succinic acid production from industrial hemp (Cannabis sativa L.).

    Science.gov (United States)

    Gunnarsson, Ingólfur B; Kuglarz, Mariusz; Karakashev, Dimitar; Angelidaki, Irini

    2015-04-01

    The aim of this study was to develop an efficient thermochemical method for treatment of industrial hemp biomass, in order to increase its bioconversion to succinic acid. Industrial hemp was subjected to various thermochemical pretreatments using 0-3% H2SO4, NaOH or H2O2 at 121-180°C prior to enzymatic hydrolysis. The influence of the different pretreatments on hydrolysis and succinic acid production by Actinobacillus succinogenes 130Z was investigated in batch mode, using anaerobic bottles and bioreactors. Enzymatic hydrolysis and fermentation of hemp material pretreated with 3% H2O2 resulted in the highest overall sugar yield (73.5%), maximum succinic acid titer (21.9 g L(-1)), as well as the highest succinic acid yield (83%). Results obtained clearly demonstrated the impact of different pretreatments on the bioconversion efficiency of industrial hemp into succinic acid. Copyright © 2015. Published by Elsevier Ltd.

  10. Thermochemical pretreatments for enhancing succinic acid production from industrial hemp (Cannabis sativa L.)

    DEFF Research Database (Denmark)

    Gunnarsson, Ingólfur Bragi; Kuglarz, Mariusz; Karakashev, Dimitar Borisov

    2015-01-01

    The aim of this study was to develop an efficient thermochemical method for treatment of industrial hemp biomass, in order to increase its bioconversion to succinic acid. Industrial hemp was subjected to various thermochemical pretreatments using 0-3% H2SO4, NaOH or H2O2 at 121-180°C prior...... to enzymatic hydrolysis. The influence of the different pretreatments on hydrolysis and succinic acid production by Actinobacillus succinogenes 130Z was investigated in batch mode, using anaerobic bottles and bioreactors. Enzymatic hydrolysis and fermentation of hemp material pretreated with 3% H2O2 resulted...... in the highest overall sugar yield (73.5%), maximum succinic acid titer (21.9gL-1), as well as the highest succinic acid yield (83%). Results obtained clearly demonstrated the impact of different pretreatments on the bioconversion efficiency of industrial hemp into succinic acid....

  11. Common missense mutation G1528C in long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency. Characterization and expression of the mutant protein, mutation analysis on genomic DNA and chromosomal localization of the mitochondrial trifunctional protein alpha subunit gene

    NARCIS (Netherlands)

    IJlst, L.; Ruiter, J. P.; Hoovers, J. M.; Jakobs, M. E.; Wanders, R. J.

    1996-01-01

    Mitochondrial trifunctional protein (MTP) is a recently identified enzyme involved in mitochondrial beta-oxidation, harboring long-chain enoyl-CoA hydratase, long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) and long-chain 3-ketothiolase activity. A deficiency of this protein is associated with

  12. Cloning and characterization of human very-long-chain acyl-CoA dehydrogenase cDNA, chromosomal assignment of the gene and identification in four patients of nine different mutations within the VLCAD gene

    DEFF Research Database (Denmark)

    Andresen, B S; Bross, P; Vianey-Saban, C

    1996-01-01

    specific distribution of VLCAD mRNA and protein in several human tissues we showed that VLCAD is most abundant in heart and skeletal muscle. This agrees well with the fact that cardiac and muscle symptoms are characteristic for patients with VLCAD deficiency. Northern blot analysis and sequencing of cloned......) based strategy to obtain the sequence of cDNAs encoding human VLCAD from placenta and fibroblasts. Alignment of the predicted amino acid sequence of human VLCAD with those of the other human ACD enzymes revealed extensive sequence homology. Moreover, human VLCAD and human acyl-CoA oxidase showed...... extensive sequence homology corroborating the notion that these genes are evolutionarily related. Southern blot analysis of genomic DNA from hybrid cell lines was used to localize the VLCAD gene to human chromosome 17p11.2-p11.13105. Using Northern and Western blot analysis to investigate the tissue...

  13. Genetics Home Reference: dihydropyrimidine dehydrogenase deficiency

    Science.gov (United States)

    ... 5-fluorouracil and capecitabine. These drugs are not broken down efficiently by people with dihydropyrimidine dehydrogenase deficiency ... of this enzyme. Because fluoropyrimidine drugs are also broken down by the dihydropyrimidine dehydrogenase enzyme, deficiency of ...

  14. Target gene approaches: Gene expression in Daphnia magna exposed to predator-borne kairomones or to microcystin-producing and microcystin-free Microcystis aeruginosa

    Directory of Open Access Journals (Sweden)

    Courts Cornelius

    2009-11-01

    Full Text Available Abstract Background Two major biological stressors of freshwater zooplankton of the genus Daphnia are predation and fluctuations in food quality. Here we use kairomones released from a planktivorous fish (Leucaspius delineatus and from an invertebrate predator (larvae of Chaoborus flavicans to simulate predation pressure; a microcystin-producing culture of the cyanobacterium Microcystis aeruginosa and a microcystin-deficient mutant are used to investigate effects of low food quality. Real-time quantitative polymerase chain reaction (QPCR allows quantification of the impact of biotic stressors on differential gene activity. The draft genome sequence for Daphnia pulex facilitates the use of candidate genes by precisely identifying orthologs to functionally characterized genes in other model species. This information is obtained by constructing phylogenetic trees of candidate genes with the knowledge that the Daphnia genome is composed of many expanded gene families. Results We evaluated seven candidate reference genes for QPCR in Daphnia magna after exposure to kairomones. As a robust approach, a combination normalisation factor (NF was calculated based on the geometric mean of three of these seven reference genes: glyceraldehyde-3-phosphate dehydrogenase, TATA-box binding protein and succinate dehydrogenase. Using this NF, expression of the target genes actin and alpha-tubulin were revealed to be unchanged in the presence of the tested kairomones. The presence of fish kairomone up-regulated one gene (cyclophilin involved in the folding of proteins, whereas Chaoborus kairomone down-regulated the same gene. We evaluated the same set of candidate reference genes for QPCR in Daphnia magna after exposure to a microcystin-producing and a microcystin-free strain of the cyanobacterium Microcystis aeruginosa. The NF was calculated based on the reference genes 18S ribosomal RNA, alpha-tubulin and TATA-box binding protein. We found glyceraldehyde-3

  15. Succinate modulates Ca(2+) transient and cardiomyocyte viability through PKA-dependent pathway.

    Science.gov (United States)

    Aguiar, Carla J; Andrade, Vanessa L; Gomes, Enéas R M; Alves, Márcia N M; Ladeira, Marina S; Pinheiro, Ana Cristina N; Gomes, Dawidson A; Almeida, Alvair P; Goes, Alfredo M; Resende, Rodrigo R; Guatimosim, Silvia; Leite, M Fatima

    2010-01-01

    GPR91 is an orphan G-protein-coupled receptor (GPCR) that has been characterized as a receptor for succinate, a citric acid cycle intermediate, in several tissues. In the heart, the role of succinate is unknown. We now report that rat ventricular cardiomyocytes express GPR91. We found that succinate, through GPR91, increases the amplitude and the rate of decline of global Ca(2+) transient, by increasing the phosphorylation levels of ryanodine receptor and phospholamban, two well known Ca(2+) handling proteins. The effects of succinate on Ca(2+) transient were abolished by pre-treatment with adenylyl cyclase and cAMP-dependent protein kinase (PKA) inhibitors. Direct PKA activation by succinate was further confirmed using a FRET-based A-kinase activity reporter. Additionally, succinate decreases cardiomyocyte viability through a caspase-3 activation pathway, effect also prevented by PKA inhibition. Taken together, these observations show that succinate acts as a signaling molecule in cardiomyocytes, modulating global Ca(2+) transient and cell viability through a PKA-dependent pathway. 2009 Elsevier Ltd. All rights reserved.

  16. Enhanced succinic acid production from corncob hydrolysate by microbial electrolysis cells.

    Science.gov (United States)

    Zhao, Yan; Cao, Weijia; Wang, Zhen; Zhang, Bowen; Chen, Kequan; Ouyang, Pingkai

    2016-02-01

    In this study, Actinobacillus succinogenes NJ113 microbial electrolysis cells (MECs) were used to enhance the reducing power responsible for succinic acid production from corncob hydrolysate. During corncob hydrolysate fermentation, electric MECs resulted in a 1.31-fold increase in succinic acid production and a 1.33-fold increase in the reducing power compared with those in non-electric MECs. When the hydrolysate was detoxified by combining Ca(OH)2, NaOH, and activated carbon, succinic acid production increased from 3.47 to 6.95 g/l. Using a constant potential of -1.8 V further increased succinic acid production to 7.18 g/l. A total of 18.09 g/l of succinic acid and a yield of 0.60 g/g total sugar were obtained after a 60-h fermentation when NaOH was used as a pH regulator. The improved succinic acid yield from corncob hydrolysate fermentation using A. succinogenes NJ113 in electric MECs demonstrates the great potential of using biomass as a feedstock to cost-effectively produce succinate. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. Study of the radiolysis of succinic acid - applications in the dosimetry of high doses

    International Nuclear Information System (INIS)

    Andrade e Silva, L.G.

    1978-01-01

    A study is made of the effect of the gama radiation dose and of particle size of succinic acid (fine powder of large crystals) in relation with the formation of CO 2 and CO + H 2 , which are the main gaseous products of radiolysis. A different yield of CO + H 2 is found when the succinic acid is used as powder compared to the material in the form of large crystals. The reason for this difference is searched, studying the influence of heating and sublimation of the succinic acid prior to irradiation. The influence, in the mentioned yield, of the surface area of succinic acid particles, of the presence of oxygen (air) and of the rapid recrystallization of the acid are also studied. The formation of intermediate species in the radiolysis of succinic acid is examined. The system used in ethanol-succinic acid at 77K. Analysis are made using an electronic paramagnetic resonance spectrometer. The possibility of using succinic acid as a dosimeter for high level gama radiation doses is discussed [pt

  18. Thermodynamic and Thermomicroscopy Study of Atorvastatin Calcium-Succinic Acid Binary Mixtures

    Directory of Open Access Journals (Sweden)

    Yudi Wicaksono

    2017-11-01

    Full Text Available Binary mixtures of pharmaceuticals significantly affect the physical and chemical properties of each component. The aim of this work was to explore the thermal behavior and solid state transformation of binary mixture of atorvastatin calcium and succinic acid. The thermodynamics of binary mixtures of atorvastatin calcium - succinic acid were determined by differential scanning calorimeter. Meanwhile, thermomicroscopy and microstructure were determined by a polarized microscope equipped with a heating stage and camera. The results showed that melting points of atorvastatin calcium and succinic acid respectively were 159.35 and 188.51 °C. The solid-liquid phase diagram of atorvastatin calcium - succinic acid indicates the existence of two eutectic points at 136.57 °C and 120.96 °C respectively on the mole fraction of atorvastatin calcium 0.3 and 0.5. Tamman diagram accurately shows mole fraction of atorvastatin calcium at eutectic point 0.33 and 0.46 respectively for eutectic points 130.0 °C and 134.0 °C. Determination of Jackson’s roughness parameter showed a value of atorvastatin calcium, succinic acid and eutectic mixtures > 2 which indicates that the interfaces of remelting crystals were smooth. Microstructure of remelting crystal of atorvastatin calcium and succinic acid respectively was irregular form and crossed plates. The results of thermomicroscopy of binary mixtures of atorvastatin calcium-succinic acid were consistent with differential scanning calorimetry curves and solid-liquid phase diagram.

  19. The succinate receptor as a novel therapeutic target for oxidative and metabolic stress-related conditions.

    Directory of Open Access Journals (Sweden)

    Ana Carolina eAriza

    2012-02-01

    Full Text Available The succinate receptor (also known as GPR91 is a G protein-coupled receptor that is closely related to the family of P2Y purinoreceptors. It is expressed in a variety of tissues, including blood cells, adipose tissue, the liver, retina and kidney. In these tissues, this receptor and its ligand succinate have recently emerged as novel mediators in local stress situations, including ischemia, hypoxia, toxicity and hyperglycemia. Amongst others, the succinate receptor is involved in recruitment of immune cells to transplanted tissues. Moreover, it was shown to play a key role in the development of diabetic retinopathy. However, most prominently, the role of locally increased succinate levels and succinate receptor activation in the kidney, stimulating the systemic and local renin-angiotensin system, starts to unfold: The succinate receptor is a key mediator in the development of hypertension and possibly fibrosis in diabetes mellitus and metabolic syndrome. This makes the succinate receptor a promising drug target to counteract or prevent cardiovascular and fibrotic defects in these expanding disorders. Recent development of SUCNR1-specific antagonists opens novel possibilities for research in models for these disorders and may eventually provide novel opportunities for the treatment of patients.

  20. Expression of Ascaris suum malic enzyme in a mutant Escherichia coli allows production of succinic acid from glucose

    Energy Technology Data Exchange (ETDEWEB)

    Stols, L.; Donnelly, M.I. [Argonne National Lab., IL (United States); Kulkarni, G.; Harris, B.G. [Univ. of North Texas, Fort Worth, TX (United States)

    1997-12-31

    The malic enzyme gene of Ascaris suum was cloned into the vector pTRC99a in two forms encoding alternative amino-termini. The resulting plasmids, pMEA1 and pMEA2, were introduced into Escherichia coli NZN111, a strain that is unable to grow fermentatively because of inactivation of the genes encoding pyruvate dissimilation. Induction of pMEA1, which encodes the native animoterminus, gave better overexpression of malic enzyme, approx 12-fold compared to uninduced cells. Under the appropriate culture conditions, expression of malic enzyme allowed the fermentative dissimilation of glucose by NZN111. The major fermentation product formed in induced cultures was succinic acid.

  1. Interaction of Palmitic Acid with Metoprolol Succinate at the Binding Sites of Bovine Serum Albumin

    Directory of Open Access Journals (Sweden)

    Mashiur Rahman

    2014-12-01

    Full Text Available Purpose: The aim of this study was to characterize the binding profile as well as to notify the interaction of palmitic acid with metoprolol succinate at its binding site on albumin. Methods: The binding of metoprolol succinate to bovine serum albumin (BSA was studied by equilibrium dialysis method (ED at 27°C and pH 7.4, in order to have an insight in the binding chemistry of the drug to BSA in presence and absence of palmitic acid. The study was carried out using ranitidine as site-1 and diazepam as site-2 specific probe. Results: Different analysis of binding of metoprolol succinate to bovine serum albumin suggested two sets of association constants: high affinity association constant (k1 = 11.0 x 105 M-1 with low capacity (n1 = 2 and low affinity association (k2 = 4.0×105 M-1 constant with high capacity (n2 = 8 at pH 7.4 and 27°C. During concurrent administration of palmitic acid and metoprolol succinate in presence or absence of ranitidine or diazepam, it was found that palmitic acid displaced metoprolol succinate from its binding site on BSA resulting reduced binding of metoprolol succinate to BSA. The increment in free fraction of metoprolol succinate was from 26.27% to 55.08% upon the addition of increased concentration of palmitic acid at a concentration of 0×10-5 M to 16×10-5 M. In presence of ranitidine and diazepam, palmitic acid further increases the free fraction of metoprolol succinate from 33.05% to 66.95% and 40.68% to 72.88%, respectively. Conclusion: This data provided the evidence of interaction at higher concentration of palmitic acid at the binding sites on BSA, which might change the pharmacokinetic properties of metoprolol succinate.

  2. Identification of the human mitochondrial FAD transporter and its potential role in multiple acyl-CoA dehydrogenase deficiency

    NARCIS (Netherlands)

    Spaan, András N.; Ijlst, Lodewijk; van Roermund, Carlo W. T.; Wijburg, Frits A.; Wanders, Ronald J. A.; Waterham, Hans R.

    2005-01-01

    Multiple acyl-CoA dehydrogenase deficiency (MADD) or glutaric aciduria type II (GAII) is most often caused by mutations in the genes encoding the alpha- or beta-subunit of electron transfer flavoprotein (ETF) or electron transfer flavoprotein dehydrogenase (ETF-DH). Since not all patients have

  3. Receptor structure-based discovery of non-metabolite agonists for the succinate receptor GPR91

    DEFF Research Database (Denmark)

    Trauelsen, Mette; Rexen Ulven, Elisabeth; Hjorth, Siv A

    2017-01-01

    OBJECTIVE: Besides functioning as an intracellular metabolite, succinate acts as a stress-induced extracellular signal through activation of GPR91 (SUCNR1) for which we lack suitable pharmacological tools. METHODS AND RESULTS: Here we first determined that the cis conformation of the succinate...... backbone is preferred and that certain backbone modifications are allowed for GPR91 activation. Through receptor modeling over the X-ray structure of the closely related P2Y1 receptor, we discovered that the binding pocket is partly occupied by a segment of an extracellular loop and that succinate...

  4. Integrated production of cellulosic bioethanol and succinic acid from industrial hemp in a biorefinery concept

    DEFF Research Database (Denmark)

    Kuglarz, Mariusz; Alvarado-Morales, Merlin; Karakashev, Dimitar Borisov

    2016-01-01

    The aim of this study was to develop integrated biofuel (cellulosic bioethanol) and biochemical (succinic acid) production from industrial hemp (Cannabis sativa L.) in a biorefinery concept. Two types of pretreatments were studied (dilute-acid and alkaline oxidative method). High cellulose recovery...... productivity. With respect to succinic acid production, the highest productivity was obtained after liquid fraction fermentation originated from steam treatment with 1.5% of acid. The mass balance calculations clearly showed that 149 kg of EtOH and 115 kg of succinic acid can be obtained per 1 ton of dry hemp....... Results obtained in this study clearly document the potential of industrial hemp for a biorefinery....

  5. CRISPR-Cas9 for the genome engineering of cyanobacteria and succinate production.

    Science.gov (United States)

    Li, Hung; Shen, Claire R; Huang, Chun-Hung; Sung, Li-Yu; Wu, Meng-Ying; Hu, Yu-Chen

    2016-11-01

    Cyanobacteria hold promise as a cell factory for producing biofuels and bio-derived chemicals, but genome engineering of cyanobacteria such as Synechococcus elongatus PCC 7942 poses challenges because of their oligoploidy nature and long-term instability of the introduced gene. CRISPR-Cas9 is a newly developed RNA-guided genome editing system, yet its application for cyanobacteria engineering has yet to be reported. Here we demonstrated that CRISPR-Cas9 system can effectively trigger programmable double strand break (DSB) at the chromosome of PCC 7942 and provoke cell death. With the co-transformation of template plasmid harboring the gene cassette and flanking homology arms, CRISPR-Cas9-mediated DSB enabled precise gene integration, ameliorated the homologous recombination efficiency and allowed the use of lower amount of template DNA and shorter homology arms. The CRISPR-Cas9-induced cell death imposed selective pressure and enhanced the chance of concomitant integration of gene cassettes into all chromosomes of PCC 7942, hence accelerating the process of obtaining homogeneous and stable recombinant strains. We further explored the feasibility of engineering cyanobacteria by CRISPR-Cas9-assisted simultaneous glgc knock-out and gltA/ppc knock-in, which improved the succinate titer to 435.0±35.0μg/L, an ≈11-fold increase when compared with that of the wild-type cells. These data altogether justify the use of CRISPR-Cas9 for genome engineering and manipulation of metabolic pathways in cyanobacteria. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  6. In-vitro responses of T lymphocytes to poly(butylene succinate) based biomaterials.

    Science.gov (United States)

    Toso, Montree; Patntirapong, Somying; Janvikul, Wanida; Singhatanadgit, Weerachai

    2017-04-01

    Polybutylene succinate (PBSu) and PBSu/β-tricalcium phosphate (TCP) composites are biocompatible and good candidates as bone graft materials. However, little is known about the responses of T lymphocytes to these biomaterials, which play an important role in the success of bone grafting. Activated T lymphocytes were cultured onto 32 mm diameter films (PBSu/TCP films), that had previously been placed in 6-well culture plates, for 8, 24 and 72 hours. A plastic-well culture plate was used as a control surface. The effects of PBSu-based biomaterials on T lymphocytes were examined by the using flow cytometry and reverse-transcription polymerase chain reaction. These biomaterials were non-toxic to T lymphocytes, allowing their normal DNA synthesis and activation. All materials induced only transient activation of T lymphocytes, which existed no longer than 72 hours. Proportions of four main CD4/CD8 T lymphocyte subpopulations were not affected by these biomaterials. Moreover, PBSu and PBSu/TCP significantly suppressed the expression of IL-1β and IL-6 genes by 15-35% and 21-26%, respectively. In contrast, a PBSu/TCP composite (at PBSu:TCP=60:40) significantly stimulated the expression of IL-10 and IL-13 genes by 17% and 19%, respectively. PBSu and PBSu/TCP composites were non-toxic to T lymphocytes and did not induce unfavorable responses of T lymphocytes. The tested biomaterials down-regulated key proinflammatory cytokine genes and up-regulated anti-inflammatory cytokine genes in T lymphocytes. These suggest that the biomaterials studied are good candidates as bone graft materials.

  7. Effects of TCDD on the expression of nuclear encoded mitochondrial genes

    International Nuclear Information System (INIS)

    Forgacs, Agnes L.; Burgoon, Lyle D.; Lynn, Scott G.; LaPres, John J.; Zacharewski, Timothy

    2010-01-01

    Generation of mitochondrial reactive oxygen species (ROS) can be perturbed following exposure to environmental chemicals such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Reports indicate that the aryl hydrocarbon receptor (AhR) mediates TCDD-induced sustained hepatic oxidative stress by decreasing hepatic ATP levels and through hyperpolarization of the inner mitochondrial membrane. To further elucidate the effects of TCDD on the mitochondria, high-throughput quantitative real-time PCR (HTP-QRTPCR) was used to evaluate the expression of 90 nuclear genes encoding mitochondrial proteins involved in electron transport, oxidative phosphorylation, uncoupling, and associated chaperones. HTP-QRTPCR analysis of time course (30 μg/kg TCDD at 2, 4, 8, 12, 18, 24, 72, and 168 h) liver samples obtained from orally gavaged immature, ovariectomized C57BL/6 mice identified 54 differentially expressed genes (|fold change| > 1.5 and P-value < 0.1). Of these, 8 exhibited a sigmoidal or exponential dose-response profile (0.03 to 300 μg/kg TCDD) at 4, 24 or 72 h. Dose-responsive genes encoded proteins associated with electron transport chain (ETC) complexes I (NADH dehydrogenase), III (cytochrome c reductase), IV (cytochrome c oxidase), and V (ATP synthase) and could be generally categorized as having proton gradient, ATP synthesis, and chaperone activities. In contrast, transcript levels of ETC complex II, succinate dehydrogenase, remained unchanged. Putative dioxin response elements were computationally found in the promoter regions of all 8 dose-responsive genes. This high-throughput approach suggests that TCDD alters the expression of genes associated with mitochondrial function which may contribute to TCDD-elicited mitochondrial toxicity.

  8. The neuronal insulin sensitizer dicholine succinate reduces stress-induced depressive traits and memory deficit: possible role of insulin-like growth factor 2

    Directory of Open Access Journals (Sweden)

    Cline Brandon H

    2012-09-01

    Full Text Available Abstract Background A number of epidemiological studies have established a link between insulin resistance and the prevalence of depression. The occurrence of depression was found to precede the onset of diabetes and was hypothesized to be associated with inherited inter-related insufficiency of the peripheral and central insulin receptors. Recently, dicholine succinate, a sensitizer of the neuronal insulin receptor, was shown to stimulate insulin-dependent H2O2 production of the mitochondrial respiratory chain leading to an enhancement of insulin receptor autophosphorylation in neurons. As such, this mechanism can be a novel target for the elevation of insulin signaling. Results Administration of DS (25 mg/kg/day, intraperitoneal in CD1 mice for 7 days prior to the onset of stress procedure, diminished manifestations of anhedonia defined in a sucrose test and behavioral despair in the forced swim test. Treatment with dicholine succinate reduced the anxiety scores of stressed mice in the dark/light box paradigm, precluded stress-induced decreases of long-term contextual memory in the step-down avoidance test and hippocampal gene expression of IGF2. Conclusions Our data suggest that dicholine succinate has an antidepressant-like effect, which might be mediated via the up-regulation of hippocampal expression of IGF2, and implicate the neuronal insulin receptor in the pathogenesis of stress-induced depressive syndrome.

  9. Metabolic Effects of a Succinic Acid

    Directory of Open Access Journals (Sweden)

    B. N. Shakh

    2014-01-01

    Full Text Available The paper discusses promises for clinical use of substrate antihypoxants.Objective: to investigate the efficacy of succinate containing  substrate  antihypoxants  on  systemic  oxygen  consumption,  blood  buffer  capacity,  and  changes  in  the  mixed venous blood level of lactate when they are used in gravely sick patients and victims with marked metabolic posthypoxic disorders.Subjects and methods. The trial enrolled 30 patients and victims who had sustained an episode of severe hypoxia of mixed genesis, the severity of which was evaluated by the APACHE II scale and amounted to 23 to 30 scores with a 46 to 70.3% risk of death. The standard infusion program in this group involved the succinate-containing drug 1.5% reamberin solution  in  a  total  dose  of  800  ml.  A  comparison  group  included  15  patients  who  had  undergone  emergency  extensive surgery for abdominal diseases. 400 ml of 10% glucose solution was used as an infusion medium. Oxygen consumption (VO2ml/min and carbon dioxide production (VCO2ml/min were measured before infusion and monitored for 2 hours. Arterial blood gases and acid-base balance (ABB parameters and mixed venous blood lactate levels were examined. Measurements were made before and 30 minutes after the infusion of reamberin or glucose solution.Results. Infusion of 1.5% reamberin solution was followed by a significant increase in minute oxygen consumption from 281.5±21.2 to 310.4±24.4 ml/min. CO2 production declined (on average, from 223.3±6.5 to 206.5±7.59 ml/min. During infusion of 10% glucose solution, all the patients of the comparison group showed a rise in oxygen consumption from 303.6±33.86 to 443.13±32.1 ml/min, i.e. about 1.5-fold. VCO2 changed similarly. The intravenous infusion of 800 ml of 1.5% reamberin solution raised arterial blood buffer capacity, which was reflected by changes in pH, BE, and HCO3. There was a clear trend for lactate values to drop in the

  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. Multifunctional nanobiocomposite of Poly[(butylene succinate)-co-adipate] and clay

    CSIR Research Space (South Africa)

    Al-Thabaiti, SA

    2015-03-01

    Full Text Available The processing and characterization of multifunctional nanobiocomposite of biodegradable poly[(butylene succinate)-co-adipate] (PBSA) and organically modified synthetic fluorine mica (OSFM) are reported. The nanobiocomposite of PBSA with OSFM...

  12. An L-glucitol oxidizing dehydrogenase from Bradyrhizobium japonicum USDA 110 for production of D-sorbose with enzymatic or electrochemical cofactor regeneration

    DEFF Research Database (Denmark)

    Gauer, Sabrina; Wang, Zhijie; Otten, Harm

    2014-01-01

    A gene in Bradyrhizobium japonicum USDA 110, annotated as a ribitol dehydrogenase (RDH), had 87 % sequence identity (97 % positives) to the N-terminal 31 amino acids of an L-glucitol dehydrogenase from Stenotrophomonas maltophilia DSMZ 14322. The 729-bp long RDH gene coded for a protein consistin...

  13. Sustainability assessment of succinic acid production technologies from biomass using metabolic engineering

    OpenAIRE

    Morales M.; Ataman M.; Badr S.; Linster S.; Kourlimpinis I.; Papadokonstantakis S.; Hatzimanikatis V.; Hungerbühlera K.

    2016-01-01

    Over the past few years bio succinic acid from renewable resources has gained increasing attention as a potential bio derived platform chemical for the detergent/surfactant ion chelator food and pharmaceutical markets. Until now much research was undertaken to lower the production costs of bio succinic acid however a multicriteria sustainability evaluation of established and upcoming production processes from a technical perspective is still lacking in the scientific literature. In this study...

  14. Thermodynamic and Thermomicroscopy Study of Atorvastatin Calcium-Succinic Acid Binary Mixtures

    OpenAIRE

    Yudi Wicaksono; Budipratiwi Wisudyaningsih; Frida Oktaningtias Widiarthi; Tri Agus Siswoyo

    2017-01-01

    Binary mixtures of pharmaceuticals significantly affect the physical and chemical properties of each component. The aim of this work was to explore the thermal behavior and solid state transformation of binary mixture of atorvastatin calcium and succinic acid. The thermodynamics of binary mixtures of atorvastatin calcium - succinic acid were determined by differential scanning calorimeter. Meanwhile, thermomicroscopy and microstructure were determined by a polarized microscope equipped with a...

  15. A Relational Database for the Discovery of Genes Encoding Amino Acid Biosynthetic Enzymes in Pathogenic Fungi

    Directory of Open Access Journals (Sweden)

    Nicholas J. Talbot

    2006-04-01

    Full Text Available Fungal phytopathogens continue to cause major economic impact, either directly, through crop losses, or due to the costs of fungicide application. Attempts to understand these organisms are hampered by a lack of fungal genome sequence data. A need exists, however, to develop specific bioinformatics tools to collate and analyse the sequence data that currently is available. A web-accessible gene discovery database (http://cogeme.ex.ac.uk/biosynthesis.html was developed as a demonstration tool for the analysis of metabolic and signal transduction pathways in pathogenic fungi using incomplete gene inventories. Using Bayesian probability to analyse the currently available gene information from pathogenic fungi, we provide evidence that the obligate pathogen Blumeria graminis possesses all amino acid biosynthetic pathways found in free-living fungi, such as Saccharomyces cerevisiae. Phylogenetic analysis was also used to deduce a gene history of succinate-semialdehyde dehydrogenase, an enzyme in the glutamate and lysine biosynthesis pathways. The database provides a tool and methodology to researchers to direct experimentation towards predicting pathway conservation in pathogenic microorganisms.

  16. Biotechnological route for sustainable succinate production utilizing oil palm frond and kenaf as potential carbon sources.

    Science.gov (United States)

    Luthfi, Abdullah Amru Indera; Manaf, Shareena Fairuz Abdul; Illias, Rosli Md; Harun, Shuhaida; Mohammad, Abdul Wahab; Jahim, Jamaliah Md

    2017-04-01

    Due to the world's dwindling energy supplies, greater thrust has been placed on the utilization of renewable resources for global succinate production. Exploration of such biotechnological route could be seen as an act of counterbalance to the continued fossil fuel dominance. Malaysia being a tropical country stands out among many other nations for its plenty of resources in the form of lignocellulosic biomass. To date, oil palm frond (OPF) contributes to the largest fraction of agricultural residues in Malaysia, while kenaf, a newly introduced fiber crop with relatively high growth rate, holds great potential for developing sustainable succinate production, apart from OPF. Utilization of non-food, inexhaustible, and low-cost derived biomass in the form of OPF and kenaf for bio-based succinate production remains largely untapped. Owing to the richness of carbohydrates in OPF and kenaf, bio-succinate commercialization using these sources appears as an attractive proposition for future sustainable developments. The aim of this paper was to review some research efforts in developing a biorefinery system based on OPF and kenaf as processing inputs. It presents the importance of the current progress in bio-succinate commercialization, in addition to describing the potential use of different succinate production hosts and various pretreatments-saccharifications under development for OPF and kenaf. Evaluations on the feasibility of OPF and kenaf as fermentation substrates are also discussed.

  17. Production of succinic acid from oil palm empty fruit bunch cellulose using Actinobacillus succinogenes

    Science.gov (United States)

    Pasma, Satriani Aga; Daik, Rusli; Maskat, Mohamad Yusof

    2013-11-01

    Succinic acid is a common metabolite in plants, animals and microorganisms. It has been used widely in agricultural, food and pharmaceutical industries. Enzymatic hydrolysate glucose from oil palm empty fruit bunch (OPEFB) cellulose was used as a substrate for succinic acid production using Actinobacillus succinogenes. Using cellulose extraction from OPEFB can enhance the production of glucose as a main substrate for succinic acid production. The highest concentration of glucose produced from enzymatic hydrolysis is 167 mg/mL and the sugar recovery is 0.73 g/g of OPEFB. By optimizing the culture medium for succinic acid fermentation with enzymatic hydrolysate of OPEFB cellulose, the nitrogen sources could be reduced to just only 2.5 g yeast extract and 2.5 g corn step liquor. Batch fermentation was carried out using enzymatic hydrolysate of OPEFB cellulose with yeast extract, corn steep liquor and the salts mixture, 23.5 g/L succinic acid was obtained with consumption of 72 g/L glucose in enzymatic hydrolysate of OPEFB cellulose at 38 hours and 37°C. This study suggests that enzymatic hydrolysate of OPEFB cellulose maybe an alternative substrate for the efficient production of succinic acid by Actinobacillus succinogenes.

  18. Continuous and batch cultures of Escherichia coli KJ134 for succinic acid fermentation: metabolic flux distributions and production characteristics.

    Science.gov (United States)

    van Heerden, Carel D; Nicol, Willie

    2013-09-17

    Succinic acid (SA) has become a prominent biobased platform chemical with global production quantities increasing annually. Numerous genetically modified E. coli strains have been developed with the main aim of increasing the SA yield of the organic carbon source. In this study, a promising SA-producing strain, E. coli KJ134 [Biotechnol. Bioeng. 101:881-893, 2008], from the Department of Microbiology and Cell Science of the University of Florida was evaluated under continuous and batch conditions using D-glucose and CO2 in a mineral salt medium. Production characteristics entailing growth and maintenance rates, growth termination points and metabolic flux distributions under growth and non-growth conditions were determined. The culture remained stable for weeks under continuous conditions. Under growth conditions the redox requirements of the reductive tricarboxylic acid (TCA) cycle was solely balanced by acetic acid (AcA) production via the pyruvate dehydrogenase route resulting in a molar ratio of SA:AcA of two. A maximum growth rate of 0.22 h(-1) was obtained, while complete growth inhibition occurred at a SA concentration of 18 g L(-1). Batch culture revealed that high-yield succinate production (via oxidative TCA or glyoxylate redox balancing) occurred under non-growth conditions where a SA:AcA molar ratio of up to five was attained, with a final SA yield of 0.94 g g(-1). Growth termination of the batch culture was in agreement with that of the continuous culture. The maximum maintenance production rate of SA under batch conditions was found to be 0.6 g g(-1) h(-1). This is twice the maintenance rate observed in the continuous runs. The study revealed that the metabolic flux of E. coli KJ134 differs significantly for growth and non-growth conditions, with non-growth conditions resulting in higher SA:AcA ratios and SA yields. Bioreaction characteristics entailing growth and maintenance rates, as well as growth termination markers will guide future fermentor

  19. Biological denitrification using poly(butanediol succinate) as electron donor.

    Science.gov (United States)

    Shen, Zhiqiang; Yin, Yanan; Wang, Jianlong

    2016-07-01

    Poly(butanediol succinate) (PBS), a biodegradable polymer, was used as both solid carbon source and biofilm carrier for biological nitrate removal process, in which PBS was filled in a packed-bed bioreactor. The denitrification performance and the microbial diversity of biofilm attached on the surface of PBS were investigated. The experimental results showed that the volumetric denitrification rate was 0.60 kg m(-3) day(-1) when NO3-N loading rate was 0.63 kg m(-3) day(-1), and the average NO2-N concentration was below 0.20 mg L(-1). The effluent pH value decreased slightly from a range of 6.98-7.87 to 6.46-7.18. The analysis of microbial community structure of biofilm by pyrosequencing method showed that Proteobacteria was the most abundant phylum (89.87 %), and β-Proteobacteria represented the most abundant class. Among the 76 identified genera, Dechloromonas (10.26 %), Alicycliphilus (9.15 %), Azospira (8.92 %), and Sinobacteraceae-uncultured (8.75 %) were the abundant genera. PBS, as a promising alternative carbon source, is a suitable solid carbon source and biofilm carrier for nitrate removal.

  20. Competitive inhibition of glutamate dehydrogenase reaction.

    Science.gov (United States)

    Choudhury, Rajarshi; Punekar, Narayan S

    2007-06-12

    Irrespective of their pyridine nucleotide specificity, all glutamate dehydrogenases share a common chemical mechanism that involves an enzyme bound 'iminoglutarate' intermediate. Three compounds, structurally related to this intermediate, were tested for the inhibition of purified NADP-glutamate dehydrogenases from two Aspergilli, as also the bovine liver NAD(P)-glutamate dehydrogenase. 2-Methyleneglutarate, closely resembling iminoglutarate, was a potent competitive inhibitor of the glutamate dehydrogenase reaction. This is the first report of a non-aromatic structure with a better glutamate dehydrogenase inhibitory potency than aryl carboxylic acids such as isophthalate. A suitably located 2-methylene group to mimic the iminium ion could be exploited to design inhibitors of other amino acid dehydrogenases.

  1. ald of Mycobacterium tuberculosis Encodes both the Alanine Dehydrogenase and the Putative Glycine Dehydrogenase

    Science.gov (United States)

    Giffin, Michelle M.; Modesti, Lucia; Raab, Ronald W.; Wayne, Lawrence G.

    2012-01-01

    The putative glycine dehydrogenase of Mycobacterium tuberculosis catalyzes the reductive amination of glyoxylate to glycine but not the reverse reaction. The enzyme was purified and identified as the previously characterized alanine dehydrogenase. The Ald enzyme was expressed in Escherichia coli and had both pyruvate and glyoxylate aminating activities. The gene, ald, was inactivated in M. tuberculosis, which resulted in the loss of all activities. Both enzyme activities were found associated with the cell and were not detected in the extracellular filtrate. By using an anti-Ald antibody, the protein was localized to the cell membrane, with a smaller fraction in the cytosol. None was detected in the extracellular medium. The ald knockout strain grew without alanine or glycine and was able to utilize glycine but not alanine as a nitrogen source. Transcription of ald was induced when alanine was the sole nitrogen source, and higher levels of Ald enzyme were measured. Ald is proposed to have several functions, including ammonium incorporation and alanine breakdown. PMID:22210765

  2. 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...

  3. 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.

  4. 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.

  5. 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.

  6. Effect of alpha-tocopherol and tocopherol succinate on lipid peroxidation in equine spermatozoa.

    Science.gov (United States)

    Almeida, Juliana; Ball, Barry A

    2005-07-01

    The objective of this study was to compare the effect of alpha-tocopherol and its ester, alpha tocopherol succinate, on lipid peroxidation and motility of equine spermatozoa. In experiment one, spermatozoa were incubated with dl-alpha-tocopherol (5, 25, 100 or 500 microM), DL-alpha tocopherol succinate (5, 25, 100 or 500 microM) or vehicle (0.5% ethanol) at 38 degrees C, and sperm motility was determined at 30, 60 and 120 min. In experiment two, spermatozoa loaded with the lipophilic probe, C11BODIPY(581/591), were incubated with dl-alpha-tocopherol (50 and 100 microM), DL-alpha-tocopherol succinate (50 and 100 microM) or ethanol (0.5%) and with the promoters cumene hydroperoxide, Fe2SO4, and ascorbate at 38 degrees C in 5% CO2. Lipid peroxidation was determined by changes in fluorescence of C11BODIPY(581/591), and motility was determined by CASA at 0, 15, 30 and 60 min. In experiment three, spermatozoa loaded with C11BODIPY(581/591) were incubated with dl-alpha-tocopherol (5, 25, 100 or 500 microM), DL-alpha-tocopherol succinate (5, 25, 100 or 500 microM) or ethanol (0.5%) at 38 degrees C and then submitted to a 4-hour incubation at room temperature. Motility and lipid peroxidation were determined at 1 and 4 h. In experiment four, the effect of DL alpha tocopherol (5, 25 or 500 microM), DL-alpha-tocopherol succinate (5, 25 or 500 microM) or ethanol (0.5%) on lipid peroxidation and motility were evaluated during storage at 5 degrees C in a skim-milk based extender. Although dl-alpha-tocopherol succinate appeared more effective than DL-alpha-tocopherol in preventing lipid peroxidation during short-term incubations, the succinate ester suppressed sperm motility compared to dl-alpha-tocopherol alone.

  7. Compatibility of ondansetron hydrochloride and methylprednisolone sodium succinate in multilayer polyolefin containers.

    Science.gov (United States)

    Bougouin, Christelle; Thelcide, Chloë; Crespin-Maillard, Fabienne; Maillard, Christian; Kinowski, Jean Marie; Favier, Mireille

    2005-10-01

    The compatibility of ondansetron hydrochloride and methylprednisolone sodium succinate in 5% dextrose injection and 0.9% sodium chloride injection was studied. Test solutions of ondansetron hydrochloride 0.16 mg/mL and methylprednisolone sodium succinate 2.4 mg/mL were prepared in triplicate and tested in duplicate. Total volumes of 4 and 2 mL of ondansetron hydrochloride solution and methylprednisolone sodium succinate solution, respectively, were added to 50-mL multilayer polyolefin bags containing 5% dextrose injection or 0.9% sodium chloride injection. Bags were stored for 24 hours at 20-25 degrees C and for 48 hours at 4-8 degrees C. Chemical compatibility was measured with high-performance liquid chromatography, and physical compatibility was determined visually. Ondansetron hydrochloride was stable for up to 24 hours at 20-25 degrees C and up to 48 hours at 4-8 degrees C. Methylprednisolone sodium succinate was stable for up to 48 hours at 4-8 degrees C. When stored at 20-25 degrees C, methylprednisolone sodium succinate was stable for up to 7 hours in 5% dextrose injection and up to 24 hours in 0.9% sodium chloride injection. Compatibility data for solutions containing ondansetron hydrochloride plus methylprednisolone sodium succinate revealed that each drug was stable for up to 24 hours at 20-25 degrees C and up to 48 hours at 4-8 degrees C. Ondansetron 0.16 mg/mL (as the hydrochloride) and methylprednisolone 2.4 mg/mL (as the sodium succinate) mixed in 50-mL multilayer polyolefin bags were stable in both 5% dextrose injection and 0.9% sodium chloride injection for up to 24 hours at 20-25 degrees C and up to 48 hours at 4-8 degrees C.

  8. Preparation of biodegradable porous poly(butylene succinate microspheres

    Directory of Open Access Journals (Sweden)

    Pepić Dragana

    2008-01-01

    Full Text Available The aim of this study was to determine the optimal conditions for the fabrication of porous microspheres based on poly(butylene succinate, PBS. The biodegradable non-porous PBS microspheres were prepared by the oil-in-water (o/w emulsion solvent evaporation method using poly(vinyl alcohol, PVA, as the surfactant. Fabrication conditions, such as stirring rate, organic/aqueous ratio, PBS concentration and surfactant (PVA concentration, which have an important influence on both the particle size and the morphology of the microspheres, were varied. Scanning electron microscopy, SEM, observations confirmed the size, size distribution and surface morphology of the microspheres. The optimal conditions for the preparation of the non-porous microspheres were found to be: concentration the PBS solution, 10 mass%; PVA concentration, 1 mass%; the organic/ aqueous ratio CHCl3/H2O = 1/20 and stirring rate 800 rpm. Porous PBS microspheres were fabricated under the optimal conditions using various amounts of hexane and poly(ethylene oxide, PEO, as porogens. The influence of the amount of porogen on the pore size and the particle size was investigated using SEM and the apparent density. The microspheres exhibited various porosities and the pore sizes. The average particle size of the microspheres with PEO as the porogen was from 100 to 122μm and that of the microspheres with hexane as the porogen was from 87 to 97μm. The apparent density of the porous microspheres with PEO as the porogen, from 0.16 to 0.23 g/cm3, was much smaller than the non-porous microspheres, 0.40 g/cm3. In the in vitro degradation experiments, the porous microspheres were incubated in phosphate buffer solution (pH 7 at 37°C. After incubating for one month, the microspheres showed significant extent of the hydrolytic degradation of the porous PBS microspheres.

  9. Promotion of growth by Coenzyme Q10 is linked to gene expression in C. elegans.

    Science.gov (United States)

    Fischer, Alexandra; Niklowitz, Petra; Menke, Thomas; Döring, Frank

    2014-10-03

    Coenzyme Q (CoQ, ubiquinone) is an essential component of the respiratory chain, a cofactor of pyrimidine biosynthesis and acts as an antioxidant in extra mitochondrial membranes. More recently CoQ has been identified as a modulator of apoptosis, inflammation and gene expression. CoQ deficient Caenorhabditis elegans clk-1 mutants show several phenotypes including a delayed postembryonic growth. Using wild type and two clk-1 mutants, here we established an experimental set-up to study the consequences of endogenous CoQ deficiency or exogenous CoQ supply on gene expression and growth. We found that a deficiency of endogenous CoQ synthesis down-regulates a cluster of genes that are important for growth (i.e., RNA polymerase II, eukaryotic initiation factor) and up-regulates oxidation reactions (i.e., cytochrome P450, superoxide dismutase) and protein interactions (i.e., F-Box proteins). Exogenous CoQ supply partially restores the expression of these genes as well as the growth retardation of CoQ deficient clk-1 mutants. On the other hand exogenous CoQ supply does not alter the expression of a further sub-set of genes. These genes are involved in metabolism (i.e., succinate dehydrogenase complex), cell signalling or synthesis of lectins. Thus, our work provides a comprehensive overview of genes which can be modulated in their expression by endogenous or exogenous CoQ. As growth retardation in CoQ deficiency is linked to the gene expression profile we suggest that CoQ promotes growth via gene expression. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Characterization of human short chain dehydrogenase/reductase SDR16C family members related to retinol dehydrogenase 10.

    Science.gov (United States)

    Adams, Mark K; Lee, Seung-Ah; Belyaeva, Olga V; Wu, Lizhi; Kedishvili, Natalia Y

    2017-10-01

    All-trans-retinoic acid (RA) is a bioactive derivative of vitamin A that serves as an activating ligand for nuclear transcription factors, retinoic acid receptors. RA biosynthesis is initiated by the enzymes that oxidize retinol to retinaldehyde. It is well established that retinol dehydrogenase 10 (RDH10, SDR16C4), which belongs to the 16C family of the short chain dehydrogenase/reductase (SDR) superfamily of proteins, is the major enzyme responsible for the oxidation of retinol to retinaldehyde for RA biosynthesis during embryogenesis. However, several lines of evidence point towards the existence of additional retinol dehydrogenases that contribute to RA biosynthesis in vivo. In close proximity to RDH10 gene on human chromosome 8 are located two genes that are phylogenetically related to RDH10. The predicted protein products of these genes, retinol dehydrogenase epidermal 2 (RDHE2, SDR16C5) and retinol dehydrogenase epidermal 2-similar (RDHE2S, SDR16C6), share 59% and 56% sequence similarity with RDH10, respectively. Previously, we showed that the single ortholog of the human RDHE2 and RDHE2S in frogs, Xenopus laevis rdhe2, oxidizes retinol to retinaldehyde and is essential for frog embryonic development. In this study, we explored the potential of each of the two human proteins to contribute to RA biosynthesis. The results of this study demonstrate that human RDHE2 exhibits a relatively low but reproducible activity when expressed in either HepG2 or HEK293 cells. Expression of the native RDHE2 is downregulated in the presence of elevated levels of RA. On the other hand, the protein encoded by the human RDHE2S gene is unstable when expressed in HEK293 cells. RDHE2S protein produced in Sf9 cells is stable but has no detectable catalytic activity towards retinol. We conclude that the human RDHE2S does not contribute to RA biosynthesis, whereas the low-activity RA-sensitive human RDHE2 may have a role in adjusting the cellular levels of RA in accord with

  11. Lateral gene transfer and gene duplication played a key role in the evolution of Mastigamoeba balamuthi hydrogenosomes

    Czech Academy of Sciences Publication Activity Database

    Nývltová, E.; Stairs, C.W.; Hrdý, I.; Rídl, Jakub; Mach, J.; Pačes, Jan; Roger, A. J.; Tachezy, J.

    2015-01-01

    Roč. 32, č. 4 (2015), s. 1039-1055 ISSN 0737-4038 R&D Projects: GA ČR(CZ) GAP305/11/1061; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:68378050 Keywords : acetylCoA synthetase * sulfate activation pathway * PFO * glycine cleavage system * hydrogenase * succinate dehydrogenase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 13.649, year: 2015

  12. Krebs cycle metabolites and preferential succinate oxidation following neonatal hypoxic-ischemic brain injury in mice.

    Science.gov (United States)

    Sahni, Prateek V; Zhang, Jimmy; Sosunov, Sergey; Galkin, Alexander; Niatsetskaya, Zoya; Starkov, Anatoly; Brookes, Paul S; Ten, Vadim S

    2018-02-01

    BackgroundReverse electron transport (RET) driven by the oxidation of succinate has been proposed as the mechanism of accelerated production of reactive oxygen species (ROS) in post-ischemic mitochondria. However, it remains unclear whether upon reperfusion, mitochondria preferentially oxidase succinate.MethodsNeonatal mice were subjected to Rice-Vannucci model of hypoxic-ischemic brain injury (HI) followed by assessment of Krebs cycle metabolites, mitochondrial substrate preference, and H 2 O 2 generation rate in the ischemic brain.ResultsWhile brain mitochondria from control mice exhibited a rotenone-sensitive complex-I-dependent respiration, HI-brain mitochondria, at the initiation of reperfusion, demonstrated complex-II-dependent respiration, as rotenone minimally affected, but inhibition of complex-II ceased respiration. This was associated with a 30-fold increase of cerebral succinate concentration and significantly elevated H 2 O 2 emission rate in HI-mice compared to controls. At 60 min of reperfusion, cerebral succinate content and the mitochondrial response to rotenone did not differ from that in controls.ConclusionThese data are the first ex vivo evidence, that at the initiation of reperfusion, brain mitochondria transiently shift their metabolism from complex-I-dependent oxidation of NADH toward complex II-linked oxidation of succinate. Our study provides a critical piece of support for existence of the RET-dependent mechanism of elevated ROS production in reperfusion.

  13. Bio-oil based biorefinery strategy for the production of succinic acid

    DEFF Research Database (Denmark)

    Wang, Caixia; Thygesen, Anders; Liu, Yilan

    2013-01-01

    Background: Succinic acid is one of the key platform chemicals which can be produced via biotechnology process instead of petrochemical process. Biomass derived bio-oil have been investigated intensively as an alternative of diesel and gasoline fuels. Bio-oil could be fractionized into organic...... was investigated. Results: The transgenic E. coli strain could grow in modified M9 medium containing 20 v/v% AP-bio-oil with an increase in OD from 0.25 to 1.09. And 0.38 g/L succinic acid was produced. With the presence of 4 g/L glucose in the medium, succinic acid concentration increased from 1.4 to 2.4 g....../L by addition of 20 v/v% AP-bio-oil. When enzymatic hydrolysate of corn stover was used as carbon source, 10.3 g/L succinic acid was produced. The obtained succinic acid concentration increased to 11.5 g/L when 12.5 v/v% AP-bio-oil was added. However, it decreased to 8 g/L when 50 v/v% AP-bio-oil was added. GC...

  14. Succinate and Lactate Production from Euglena gracilis during Dark, Anaerobic Conditions

    Science.gov (United States)

    Tomita, Yuko; Yoshioka, Kazumasa; Iijima, Hiroko; Nakashima, Ayaka; Iwata, Osamu; Suzuki, Kengo; Hasunuma, Tomohisa; Kondo, Akihiko; Hirai, Masami Yokota; Osanai, Takashi

    2016-01-01

    Euglena gracilis is a eukaryotic, unicellular phytoflagellate that has been widely studied in basic science and applied science. Under dark, anaerobic conditions, the cells of E. gracilis produce a wax ester that can be converted into biofuel. Here, we demonstrate that under dark, anaerobic conditions, E. gracilis excretes organic acids, such as succinate and lactate, which are bulk chemicals used in the production of bioplastics. The levels of succinate were altered by changes in the medium and temperature during dark, anaerobic incubation. Succinate production was enhanced when cells were incubated in CM medium in the presence of NaHCO3. Excretion of lactate was minimal in the absence of external carbon sources, but lactate was produced in the presence of glucose during dark, anaerobic incubation. E. gracilis predominantly produced L-lactate; however, the percentage of D-lactate increased to 28.4% in CM medium at 30°C. Finally, we used a commercial strain of E. gracilis for succinate production and found that nitrogen-starved cells, incubated under dark, anaerobic conditions, produced 869.6 mg/L succinate over a 3-day incubation period, which was 70-fold higher than the amount produced by nitrogen-replete cells. This is the first study to demonstrate organic acid excretion by E. gracilis cells and to reveal novel aspects of primary carbon metabolism in this organism. PMID:28066371

  15. Efficient utilization of cassava pulp for succinate production by metabolically engineered Escherichia coli KJ122.

    Science.gov (United States)

    Sawisit, Apichai; Jantama, Sirima Suvarnakuta; Kanchanatawee, Sunthorn; Jantama, Kaemwich

    2015-01-01

    A metabolically engineered Escherichia coli KJ122 was efficiently utilized for succinate production from cassava pulp during batch separate hydrolysis and fermentation (SHF) under simple anaerobic conditions. Succinate concentration of 41.46 ± 0.05 g/L with yield and productivity of 82.33 ± 0.14 g/100 g dry pulp and 0.84 ± 0.02 g/L/h was obtained. In batch simultaneous saccharification and fermentation (SSF), hydrolysis of 12 % (w/v) cassava pulp with an enzyme loading of 2 % AMG + 3 % Cel (v/w) at pH 6.5 was optimized at 39 °C. Succinate concentration of 80.86 ± 0.49 g/L with a yield of 70.34 ± 0.37 g/100 g dry pulp and a productivity of 0.84 ± 0.01 g/L/h was attained using E. coli KJ122. Fed-batch SSF significantly enhanced succinate concentration to 98.63 ± 0.12 g/L at yield and productivity of 71.64 ± 0.97 g/100 g dry pulp and 1.03 ± 0.01 g/L/h. This result indicated an efficient and economical succinate production from cassava pulp using SHF and SSF by the use of E. coli KJ122.

  16. Genotoxicity of meso-2,3-dimercapto succinic acid-coated silver sulfide quantum dot

    Directory of Open Access Journals (Sweden)

    Deniz Özkan Vardar

    2015-06-01

    Full Text Available Nanotecnology products have been used in wide applications in chemistry, electronics, energy generation, and medicine. Despite significant interest in developing quantum dots (QDs for biomedical applications, many researchers are convinced that QDs will never be used for the treatment of patients because of their potential toxicity. In various in vitro cell culture studies, the cytotoxic properties of some QD have been demonstrated and they have been suggested to be toxic in humans. In this study, the cytotoxic properties of Ag2S-(Meso-2,3-Dimercapto Succinic acid nanomaterials in V79 cells (Chinese lung fibroblast cell line were determined by MTT assay. The genotoxic effects of Ag2S-(Meso-2,3-Dimercapto Succinic acid were evaluated by the alkaline single cell gel electrophoresis. The cells were treated with Ag2S-(Meso-2,3-Dimercapto Succinic acid at the concentrations of 5- 2000 µg/ml. No cytotoxic effect of Ag2S-(Meso-2,3-Dimercapto Succinic acid at all concentrations studied was observed. No significant increases in DNA damage were found at the studied concentrations when compared to negative control in V79 cells. In conclusion, further in vitro and in vivo studies are required to determine the safety doses of Ag2S-(Meso-2,3-Dimercapto Succinic acid.

  17. GABA metabolism pathway genes, UGA1 and GAD1, regulate replicative lifespan in Saccharomycescerevisiae

    International Nuclear Information System (INIS)

    Kamei, Yuka; Tamura, Takayuki; Yoshida, Ryo; Ohta, Shinji; Fukusaki, Eiichiro; Mukai, Yukio

    2011-01-01

    Highlights: →We demonstrate that two genes in the yeast GABA metabolism pathway affect aging. → Deletion of the UGA1 or GAD1 genes extends replicative lifespan. → Addition of GABA to wild-type cultures has no effect on lifespan. → Intracellular GABA levels do not differ in longevity mutants and wild-type cells. → Levels of tricarboxylic acid cycle intermediates positively correlate with lifespan. -- Abstract: Many of the genes involved in aging have been identified in organisms ranging from yeast to human. Our previous study showed that deletion of the UGA3 gene-which encodes a zinc-finger transcription factor necessary for γ-aminobutyric acid (GABA)-dependent induction of the UGA1 (GABA aminotransferase), UGA2 (succinate semialdehyde dehydrogenase), and UGA4 (GABA permease) genes-extends replicative lifespan in the budding yeast Saccharomycescerevisiae. Here, we found that deletion of UGA1 lengthened the lifespan, as did deletion of UGA3; in contrast, strains with UGA2 or UGA4 deletions exhibited no lifespan extension. The Δuga1 strain cannot deaminate GABA to succinate semialdehyde. Deletion of GAD1, which encodes the glutamate decarboxylase that converts glutamate into GABA, also increased lifespan. Therefore, two genes in the GABA metabolism pathway, UGA1 and GAD1, were identified as aging genes. Unexpectedly, intracellular GABA levels in mutant cells (except for Δuga2 cells) did not differ from those in wild-type cells. Addition of GABA to culture media, which induces transcription of the UGA structural genes, had no effect on replicative lifespan of wild-type cells. Multivariate analysis of 1 H nuclear magnetic resonance spectra for the whole-cell metabolite levels demonstrated a separation between long-lived and normal-lived strains. Gas chromatography-mass spectrometry analysis of identified metabolites showed that levels of tricarboxylic acid cycle intermediates positively correlated with lifespan extension. These results strongly suggest

  18. GABA metabolism pathway genes, UGA1 and GAD1, regulate replicative lifespan in Saccharomycescerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Kamei, Yuka; Tamura, Takayuki [Department of Bioscience, Faculty of Bioscience, Nagahama Institute of Bio-Science and Technology, 1266 Tamura, Nagahama, Shiga 526-0829 (Japan); Yoshida, Ryo [Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871 (Japan); Ohta, Shinji [Department of Bioscience, Faculty of Bioscience, Nagahama Institute of Bio-Science and Technology, 1266 Tamura, Nagahama, Shiga 526-0829 (Japan); Fukusaki, Eiichiro [Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871 (Japan); Mukai, Yukio, E-mail: y_mukai@nagahama-i-bio.ac.jp [Department of Bioscience, Faculty of Bioscience, Nagahama Institute of Bio-Science and Technology, 1266 Tamura, Nagahama, Shiga 526-0829 (Japan)

    2011-04-01

    Highlights: {yields}We demonstrate that two genes in the yeast GABA metabolism pathway affect aging. {yields} Deletion of the UGA1 or GAD1 genes extends replicative lifespan. {yields} Addition of GABA to wild-type cultures has no effect on lifespan. {yields} Intracellular GABA levels do not differ in longevity mutants and wild-type cells. {yields} Levels of tricarboxylic acid cycle intermediates positively correlate with lifespan. -- Abstract: Many of the genes involved in aging have been identified in organisms ranging from yeast to human. Our previous study showed that deletion of the UGA3 gene-which encodes a zinc-finger transcription factor necessary for {gamma}-aminobutyric acid (GABA)-dependent induction of the UGA1 (GABA aminotransferase), UGA2 (succinate semialdehyde dehydrogenase), and UGA4 (GABA permease) genes-extends replicative lifespan in the budding yeast Saccharomycescerevisiae. Here, we found that deletion of UGA1 lengthened the lifespan, as did deletion of UGA3; in contrast, strains with UGA2 or UGA4 deletions exhibited no lifespan extension. The {Delta}uga1 strain cannot deaminate GABA to succinate semialdehyde. Deletion of GAD1, which encodes the glutamate decarboxylase that converts glutamate into GABA, also increased lifespan. Therefore, two genes in the GABA metabolism pathway, UGA1 and GAD1, were identified as aging genes. Unexpectedly, intracellular GABA levels in mutant cells (except for {Delta}uga2 cells) did not differ from those in wild-type cells. Addition of GABA to culture media, which induces transcription of the UGA structural genes, had no effect on replicative lifespan of wild-type cells. Multivariate analysis of {sup 1}H nuclear magnetic resonance spectra for the whole-cell metabolite levels demonstrated a separation between long-lived and normal-lived strains. Gas chromatography-mass spectrometry analysis of identified metabolites showed that levels of tricarboxylic acid cycle intermediates positively correlated with lifespan

  19. atp mutants of Escherichia coli fail to grow on succinate due to a transport deficiency

    DEFF Research Database (Denmark)

    Boogerd, Fred; Boe, Lars; Michelsen, Ole

    1998-01-01

    Escherichia coli atp mutants, which lack a functional Hf-ATPase complex, are capable of growth on glucose but not on succinate or other C-4-dicarboxylates (Suc(-) phenotype). Suc(+) revertants of an atp deletion strain were isolated which were capable of growth on succinate even though they lack......) was expressed in trans, the Suc(-) phenotype of the atp deletion strain reverted to Suc(+), which shows that the reason why the E. coli atp mutant is unable to grow aerobically on C-4-dicarboxylates is insufficient transport capacity for these substrates....

  20. Receptor structure-based discovery of non-metabolite agonists for the succinate receptor GPR91

    DEFF Research Database (Denmark)

    Trauelsen, Mette; Rexen Ulven, Elisabeth; Hjorth, Siv A

    2017-01-01

    backbone is preferred and that certain backbone modifications are allowed for GPR91 activation. Through receptor modeling over the X-ray structure of the closely related P2Y1 receptor, we discovered that the binding pocket is partly occupied by a segment of an extracellular loop and that succinate...... therefore binds in a very different mode than generally believed. Importantly, an empty side-pocket is identified next to the succinate binding site. All this information formed the basis for a substructure-based search query, which, combined with molecular docking, was used in virtual screening of the ZINC...

  1. Synthesis of tocopheryl succinate phospholipid conjugates and monitoring of phospholipase A2 activity

    DEFF Research Database (Denmark)

    Pedersen, Palle Jacob; Viart, Helene Marie-France; Melander, Fredrik

    2012-01-01

    Tocopheryl succinates (TOSs) are, in contrast to tocopherols, highly cytotoxic against many cancer cells. In this study the enzyme activity of secretory phospholipase A2 towards various succinate-phospholipid conjugates has been investigated. The synthesis of six novel phospholipids is described,...

  2. Modification of cellulose with succinic anhydride in TBAA/DMSO mixed solvent under catalyst-free conditions

    Science.gov (United States)

    Homogeneous modification of cellulose with succinic anhydride was performed in tetrabutylammonium acetate (TBAA)/dimethyl sulfoxide (DMSO) mixed solvent. The molar ratio of succinic anhydride (SA) to free hydroxyl groups in the anhydroglucose units (AGU) and TBAA dosage were investigated as paramete...

  3. Modification of cellulose with succinic anhydride in TBAA/DMSO mixed solvent under catalyst-free conditions

    Science.gov (United States)

    Ping-Ping Xin; Yao-Bing Huang; Chung-Yun Hse; Huai N. Cheng; Chaobo Huang; Hui. Pan

    2017-01-01

    Homogeneous modification of cellulose with succinic anhydride was performed using tetrabutylammonium acetate (TBAA)/dimethyl sulfoxide (DMSO) mixed solvent. The molar ratio of succinic anhydride (SA) to free hydroxyl groups in the anhydroglucose units (AGU), TBAA dosage, reaction temperature, and reaction time were investigated. The highest degree of substitution (DS)...

  4. 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.

  5. Expression of Aeromonas caviae ST pyruvate dehydrogenase complex components mediate tellurite resistance in Escherichia coli

    International Nuclear Information System (INIS)

    Castro, Miguel E.; Molina, Roberto C.; Diaz, Waldo A.; Pradenas, Gonzalo A.; Vasquez, Claudio C.

    2009-01-01

    Potassium tellurite (K 2 TeO 3 ) is harmful to most organisms and specific mechanisms explaining its toxicity are not well known to date. We previously reported that the lpdA gene product of the tellurite-resistant environmental isolate Aeromonas caviae ST is involved in the reduction of tellurite to elemental tellurium. In this work, we show that expression of A. caviae ST aceE, aceF, and lpdA genes, encoding pyruvate dehydrogenase, dihydrolipoamide transacetylase, and dihydrolipoamide dehydrogenase, respectively, results in tellurite resistance and decreased levels of tellurite-induced superoxide in Escherichia coli. In addition to oxidative damage resulting from tellurite exposure, a metabolic disorder would be simultaneously established in which the pyruvate dehydrogenase complex would represent an intracellular tellurite target. These results allow us to widen our vision regarding the molecular mechanisms involved in bacterial tellurite resistance by correlating tellurite toxicity and key enzymes of aerobic metabolism.

  6. Human dehydrogenase/reductase (SDR family) member 11 is a novel type of 17β-hydroxysteroid dehydrogenase.

    Science.gov (United States)

    Endo, Satoshi; Miyagi, Namiki; Matsunaga, Toshiyuki; Hara, Akira; Ikari, Akira

    2016-03-25

    We report characterization of a member of the short-chain dehydrogenase/reductase superfamily encoded in a human gene, DHRS11. The recombinant protein (DHRS11) efficiently catalyzed the conversion of the 17-keto group of estrone, 4- and 5-androstenes and 5α-androstanes into their 17β-hydroxyl metabolites with NADPH as a coenzyme. In contrast, it exhibited reductive 3β-hydroxysteroid dehydrogenase activity toward 5β-androstanes, 5β-pregnanes, 4-pregnenes and bile acids. Additionally, DHRS11 reduced α-dicarbonyls (such as diacetyl and methylglyoxal) and alicyclic ketones (such as 1-indanone and loxoprofen). The enzyme activity was inhibited in a mixed-type manner by flavonoids, and competitively by carbenoxolone, glycyrrhetinic acid, zearalenone, curcumin and flufenamic acid. The expression of DHRS11 mRNA was observed widely in human tissues, most abundantly in testis, small intestine, colon, kidney and cancer cell lines. Thus, DHRS11 represents a novel type of 17β-hydroxysteroid dehydrogenase with unique catalytic properties and tissue distribution. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Gene expression analyses reveal metabolic specifications in acute O2-sensing chemoreceptor cells.

    Science.gov (United States)

    Gao, Lin; Bonilla-Henao, Victoria; García-Flores, Paula; Arias-Mayenco, Ignacio; Ortega-Sáenz, Patricia; López-Barneo, José

    2017-09-15

    Glomus cells in the carotid body (CB) and chromaffin cells in the adrenal medulla (AM) are essential for reflex cardiorespiratory adaptation to hypoxia. However, the mechanisms whereby these cells detect changes in O 2 tension are poorly understood. The metabolic properties of acute O 2 -sensing cells have been investigated by comparing the transcriptomes of CB and AM cells, which are O 2 -sensitive, with superior cervical ganglion neurons, which are practically O 2 -insensitive. In O 2 -sensitive cells, we found a characteristic prolyl hydroxylase 3 down-regulation and hypoxia inducible factor 2α up-regulation, as well as overexpression of genes coding for three atypical mitochondrial electron transport subunits and pyruvate carboxylase, an enzyme that replenishes tricarboxylic acid cycle intermediates. In agreement with this observation, the inhibition of succinate dehydrogenase impairs CB acute O 2 sensing. The responsiveness of peripheral chemoreceptor cells to acute hypoxia depends on a 'signature metabolic profile'. Acute O 2 sensing is a fundamental property of cells in the peripheral chemoreceptors, e.g. glomus cells in the carotid body (CB) and chromaffin cells in the adrenal medulla (AM), and is necessary for adaptation to hypoxia. These cells contain O 2 -sensitive ion channels, which mediate membrane depolarization and transmitter release upon exposure to hypoxia. However, the mechanisms underlying the detection of changes in O 2 tension by cells are still poorly understood. Recently, we suggested that CB glomus cells have specific metabolic features that favour the accumulation of reduced quinone and the production of mitochondrial NADH and reactive oxygen species during hypoxia. These signals alter membrane ion channel activity. To investigate the metabolic profile characteristic of acute O 2 -sensing cells, we used adult mice to compare the transcriptomes of three cell types derived from common sympathoadrenal progenitors, but exhibiting variable

  8. 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...

  9. Investigations regarding the anthropic impact on the Krebs cycle dehydrogenases system on certain wood-species in mining areas, Suceava county

    Directory of Open Access Journals (Sweden)

    Marius Viorel Oniciuc

    2013-03-01

    Full Text Available The Krebs cycle, a second stage of cellular respiration that occurs in the mitochondrion of the leafcell and consist in a multistep processes plays a central role in catabolism of organic fuel molecules. The miningextraction technologies for both underground and surface, the preparation of copper ore and barite applied in Tarnia,respectively to the sulphur in Calimani Mountain and the excess of these elements in natural environment may causemalfunction of ecosystem. The dehydrogenases of Krebs cycle can give information on the type and the duration of theeffects of pollutants on the metabolic activity in leaves, to subsequent area pollution, therefore, the aim of the presentstudy has been to determine these effects on this enzymatic system activity. For this reason, the isocitrate dehydrogenase,the -ketoglutate dehydrogenase, the succinate ehydrogenase and the malate dehydrogenase activity was determined using the spectrophotometric method with triphenyl-tetrazolium and the analysis of experimental results shows the differences from one sample to another sample of closely related species specificity, but also the effect of environmentalfactors.

  10. 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.

  11. Genetics Home Reference: dihydrolipoamide dehydrogenase deficiency

    Science.gov (United States)

    ... Lacaille F, de Keyzer Y, Di Martino V, de Lonlay P. Dihydrolipoamide dehydrogenase deficiency: a still overlooked cause of recurrent acute liver failure and Reye-like syndrome. Mol Genet Metab. 2013 May;109(1):28- ...

  12. Histochemical localization of cytokinin oxidase/dehydrogenase ...

    African Journals Online (AJOL)

    Jane

    2011-08-15

    dehydrogenase, Withania somnifera, CKX localization. INTRODUCTION. Cytokinin (Ck) is a plant hormone that plays a crucial role in many fundamental processes of plant development throughout the life cycle. These include ...

  13. 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.

  14. Isocitrate dehydrogenase mutations in gliomas

    Science.gov (United States)

    Waitkus, Matthew S.; Diplas, Bill H.; Yan, Hai

    2016-01-01

    Over the last decade, extraordinary progress has been made in elucidating the underlying genetic causes of gliomas. In 2008, our understanding of glioma genetics was revolutionized when mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) were identified in the vast majority of progressive gliomas and secondary glioblastomas (GBMs). IDH enzymes normally catalyze the decarboxylation of isocitrate to generate α-ketoglutarate (αKG), but recurrent mutations at Arg132 of IDH1 and Arg172 of IDH2 confer a neomorphic enzyme activity that catalyzes reduction of αKG into the putative oncometabolite D-2-hydroxyglutate (D2HG). D2HG inhibits αKG-dependent dioxygenases and is thought to create a cellular state permissive to malignant transformation by altering cellular epigenetics and blocking normal differentiation processes. Herein, we discuss the relevant literature on mechanistic studies of IDH1/2 mutations in gliomas, and we review the potential impact of IDH1/2 mutations on molecular classification and glioma therapy. PMID:26188014

  15. 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...

  16. 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

  17. Succinic Acid as a Byproduct in a Corn-based Ethanol Biorefinery

    Energy Technology Data Exchange (ETDEWEB)

    MBI International

    2007-12-31

    MBI endeavored to develop a process for succinic acid production suitable for integration into a corn-based ethanol biorefinery. The project investigated the fermentative production of succinic acid using byproducts of corn mill operations. The fermentation process was attuned to include raw starch, endosperm, as the sugar source. A clean-not-sterile process was established to treat the endosperm and release the monomeric sugars. We developed the fermentation process to utilize a byproduct of corn ethanol fermentations, thin stillage, as the source of complex nitrogen and vitamin components needed to support succinic acid production in A. succinogenes. Further supplementations were eliminated without lowering titers and yields and a productivity above 0.6 g l-1 hr-1was achieved. Strain development was accomplished through generation of a recombinant strain that increased yields of succinic acid production. Isolation of additional strains with improved features was also pursued and frozen stocks were prepared from enriched, characterized cultures. Two recovery processes were evaluated at pilot scale and data obtained was incorporated into our economic analyses.

  18. Electrospun Nanoporous Poly(butylenes succinate-co-bytylene terephthalate Nonwoven Mats

    Directory of Open Access Journals (Sweden)

    Liang Wang

    2011-01-01

    Full Text Available A traditional Chinese drug “Yunnan Baiyao” is used as an additive in poly(butylenes succinate-co-bytylene terephthalate (PBST solution, which is a kind of biodegradable aliphatic-aromatic copolyesters, to produce microspheres with nanoporosity by electrospinning; the tunable size of nanoporosity can be controlled by changing the voltage applied in the electrospinning process.

  19. Succinic acid functionalized silver nanoparticles (Suc-Ag NPs) for colorimetric sensing of melamine

    Science.gov (United States)

    Rajar, Kausar; Sirajuddin; Balouch, Aamna; Bhanger, M. I.; Shah, Muhammad Tariq; Shaikh, Tayyaba; Siddiqui, Samia

    2018-03-01

    In this study, a quantitative colorimetric sensing strategy is developed for the rapid, sensitive and selective determination of melamine. The sensing system relies on the application of succinic acid as a selective recognition probe functionalized over Ag NPs. The synthesized Ag NPs were modified with cysteamine to induce positively charged atmosphere which allowed easy and favorable functionalization of succinic acid. The di-carboxyl nature of succinic acid enabled its binding to both cysteamine and melamine. The strong and favorable linkage between succinic acids carbonyl and amine moieties of melamine triggered aggregation of silver NPs producing a significant shift in the measured absorption excitation. This change in the excitation along with the colorimetric response was found linearly proportional to the melamine concentration in the range of 0.1-1.2 μM. The developed sensor system is simple and unlike electrostatic attraction based sensor system utilize selective linkage for the recognition of melamine. In addition to this, the developed optical probe can efficiently be used for the determination of melamine in milk samples.

  20. Succinate-based preparation alleviates manifestations of the climacteric syndrome in women.

    Science.gov (United States)

    Peskov, A B; Maevskii, E I; Uchitel', M L; Sakharova, N Yu; Vize-Khripunova, M A

    2005-09-01

    Clinical placebo-controlled study of Enerlit-Clima (bioactive succinate-based food additive) a showed positive effect of the preparation on general clinical and psychoemotional manifestations of the climacteric syndrome. A trend to an increase in estradiol level in early pathological climacteric and normalization of the endometrial status were observed.

  1. Viscoelastic and electrical properties of carbon nanotubes filled poly(butylene succinate)

    CSIR Research Space (South Africa)

    Bandyopadhyay, J

    2014-03-01

    Full Text Available The carbon nanotubes (CNTs)-containing composites of poly(butylene succinate) (PBS) were prepared by melt-blending in a batch mixer with three concentrations by weight of CNTs: 1, 2 and 3 %. State of dispersion-distribution of the CNTs in the PBS...

  2. 21 CFR 172.765 - Succistearin (stearoyl propylene glycol hydrogen succinate).

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Succistearin (stearoyl propylene glycol hydrogen... propylene glycol hydrogen succinate). The food additive succistearin (stearoyl propylene glycol hydrogen... or C18 fatty acid chain length), and propylene glycol. (b) The additive meets the following...

  3. Tocopherol-associated protein-1 accelerates apoptosis induced by alpha-tocopheryl succinate in mesothelioma cells

    Czech Academy of Sciences Publication Activity Database

    Neužil, Jiří; Dong, L.F.; Wang, X.F.; Zingg, J.M.

    2006-01-01

    Roč. 343, č. 4 (2006), s. 1113-1117 ISSN 0006-291X Institutional research plan: CEZ:AV0Z50520514 Keywords : apoptosis * tocopherol-associated protein * alpha-tocopheryl succinate Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.855, year: 2006

  4. Alpha-tocopheryl succinate inhibits malignant mesothelioma by disrupting the fibroblast growth factor autocrine loop

    Czech Academy of Sciences Publication Activity Database

    Stapelberg, M.; Gellert, N.; Swettenham, E.; Tomasetti, M.; Witting, P. K.; Procopio, A.; Neužil, Jiří

    2005-01-01

    Roč. 280, č. 27 (2005), s. 25369-25376 ISSN 0021-9258 Institutional research plan: CEZ:AV0Z50520514 Keywords : alpha-tocopheryl succinate * malignant mesothelioma * fibroblast growth factor Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.854, year: 2005

  5. Fourier Transform Infrared Spectroscopy Study on Cation adsorption on Viscose Rayon Succinate

    Directory of Open Access Journals (Sweden)

    D Khasbaatar

    2014-09-01

    Full Text Available Ion-exchange materials have been considered as suitable material for the recovery of heavy metals in water. A viscose rayon succinate, synthesized from viscose rayon and succinic anhydride in presence of DMSO, to remove trace bivalent metal ions such as Ag+, Cu2+, Ni2+, Pb2+, Zn2+ and Cr3+, was studied using FT-IR for the behavior of metal adsorption. Both esterification and carboxyl bonding of viscose rayon succinate were assigned essentially at 1729 and 1693cm-1, respectively. And the essential band of bonding between metal and the material was determined at 1625cm-1. The available adsorption capacity of this fiber was 6.2 mequiv/g. The adsorption of metal ions on the viscose rayon succinate follows the order of Cu2+>Cr3+>Ni2+>Pb2+>Zn2+>Ag+ with maximum adsorptions capacities 4.2, 1.42, 0.91, 0.83, 0.69 and 0.35 mmol/g, respectively.DOI: http://dx.doi.org/10.5564/mjc.v12i0.189 Mongolian Journal of Chemistry Vol.12 2011: 136-141

  6. Receptor structure-based discovery of non-metabolite agonists for the succinate receptor GPR91

    Directory of Open Access Journals (Sweden)

    Mette Trauelsen

    2017-12-01

    Conclusions: These novel, synthetic non-metabolite GPR91 agonists will be valuable both as pharmacological tools to delineate the GPR91-mediated functions of succinate and as leads for the development of GPR91-targeted drugs to potentially treat low grade metabolic inflammation and diabetic complications such as retinopathy and nephropathy.

  7. Cell recycled culture of succinic acid-producing Anaerobiospirillum succiniciproducens using an internal membrane filtration system.

    Science.gov (United States)

    Lee, Pyung-Cheon; Lee, Sang-Yup; Chang, Ho-Nam

    2008-07-01

    Cell recycled culture of succinic acid-producing Anaerobiospirillum succiniciproducens was anaerobically carried out using an internal membrane filter module in order to examine the physiological response of A. succiniciproducens to a high-cell-density environment. The optimal growth of A. succiniciproducens and its enhanced succinic acid productivity were observed under CO2-rich conditions, established by adding NaHCO3 and Na2CO3, in the cell recycled system. A. succiniciproducens grew up to 6.50 g-DCW/l, the highest cell concentration obtained so far, in cell recycled cultures. The cells did not change their morphology, which is known to be easily changed in unfavorable or stress environments. The maximum productivity of succinic acid was about 3.3 g/l/h, which is 3.3 times higher than those obtained in batch cultures. These results can serve as a guide for designing highly efficient cell recycled systems for succinic acid at a commercial level.

  8. Efficient production of succinic acid in immobilized fermentation with crude glycerol from Escherichia coli

    Directory of Open Access Journals (Sweden)

    Nik Nor Aziati, A.A.

    2017-10-01

    Full Text Available The increase in the price of commercial succinic acid has necessitated the need for its synthesis from waste materials such as glycerol. Glycerol residue is a waste product of Oleochemical production which is cheaply available and a very good source of carbon. The use of immobilized cells can further reduce the overall cost of the production process. This study primarily aims to produce succinic acid from glycerol residue through the use of immobilized Escherichia coli in a batch fermentation process. The parameters which affect bacterial fermentation process such as the mass substrate, temperature, inoculum size and duration of fermentation were screened using One-Factor-At-a-Time (OFAT method. The result of the screening process shows that a substrate (glycerol concentration of 30 g, inoculum size 20% v/v, and time 4 h produced the maximum succinic acid concentration of 117.99 g/L. The immobilized cells were found to be stable as well as retain their fermentative ability up to the 6th cycle of recycling, thereby presenting as an advantage over the free cell system. Therefore, conclude that using immobilized cells can contribute immensely to the cost-effective production of succinic acid from glycerol residue.

  9. Formation and stability of Vitamin E enriched nanoemulsions stabilized by Octenyl Succinic Anhydride modified starch

    Science.gov (United States)

    Vitamin E (VE) is highly susceptible to autoxidation; therefore, it requires systems to encapsulate and protect it from autoxidation.In this study,we developed VE delivery systems, which were stabilized by Capsul® (MS), a starch modified with octenyl succinic anhydride. Influences of interfacial ten...

  10. Volatility of organic aerosol: evaporation of ammonium sulfate/succinic acid aqueous solution droplets.

    Science.gov (United States)

    Yli-Juuti, Taina; Zardini, Alessandro A; Eriksson, Axel C; Hansen, Anne Maria K; Pagels, Joakim H; Swietlicki, Erik; Svenningsson, Birgitta; Glasius, Marianne; Worsnop, Douglas R; Riipinen, Ilona; Bilde, Merete

    2013-01-01

    Condensation and evaporation modify the properties and effects of atmospheric aerosol particles. We studied the evaporation of aqueous succinic acid and succinic acid/ammonium sulfate droplets to obtain insights on the effect of ammonium sulfate on the gas/particle partitioning of atmospheric organic acids. Droplet evaporation in a laminar flow tube was measured in a Tandem Differential Mobility Analyzer setup. A wide range of droplet compositions was investigated, and for some of the experiments the composition was tracked using an Aerosol Mass Spectrometer. The measured evaporation was compared to model predictions where the ammonium sulfate was assumed not to directly affect succinic acid evaporation. The model captured the evaporation rates for droplets with large organic content but overestimated the droplet size change when the molar concentration of succinic acid was similar to or lower than that of ammonium sulfate, suggesting that ammonium sulfate enhances the partitioning of dicarboxylic acids to aqueous particles more than currently expected from simple mixture thermodynamics. If extrapolated to the real atmosphere, these results imply enhanced partitioning of secondary organic compounds to particulate phase in environments dominated by inorganic aerosol.

  11. Viscoelastic properties of poly(butylene succinate)-co-adipate) nanocomposites

    CSIR Research Space (South Africa)

    Al-Thabaiti, SA

    2015-03-01

    Full Text Available This article reports the viscoelastic properties of poly[(butylene succinate)-co-adipate] (PBSA) nanocomposites. The nanocomposites of PBSA with various loadings of organically modified clay were prepared by melt-mixing in a batch-mixer. The solid...

  12. Integration of Succinic Acid Production in a Dry Mill Ethanol Facility

    Energy Technology Data Exchange (ETDEWEB)

    None

    2006-08-01

    This project seeks to address both issues for a dry mill ethanol biorefinery by lowering the cost of sugars with the development of an advanced pretreatment process, improving the economics of succinic acid (SA), and developing a model of an ethanol dry mill to evaluate the impact of adding different products and processes to a dry mill.

  13. Volatility of Organic Aerosol: Evaporation of Ammonium Sulfate/Succinic Acid Aqueous Solution Droplets

    Science.gov (United States)

    2013-01-01

    Condensation and evaporation modify the properties and effects of atmospheric aerosol particles. We studied the evaporation of aqueous succinic acid and succinic acid/ammonium sulfate droplets to obtain insights on the effect of ammonium sulfate on the gas/particle partitioning of atmospheric organic acids. Droplet evaporation in a laminar flow tube was measured in a Tandem Differential Mobility Analyzer setup. A wide range of droplet compositions was investigated, and for some of the experiments the composition was tracked using an Aerosol Mass Spectrometer. The measured evaporation was compared to model predictions where the ammonium sulfate was assumed not to directly affect succinic acid evaporation. The model captured the evaporation rates for droplets with large organic content but overestimated the droplet size change when the molar concentration of succinic acid was similar to or lower than that of ammonium sulfate, suggesting that ammonium sulfate enhances the partitioning of dicarboxylic acids to aqueous particles more than currently expected from simple mixture thermodynamics. If extrapolated to the real atmosphere, these results imply enhanced partitioning of secondary organic compounds to particulate phase in environments dominated by inorganic aerosol. PMID:24107221

  14. Amorphous/crystal and polymer/filler interphases in biocomposites from poly(butylene succinate)

    Energy Technology Data Exchange (ETDEWEB)

    Signori, Francesca [Consiglio Nazionale delle Ricerche - Istituto per i Processi Chimico-Fisici (CNR-IPCF), Via G. Moruzzi 1, I-56124 Pisa (Italy); Pelagaggi, Martina [Universita di Pisa - Dipartimento di Chimica e Chimica Industriale, Via Risorgimento 35, I-56126 Pisa (Italy); Bronco, Simona [Consiglio Nazionale delle Ricerche - Istituto per i Processi Chimico-Fisici (CNR-IPCF), Via G. Moruzzi 1, I-56124 Pisa (Italy); Righetti, Maria Cristina, E-mail: righetti@ipcf.cnr.it [Consiglio Nazionale delle Ricerche - Istituto per i Processi Chimico-Fisici (CNR-IPCF), Via G. Moruzzi 1, I-56124 Pisa (Italy)

    2012-09-10

    Highlights: Black-Right-Pointing-Pointer The existence of intermolecular interactions between poly(butylene succinate) and hemp fibres was proved from specific heat capacities data. Black-Right-Pointing-Pointer Different degrees of mobility of the poly(butylene succinate) amorphous segments were evidenced at the amorphous/crystal interphase. Black-Right-Pointing-Pointer Devitrification of the rigid amorphous fraction in poly(butylene succinate) was found to occur before and simultaneously with the fusion. - Abstract: Poly(butylene succinate)-hemp composites (PBS-hemp), with hemp content in the range 0-40 wt.%, were prepared in the melt and characterized. This paper focuses on the detailed analysis of the thermal behaviour of the PBS-hemp composites, investigated by differential scanning calorimetry (DSC), to enlighten the polymer/fibre interphase features. The occurrence of specific intermolecular interactions between PBS and hemp was assessed from specific heat capacity data. Different degrees of mobility of the PBS amorphous segments were found at the amorphous/crystal interphases. A broadening of the bulk glass transition was observed, and attributed to the presence of polymer segments slightly constrained. Moreover, a rigid amorphous fraction that devitrifies at temperatures higher than the bulk glass transition, partly before the melting region and partly simultaneously with the fusion, was observed and quantified, and attributed to the presence of major constraints probably occurring in geometrically restricted areas.

  15. Succinic acid production on xylose-enriched biorefinery streams by Actinobacillus succinogenes in batch fermentation.

    Science.gov (United States)

    Salvachúa, Davinia; Mohagheghi, Ali; Smith, Holly; Bradfield, Michael F A; Nicol, Willie; Black, Brenna A; Biddy, Mary J; Dowe, Nancy; Beckham, Gregg T

    2016-01-01

    Co-production of chemicals from lignocellulosic biomass alongside fuels holds promise for improving the economic outlook of integrated biorefineries. In current biochemical conversion processes that use thermochemical pretreatment and enzymatic hydrolysis, fractionation of hemicellulose-derived and cellulose-derived sugar streams is possible using hydrothermal or dilute acid pretreatment (DAP), which then offers a route to parallel trains for fuel and chemical production from xylose- and glucose-enriched streams. Succinic acid (SA) is a co-product of particular interest in biorefineries because it could potentially displace petroleum-derived chemicals and polymer precursors for myriad applications. However, SA production from biomass-derived hydrolysates has not yet been fully explored or developed. Here, we employ Actinobacillus succinogenes 130Z to produce succinate in batch fermentations from various substrates including (1) pure sugars to quantify substrate inhibition, (2) from mock hydrolysates similar to those from DAP containing single putative inhibitors, and (3) using the hydrolysate derived from two pilot-scale pretreatments: first, a mild alkaline wash (deacetylation) followed by DAP, and secondly a single DAP step, both with corn stover. These latter streams are both rich in xylose and contain different levels of inhibitors such as acetate, sugar dehydration products (furfural, 5-hydroxymethylfurfural), and lignin-derived products (ferulate, p-coumarate). In batch fermentations, we quantify succinate and co-product (acetate and formate) titers as well as succinate yields and productivities. We demonstrate yields of 0.74 g succinate/g sugars and 42.8 g/L succinate from deacetylated DAP hydrolysate, achieving maximum productivities of up to 1.27 g/L-h. Moreover, A. succinogenes is shown to detoxify furfural via reduction to furfuryl alcohol, although an initial lag in succinate production is observed when furans are present. Acetate seems to be the

  16. Alcohol consumption and type 2 diabetes: Influence of genetic variation in alcohol dehydrogenase

    NARCIS (Netherlands)

    Beulens, J.W.J.; Rimm, E.B.; Hendriks, H.F.J.; Hu, F.B.; Manson, J.E.; Hunter, D.J.; Mukamal, K.J.

    2007-01-01

    OBJECTIVE - We sought to investigate whether a polymorphism in the alcohol dehydrogenase 1c (ADH1C) gene modifies the association between alcohol consumption and type 2 diabetes. RESEARCH DESIGN AND METHODS - In nested case-control studies of 640 women with incident diabetes and 1,000 control

  17. Glycerol dehydrogenase, encoded by gldB is essential to osmotolerance in Aspergillus nidulans

    NARCIS (Netherlands)

    Vries, de R.P.; Flitter, S.J.; Vondervoort, van de P.J.I.; Chaveroche, M.K.; Fontaine, T.; Fillinger, S.; Ruijter, G.J.G.; Enfert, d' C.; Visser, J.

    2003-01-01

    We have characterized the Aspergillus nidulans gldB gene encoding a NADP(+) -dependent glycerol dehydrogenase. A basal expression level was observed for gldB , which increased significantly under conditions of hyper-osmotic shock (1 M NaCl). Growth of strains in which gldB was disrupted was severely

  18. Isolated 2-methylbutyrylglycinuria caused by short/branched-chain acyl-CoA dehydrogenase deficiency

    DEFF Research Database (Denmark)

    Andresen, B S; Christensen, E; Corydon, T J

    2000-01-01

    -CoA dehydrogenase and that both wild-type proteins are imported into mitochondria and form tetramers. In conclusion, we report the first mutation in the SBCAD gene, show that it results in an isolated defect in isoleucine catabolism, and indicate that ACAD-8 is a mitochondrial enzyme that functions in valine...

  19. Watermelon glyoxysomal malate dehydrogenase is sorted to peroxisomes of the methylotrophic yeast, Hansenula polymorpha

    NARCIS (Netherlands)

    Klei, I.J. van der; Faber, K.N.; Keizer-Gunnink, I.; Gietl, C.; Harder, W.; Veenhuis, M.

    1993-01-01

    We have studied the fate of the watermelon (Citrullus vulgaris Schrad.) glyoxysomal enzyme, malate dehydrogenase (gMDH), after synthesis in the methylotrophic yeast, Hansenula polymorpha. The gene encoding the precursor form of gMDH (pre-gMDH) was cloned in an H. polymorpha expression vector

  20. Expression of Cellobiose Dehydrogenase from Neurospora crassa in Pichia pastoris and its purification and characterization

    Science.gov (United States)

    A gene encoding cellobiose dehydrogenase (CDH) from Neurospora crassa strain FGSC 2489 has been cloned and expressed in the heterologous host Pichia pastoris, under the control of the AOX1 methanol inducible promoter. Recombinant CDH without the native signal sequence and fused with a his6-tag (rNC-...

  1. CLINICAL AND ECONOMICAL ASSESSMENTS OF METOPROLOL TARTRATE/SUCCINATE USAGE IN PATIENTS WITH ISCHEMIC HEART DISEASE

    Directory of Open Access Journals (Sweden)

    M. V. Soura

    2008-01-01

    Full Text Available Clinical and clinicoeconomical studies review is presented as well as results of author’s comparative cost analysis on metoprolol tartrate (Betaloc and metoprolol succinate (Betaloc ZOK usage in patients with ischemic heart disease. Efficacy of metoprolol therapy is proven in randomized clinical studies in patients with angina and myocardial infarction (MI. In angina patients metoprolol prevents cardiac attacks, MI, reduces nitroglycerine consumption, increases exercise tolerability, prolongs the exercise time before ST segment depression (succinate better than tartrate, decrease of angina intensity. In MI patients metoprolol therapy reduces mortality, sudden death, recurring MI and the rate of early post MI angina attacks. Nowadays metoprolol is the only β-blocker having indication on secondary MI prevention. Besides for the present metoprolol succinate is the only β-blocker with proven direct antisclerosis effect. According to Swedish clinicoeconomical study in patients after MI secondary prevention with metoprolol therapy saves the costs in comparison with placebo. American clinicoeconomical model of metoprolol and atenolol usage in all patients with MI could result in significant reduction in mortality and recurring MI rate, prolong the life and improve its quality, save financial resources. The cost of monthly treatment of angina patient with metoprolol tartrate (Betaloc and metoprolol succinate (Betaloc ZOK is 135 and 354 rubles, respectively. The price range of comparative β-blockers in ascending order is the following: atenolol (Atenolol Nicomed → metoprolol tartrate (Betaloc → metoprolol succinate (Betaloc ZOK → bisoprolol (Concor → nebivolol (Nebilet. In conclusion, metoprolol therapy is the one of mostly economically reasonable approach.

  2. Succination of Protein Disulfide Isomerase Links Mitochondrial Stress and Endoplasmic Reticulum Stress in the Adipocyte During Diabetes.

    Science.gov (United States)

    Manuel, Allison M; Walla, Michael D; Faccenda, Adam; Martin, Stephanie L; Tanis, Ross M; Piroli, Gerardo G; Adam, Julie; Kantor, Boris; Mutus, Bulent; Townsend, Danyelle M; Frizzell, Norma

    2017-12-01

    Protein succination by fumarate increases in the adipose tissue of diabetic mice and in adipocytes matured in high glucose as a result of glucotoxicity-driven mitochondrial stress. The endoplasmic reticulum (ER) oxidoreductase protein disulfide isomerase (PDI) is succinated in adipocytes that are matured in high glucose, and in this study we investigated whether succination would alter PDI oxidoreductase activity, directly linking mitochondrial stress and ER stress. Protein succination and the ER stress marker C/EBP homologous protein (CHOP) were diminished after pharmaceutical targeting of mitochondrial stress with the chemical uncoupler niclosamide in adipocytes matured in high-glucose concentrations. PDI was succinated by fumarate on both CXXC-containing active sites, contributing to reduced enzymatic activity. Succinated PDI decreased reductase activity in adipocytes matured in high glucose, and in db/db epididymal adipose tissue, in association with increased levels of CHOP. PDI succination was increased in fumarase knockdown adipocytes, leading to reduced PDI oxidoreductase activity, increased CHOP levels, and pro-inflammatory cytokine secretion, confirming the specific role of elevated fumarate levels in contributing to ER stress. In addition, PDI succination and ER stress were decreased, and PDI reductase activity was restored when exposure to chronic high glucose was limited, highlighting the importance of calorie restriction in the improvement of adipocyte metabolic function. These experiments identify PDI succination as a novel biochemical mechanism linking altered mitochondrial metabolism to ER stress in the adipocyte during diabetes. The current study demonstrates that early biochemical changes in mitochondrial metabolism have important implications for the development of adipocyte stress. Antioxid. Redox Signal. 27, 1281-1296.

  3. 2-methylbutyryl-CoA dehydrogenase deficiency associated with autism and mental retardation: a case report

    DEFF Research Database (Denmark)

    Kanavin, Øjvind; Woldseth, Berit; Jellum, Egil

    2007-01-01

    previously reported cases with SBCADD, both originating from Somalia and Eritrea, indicating that it is relatively prevalent in this population. Autism has not previously been described with mutations in this gene, thus expanding the clinical spectrum of SBCADD. PMID: 17883863 [PubMed - in process]......ABSTRACT: BACKGROUND: 2-methylbutyryl-CoA dehydrogenase deficiency or short/branched chain acyl-CoA dehydrogenase deficiency (SBCADD) is caused by a defect in the degradation pathway of the amino acid L-isoleucine. METHODS: We report a four-year-old mentally retarded Somali boy with autism...

  4. 2-methylbutyryl-CoA dehydrogenase deficiency associated with autism and mental retardation

    DEFF Research Database (Denmark)

    Kanavin, Oivind J; Woldseth, Berit; Jellum, Egil

    2007-01-01

    BACKGROUND: 2-methylbutyryl-CoA dehydrogenase deficiency or short/branched chain acyl-CoA dehydrogenase deficiency (SBCADD) is caused by a defect in the degradation pathway of the amino acid L-isoleucine. METHODS: We report a four-year-old mentally retarded Somali boy with autism and a history...... cases with SBCADD, both originating from Somalia and Eritrea, indicating that it is relatively prevalent in this population. Autism has not previously been described with mutations in this gene, thus expanding the clinical spectrum of SBCADD....

  5. Cloning, overexpression, purification, crystallization and preliminary X-ray analysis of 3-ketosteroid Delta(4)-(5 alpha)-dehydrogenase from Rhodococcus jostii RHA1

    NARCIS (Netherlands)

    van Oosterwijk, Niels; Knol, Jan; Dijkhuizen, Lubbert; van der Geize, Robert; Dijkstra, Bauke

    2011-01-01

    3-Ketosteroid dehydrogenases are flavoproteins which play key roles in steroid ring degradation. The enzymes are abundantly present in actinobacteria, including the catabolic powerhouse Rhodococcus jostii and the pathogenic species R. equi and Mycobacterium tuberculosis. The gene for 3-ketosteroid

  6. The effect of pH and ADP on ammonia affinity for human glutamate dehydrogenases

    DEFF Research Database (Denmark)

    Zaganas, Ioannis; Pajecka, Kamilla; Nielsen, Camilla Wendel

    2013-01-01

    Glutamate dehydrogenase (GDH) uses ammonia to reversibly convert α-ketoglutarate to glutamate using NADP(H) and NAD(H) as cofactors. While GDH in most mammals is encoded by a single GLUD1 gene, humans and other primates have acquired a GLUD2 gene with distinct tissue expression profile. The two h...... of the kidney during systemic acidosis. The reverse could apply for conditions of local or systemic hyperammonemia or alkalosis....

  7. 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.

  8. The influence of antioxidant and post-synthetic treatment on the properties of biodegradable poly(butylene succinate)s modified with poly(propylene oxide)

    OpenAIRE

    DRAGANA PEPIC; MARIJA RADOICIC; MARIJA S. NIKOLIC; JASNA DJONLAGIC

    2007-01-01

    Novel poly(ester–ether)s based on poly(butylene succinate) (PBS) as the hard segments and 30 mass % of poly(propylene oxide) (PPO) as the soft segments were synthesized with varying amount of the antioxidant (N,N'-diphenyl-p-phenylenediamine, DPPD). The influences of the addition of DPPD and the impact of post-synthetic treatment by precipitation on the molecular structure, thermal and physical properties, as well as on the storage stability of the biodegradable aliphatic copolyesters, were i...

  9. Enhancement of Ganoderic Acid Accumulation by Overexpression of an N-Terminally Truncated 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase Gene in the Basidiomycete Ganoderma lucidum

    Science.gov (United States)

    Xu, Jun-Wei; Xu, Yi-Ning

    2012-01-01

    Ganoderic acids produced by Ganoderma lucidum, a well-known traditional Chinese medicinal mushroom, exhibit antitumor and antimetastasis activities. Genetic modification of G. lucidum is difficult but critical for the enhancement of cellular accumulation of ganoderic acids. In this study, a homologous genetic transformation system for G. lucidum was developed for the first time using mutated sdhB, encoding the iron-sulfur protein subunit of succinate dehydrogenase, as a selection marker. The truncated G. lucidum gene encoding the catalytic domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) was overexpressed by using the Agrobacterium tumefaciens-mediated transformation system. The results showed that the mutated sdhB successfully conferred carboxin resistance upon transformation. Most of the integrated transfer DNA (T-DNA) appeared as a single copy in the genome. Moreover, deregulated constitutive overexpression of the HMGR gene led to a 2-fold increase in ganoderic acid content. It also increased the accumulation of intermediates (squalene and lanosterol) and the upregulation of downstream genes such as those of farnesyl pyrophosphate synthase, squalene synthase, and lanosterol synthase. This study demonstrates that transgenic basidiomycete G. lucidum is a promising system to achieve metabolic engineering of the ganoderic acid pathway. PMID:22941092

  10. Lactate dehydrogenase A silencing in IDH mutant gliomas.

    Science.gov (United States)

    Chesnelong, Charles; Chaumeil, Myriam M; Blough, Michael D; Al-Najjar, Mohammad; Stechishin, Owen D; Chan, Jennifer A; Pieper, Russell O; Ronen, Sabrina M; Weiss, Samuel; Luchman, H Artee; Cairncross, J Gregory

    2014-05-01

    Mutations of the isocitrate dehydrogenase 1 and 2 gene (IDH1/2) were initially thought to enhance cancer cell survival and proliferation by promoting the Warburg effect. However, recent experimental data have shown that production of 2-hydroxyglutarate by IDH mutant cells promotes hypoxia-inducible factor (HIF)1α degradation and, by doing so, may have unexpected metabolic effects. We used human glioma tissues and derived brain tumor stem cells (BTSCs) to study the expression of HIF1α target genes in IDH mutant ((mt)) and IDH wild-type ((wt)) tumors. Focusing thereafter on the major glycolytic enzyme, lactate dehydrogenase A (LDHA), we used standard molecular methods and pyrosequencing-based DNA methylation analysis to identify mechanisms by which LDHA expression was regulated in human gliomas. We found that HIF1α-responsive genes, including many essential for glycolysis (SLC2A1, PDK1, LDHA, SLC16A3), were underexpressed in IDH(mt) gliomas and/or derived BTSCs. We then demonstrated that LDHA was silenced in IDH(mt) derived BTSCs, including those that did not retain the mutant IDH1 allele (mIDH(wt)), matched BTSC xenografts, and parental glioma tissues. Silencing of LDHA was associated with increased methylation of the LDHA promoter, as was ectopic expression of mutant IDH1 in immortalized human astrocytes. Furthermore, in a search of The Cancer Genome Atlas, we found low expression and high methylation of LDHA in IDH(mt) glioblastomas. To our knowledge, this is the first demonstration of downregulation of LDHA in cancer. Although unexpected findings, silencing of LDHA and downregulation of several other glycolysis essential genes raise the intriguing possibility that IDH(mt) gliomas have limited glycolytic capacity, which may contribute to their slow growth and better prognosis.

  11. Isolation and biochemical characterization of a glucose dehydrogenase from a hay infusion metagenome.

    Directory of Open Access Journals (Sweden)

    Alexander Basner

    Full Text Available Glucose hydrolyzing enzymes are essential to determine blood glucose level. A high-throughput screening approach was established to identify NAD(P-dependent glucose dehydrogenases for the application in test stripes and the respective blood glucose meters. In the current report a glucose hydrolyzing enzyme, derived from a metagenomic library by expressing recombinant DNA fragments isolated from hay infusion, was characterized. The recombinant clone showing activity on glucose as substrate exhibited an open reading frame of 987 bp encoding for a peptide of 328 amino acids. The isolated enzyme showed typical sequence motifs of short-chain-dehydrogenases using NAD(P as a co-factor and had a sequence similarity between 33 and 35% to characterized glucose dehydrogenases from different Bacillus species. The identified glucose dehydrogenase gene was expressed in E. coli, purified and subsequently characterized. The enzyme, belonging to the superfamily of short-chain dehydrogenases, shows a broad substrate range with a high affinity to glucose, xylose and glucose-6-phosphate. Due to its ability to be strongly associated with its cofactor NAD(P, the enzyme is able to directly transfer electrons from glucose oxidation to external electron acceptors by regenerating the cofactor while being still associated to the protein.

  12. 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.

  13. Increased superoxide accumulation in pyruvate dehydrogenase complex deficient fibroblasts.

    Science.gov (United States)

    Glushakova, Lyudmyla G; Judge, Sharon; Cruz, Alex; Pourang, Deena; Mathews, Clayton E; Stacpoole, Peter W

    2011-11-01

    The pyruvate dehydrogenase complex (PDC) oxidizes pyruvate to acetyl CoA and is critically important in maintaining normal cellular energy homeostasis. Loss-of-function mutations in PDC give rise to congenital lactic acidosis and to progressive cellular energy failure. However, the subsequent biochemical consequences of PDC deficiency that may contribute to the clinical manifestations of the disorder are poorly understood. We postulated that altered flux through PDC would disrupt mitochondrial electron transport, resulting in oxidative stress. Compared to cells from 4 healthy subjects, primary cultures of skin fibroblasts from 9 patients with variable mutations in the gene encoding the alpha subunit (E1α) of pyruvate dehydrogenase (PDA1) demonstrated reduced growth and viability. Superoxide (O(2)(.-)) from the Qo site of complex III of the electron transport chain accumulated in these cells and was associated with decreased activity of manganese superoxide dismutase. The expression of uncoupling protein 2 was also decreased in patient cells, but there were no significant changes in the expression of cellular markers of protein or DNA oxidative damage. The expression of hypoxia transcription factor 1 alpha (HIF1α) also increased in PDC deficient fibroblasts. We conclude that PDC deficiency is associated with an increase in O(2)(.-) accumulation coupled to a decrease in mechanisms responsible for its removal. Increased HIF1α expression may contribute to the increase in glycolytic flux and lactate production in PDC deficiency and, by trans-activating pyruvate dehydrogenase kinase, may further suppress residual PDC activity through phosphorylation of the E1α subunit. Copyright © 2011 Elsevier Inc. All rights reserved.

  14. Enterobacter sp. LU1 as a novel succinic acid producer - co-utilization of glycerol and lactose.

    Science.gov (United States)

    Podleśny, Marcin; Jarocki, Piotr; Wyrostek, Jakub; Czernecki, Tomasz; Kucharska, Jagoda; Nowak, Anna; Targoński, Zdzisław

    2017-03-01

    Succinic acid is an important C4-building chemical platform for many applications. A novel succinic acid-producing bacterial strain was isolated from goat rumen. Phylogenetic analysis based on the 16S rRNA sequence and physiological analysis indicated that the strain belongs to the genus Enterobacter. This is the first report of a wild bacterial strain from the genus Enterobacter that is capable of efficient succinic acid production. Co-fermentation of glycerol and lactose significantly improved glycerol utilization under anaerobic conditions, debottlenecking the utilization pathway of this valuable biodiesel waste product. Succinic acid production reached 35 g l -1 when Enterobacter sp. LU1 was cultured in medium containing 50 g l -1 of glycerol and 25 g l -1 of lactose as carbon sources. © 2016 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  15. The impact of nanoclay on the crystal growth kinetics and morphology of biodegradable poly(ethylene succinate) composite

    CSIR Research Space (South Africa)

    Bandyopadhyay, J

    2012-07-01

    Full Text Available The impact of nanoclay on the isothermal crystal growth kinetics and morphology of biodegradable poly(ethylene succinate) (PES) is reported. A PES composite (PESNC) containing 5 wt% organically modified montmorillonite, was prepared via solvent...

  16. [The effect of succinate on respiration, transamination, and pyruvate formation in cells of the yeast Dipodascus magnusii].

    Science.gov (United States)

    Il'chenko, A P; Ogorelyshev, D I; Shishkanova, N V; Sokolov, A P; Finogenova, T V; Kondrashova, M N

    2005-01-01

    The effect of succinate on the growth and respiration of the yeast Dipodascus magnusii VKM Y-1072, which is auxotrophic for thiamine and biotin, was studied. The addition of succinate to a culture grown on glucose was found to activate the respiration of cells on various substrates by enhancing the processes related to transamination reactions. In this case, aerobic fermentation (ethanol production) decreased, whereas pyruvate production increased. When succinate was added to the medium as the sole carbon source, it supported yeast growth in the absence of one of the two vitamins, thiamine or biotin, but not both. The yeast metabolism was completely respiratory, without any signs of aerobic fermentation. A drastic rise in pyruvate production in the yeast grown on glucose in the presence of succinate and the absence of biotin are also indicative of metabolic changes.

  17. Stability of coordination compounds of Ni2+ and Co2+ ions with succinic acid anion in water-ethanol solvents

    Science.gov (United States)

    Tukumova, N. V.; Dieu Thuan, Tran Thi; Usacheva, T. R.; Koryshev, N. E.; Sharnin, V. A.

    2017-04-01

    Stability constants of the coordination compounds of nickel(II) and cobalt(II) ions with succinic acid anion in water-ethanol solvents are determined via potentiometric titration at ionic strength of 0.1 and at T = 298.15 K. It is found that logβ values of monoligand complexes of these ions and succinic acid anions rise along with the content of ethanol in solution ( X EtOH = 0-0.7 mole fractions). Based on an analysis of the thermodynamic characteristics of the solvation of the reagents involved in complex formation, it is found that the increased stability of succinate complexes of nickel(II) and cobalt(II) ions in water-ethanol solvents is mainly determined by the weakening of the solvation of succinic acid anion (Y2-).

  18. Comparison of high-resolution melting analysis with direct sequencing for the detection of recurrent mutations in DNA methyltransferase 3A and isocitrate dehydrogenase 1 and 2 genes in acute myeloid leukemia patients.

    Science.gov (United States)

    Gorniak, Patryk; Ejduk, Anna; Borg, Katarzyna; Makuch-Lasica, Hanna; Nowak, Grazyna; Lech-Maranda, Ewa; Prochorec-Sobieszek, Monika; Warzocha, Krzysztof; Juszczynski, Przemyslaw

    2016-02-01

    Acute myeloid leukemia (AML) cells harbor frequent mutations in genes responsible for epigenetic modifications. Increasing evidence of clinical role of DNMT3A and IDH1/2 mutations highlights the need for a robust and inexpensive test to identify these mutations in routine diagnostic work-up. Herein, we compared routinely used direct sequencing method with high-resolution melting (HRM) assay for screening DNMT3A and IDH1/2 mutations in patients with AML. We show very high concordance between HRM and Sanger sequencing (100% samples for IDH2-R140 and DNMT3-R882 mutations, 99% samples for IDH1-R132 and IDH2-R172 mutations). HRM method reported no false-negative results, suggesting that it can be used for mutations screening. Moreover, HRM displayed much higher sensitivity in comparison with DNA sequencing in all assessed loci. With Sanger sequencing, robust calls were observed when the sample contained 50% of mutant DNA in the background of wild-type DNA. In marked contrast, the detection limit of HRM improved down to 10% of mutated DNA. Given the ubiquitous presence of wild-type DNA background in bone marrow aspirates and clonal variations regarding mutant allele burden, these results favor HRM as a sensitive, specific, labor-, and cost-effective tool for screening and detection of mutations in IDH1/2 and DNMT3A genes in patients with AML. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  19. Enhanced succinic acid production under acidic conditions by introduction of glutamate decarboxylase system in E. coli AFP111.

    Science.gov (United States)

    Wu, Mingke; Li, Xiaozhan; Guo, Shunfeng; Lemma, Wubliker Dessie; Zhang, Wenming; Ma, Jiangfeng; Jia, Honghua; Wu, Hao; Jiang, Min; Ouyang, Pingkai

    2017-04-01

    Biological synthesis of succinic acid at low pH values was favored since it not only decreased investment cost but also simplified downstream purification process. In this study, the feasibility of using glutamate decarboxylase system to improve succinic acid production of Escherichia coli AFP111, a succinate-producing candidate with mutations in pfl, ldhA, and ptsG, under acidic conditions was investigated. By overexpressing gadBC operon in AFP111, a recombinant named as BA201 (AFP111/pMD19T-gadBC) was constructed. Fermentation at pH 5.6 showed that 30 g L -1 glucose was consumed and 26.58 g L -1 succinic acid was produced by BA201, which was 1.22- and 1.32-fold higher than that by the control BA200 (AFP111/pMD19T) containing the empty vector. Analysis of intracellular enzymes activities and ATP concentrations revealed that the activities of key enzymes involved in glucose uptake and products synthesis and intracellular ATP levels were all increased after overexpression of gadBC which were benefit for cell metabolism under weak acidic conditions. To further improve the succinic acid titer by recombinant BA201 at pH 5.6, the extracellular glutamate concentration was optimized and the final succinic acid titer increased 20.4% to 32.01 g L -1 . Besides, the fermentation time was prolonged by repetitive fermentation and additional 15.78 g L -1 succinic acid was produced by recovering cells into fresh medium. The results here demonstrated a potential strategy of overexpressing gadBC for increased succinic acid production of E. coli AFP111 under weak acidic conditions.

  20. Glusoce-6-phosphate dehydrogenase- History and diagnosis

    Directory of Open Access Journals (Sweden)

    K Gautam

    2016-09-01

    Full Text Available Glucose-6-phosphate dehydrogenase deficiency is the most common enzymatic defect of red blood cells, which increases the vulnerability of erythrocytes to oxidative stress leading to hemolytic anemia. Since its identification more than 60 years ago, much has been done with respect to its clinical diagnosis, laboratory diagnosis and treatment. Association of G6PD is not just limited to anti malarial drugs, but a vast number of other diseases. In this article, we aimed to review the history of Glucose-6-phosphate dehydrogenase, the diagnostic methods available along with its association with other noncommunicable diseases. 

  1. Inducible xylitol dehydrogenases in enteric bacteria.

    OpenAIRE

    Doten, R C; Mortlock, R P

    1985-01-01

    Morganella morganii ATCC 25829, Providencia stuartii ATCC 25827, Serratia marcescens ATCC 13880, and Erwinia sp. strain 4D2P were found to induce a xylitol dehydrogenase when grown on a xylitol-containing medium. The xylitol dehydrogenases were partially purified from the four strains, and those from M. morganii ATCC 25829, P. stuartii ATCC 25827, and S. marcescens ATCC 13880 were all found to oxidize xylitol to D-xylulose. These three enzymes had KmS for xylitol of 7.1 to 16.4 mM and molecul...

  2. Miscibility, crystallization and mechanical properties of biodegradable blends of poly(L-lactic acid) and poly(butylene succinate-b-ethylene succinate) multiblock copolymer

    International Nuclear Information System (INIS)

    Jiao, Ling; Huang, Cai-Li; Zeng, Jian-Bing; Wang, Yu-Zhong; Wang, Xiu-Li

    2012-01-01

    Highlights: ► The blend of PLLA and PBES showed limited miscibility. ► The crystallization rate of PLLA was accelerated by blending with PBES. ► The crystal structures of PLLA and PBES did not change. - Abstract: Poly(L-lactic acid) (PLLA) is regarded as one of the most promising biobased and biodegradable polymers. However, its application was restricted due to the brittle nature. In the present study, PLLA was blended with a novel biodegradable flexible multiblock copolymer, poly(butylene succinate-b-ethylene succinate) (PBES) to produce new biodegradable materials. PLLA/PBES blends with different composition were prepared by solution blending and casting method with chloroform as a mutual solvent. Miscibility, crystallization behavior, and mechanical properties of the blends were investigated by differential scanning calorimetry (DSC), wide-angle X-ray diffraction (WAXD), and tensile tests. The results indicated that PLLA and PBES showed limited miscibility in the amorphous phase. The crystallization rate of PLLA was accelerated with the increase of PBES in the blends while the crystallization mechanism did not change. The results of tensile tests suggest that the blends showed longer elongation at break than neat PLLA. The elongation at break of PLLA was obtained to be 10%, and those of PLLA/PBES 80/20, 60/40, 40/60 and 20/80 were 29, 110, 442, and 455%, respectively.

  3. Reactive processing of maleic anhydride-grafted poly(butylene succinate and the compatibilizing effect on poly(butylene succinate nanocomposites

    Directory of Open Access Journals (Sweden)

    Z. A. Mohd Ishak

    2013-04-01

    Full Text Available In this study, maleic anhydride-grafted poly(butylene succinate (PBS-g-MA was synthesized via reactive meltgrafting process using different initiator contents. The grafting efficiency was increased with the initiator content, manifested by the higher degree of grafting in PBS-g-MA. The grafting reaction was confirmed through Fourier transform infrared (FTIR spectroscopy and nuclear magnetic resonance (NMR spectroscopy. Then, PBS-g-MA was incorporated into organo-montmorillonite (OMMT filled poly(butylene succinate (PBS nanocomposites as compatibilizer. Mechanical properties of PBS nanocomposites were enhanced after compatibilized with PBS-g-MA, due to the better dispersion of OMMT in PBS matrix and the improved filler-matrix interfacial interactions. This was verifiable through X-ray diffraction (XRD, transmission electron microscopy (TEM and scanning electron microscopy (SEM. Differential scanning calorimetry (DSC showed that the degree of crystallinity and melting temperature increased after addition of PBS-g-MA. However, the presence of PBS-g-MA did not favor the thermal stability of the nanocomposites, as reported in the thermogravimetry (TGA.

  4. Influence of VO2+ ions on structural and optical properties of potassium succinate-succinic acid single crystal for non-linear optical applications

    Science.gov (United States)

    Juliet sheela, K.; Subramanian, P.

    2018-04-01

    A transparent and good optical quality semi organic single crystal of vanadium doped potassium succinate-succinic acid (KSSA) was synthesized by slow evaporation technique at room temperature. The structural perfection was supported by the powder XRD of the KSSA-VO2+ single crystal. Optical behavior of the material was discovered from the absorption and transmission spectra of UV-vis-NIR characterization. Functional group and presence of metal ion in the specimen are depicted from FTIR traces. From the photoluminescence studies, emission of wavelength in the violet region (418 nm) at the excitation of 243 nm could be ascertained. EDAX, SEM measurements identify presence of elements and pictures the step-line growth and the imperfection presents in the grown crystal. EPR analysis extracts the information about the local site symmetry around the impurity ion, molecular orbital coefficients, admixture coefficients and ground state wave function of VO2+ doped KSSA single crystal. Second harmonic generation (SHG) efficiency of the grown crystal was investigated to explore the NLO characteristic of the material.

  5. IR Laser Ablative Decomposition and Depolymerization/Repolymerization of Poly(ethylene succinate)

    Czech Academy of Sciences Publication Activity Database

    Pokorná, Dana; Galíková, Anna; Šubrt, Jan; Blechta, Vratislav; Pola, Josef

    2006-01-01

    Roč. 91, č. 12 (2006), s. 3383-3389 ISSN 0141-3910 R&D Projects: GA ČR(CZ) GA104/04/2028 Institutional research plan: CEZ:AV0Z40720504; CEZ:AV0Z40320502 Keywords : poly(ethylene succinate) * laser degradation * laser-produced films Subject RIV: CC - Organic Chemistry Impact factor: 2.174, year: 2006

  6. Alpha-Tocopheryl succinate causes mitochondrial permeabilization by preferential formation of Bak channels

    Czech Academy of Sciences Publication Activity Database

    Procházka, L.; Dong, L.F.; Vališ, Karel; Freeman, R.; Ralph, S.J.; Turánek, J.; Neužil, Jiří

    2010-01-01

    Roč. 15, č. 7 (2010), s. 782-794 ISSN 1360-8185 R&D Projects: GA AV ČR(CZ) KAN200520703; GA AV ČR(CZ) IAA500520702; GA AV ČR(CZ) IAA500520602 Institutional research plan: CEZ:AV0Z50520701 Keywords : Vitamin E succinate * apoptosis * Noxa-Bak Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.397, year: 2010

  7. Volatility of NH3 from internally mixed sodium succinate-NH4SO4 particles

    Science.gov (United States)

    Wang, Na; Zhang, Yunhong

    2016-04-01

    Contributing the complicacy of atmospheric constituents, aerosol particles may undergo complicated heterogeneous reactions that have profound consequences on their hygroscopic properties and volatility. Ammonia (NH3) is a ubiquitous trace atmospheric gas in the troposphere and has negative effects on human health and climate forcing of ambient aerosols. In addition, atmospheric cycle of NH3 is complex in atmosphere, therefore it necessary to get insights to the complexity of gas-to-aerosol NH3 partitioning, which results in large uncertainties in the sources and distributions of NH3. By using in-situ Fourier transform infrared spectroscopy and attenuated total reflection (FTIR-ATR), we report here the volatility of NH3 from the laboratory generated sodium succinate with ammonium sulfate ((NH4)2SO4) at a 1:1 molar ratio as well as its effect on the hygroscopicity of the mixtures. The loss of the NH4+ peak at 1451cm-1 and the formation of peaks at 1718 and 1134 cm-1 due to C = O stretching asymmetric vibration of -COOH and ν3 (SO42-) stretching of sodium sulfate indicate that sodium succinate reacts with (NH4)2SO4, releasing NH3 and forming succinic acid and sodium sulfate on dehydration process. The formation of less hygroscopic succinic acid and volatility of NH3 in mixtures leads to a significant decrease in the total water content. To the best of our knowledge, this is the first report of the reaction between (NH4)2SO4 and dicarboxylate salts, which may represent an important particle-gas partitioning for ammonia and thus elucidate another underlying ammonia cycle in atmosphere. These results could be helpful to understand the mutual transformation process of dicarboxylic acids and dicarboxylate salts.

  8. Effect of vitamin E succinate on inflammatory cytokines induced by high-intensity interval training

    Directory of Open Access Journals (Sweden)

    Hadi Sarir

    2015-01-01

    Full Text Available Aim and Scope: The anti-inflammatory effect of vitamin E under moderate exercises has been evaluated. However, the effect of vitamin E succinate, which has more potent anti-inflammatory effect than other isomers of vitamin E has not been evaluated. Therefore, the aim of the present study was to evaluate the effects of vitamin E succinate on tumor necrosis factor alpha (TNF-a and interleukin-6 (IL-6 production induced by high-intensity interval training (HIIT. Materials and Methods: In the present study, 24 rats were randomly divided into control (C, supplementation (S, HIIT, and HIIT + supplementation (HIIT+S groups. HIIT training protocol on a treadmill (at a speed of 40-54 m/min and vitamin E succinate supplementation (60 mg/kg/day was conducted for 6 weeks. Results: Serum IL-6 in the HIIT group significantly increased compared with the C group (350.42 ± 123.31 pg/mL vs 158.60 ± 41.96 pg/mL; P = 0.002. Also, serum TNF-a concentrations significantly enhanced (718.15 ± 133.42 pg/mL vs 350.87 ± 64.93 pg/mL; P = 0.001 in the HIIT group compared with the C group. Treatment of the training group with vitamin E numerically reduced IL-6 and TNF-a when compared with the HIIT group (217.31 ± 29.21 and 510.23 ± 217.88, respectively, P > 0.05. However, no significant changes were observed in serum TNF-a (P = 0.31 and IL-6 (P = 0.52 concentrations in the HIIT + S group compared with the C group. Conclusion: HIIT-induced IL-6 and TNF-α decreased by administration of Vitamin E succinate.

  9. Polymeric nanoparticles loaded with dexamethasone or α-tocopheryl succinate to prevent cisplatin-induced ototoxicity.

    Science.gov (United States)

    Martín-Saldaña, Sergio; Palao-Suay, Raquel; Aguilar, María Rosa; Ramírez-Camacho, Rafael; San Román, Julio

    2017-04-15

    The aim of this work is the development of highly protective agents to be administered locally within the middle ear to avoid cisplatin-induced ototoxicity, which affects to 100% of the clinical patients at ultra-high concentrations (16mg/kg). The protective agents are based on polymeric nanoparticles loaded with dexamethasone or α-tocopheryl succinate as anti-inflammarory and anti-apoptotic molecules. Dexamethasone and α-tocopheryl succinate are poorly soluble in water and present severe side effects when systemic administered during long periods of time. Their incorporation in the hydrophobic core of nanoparticles with the appropriate hydrodynamic properties provides the desired effects in vitro (lower cisplatin-induced toxicity, decreasing of caspase 3/7 activity, and lower IL-1β release) and in vivo (reducing the hearing loss at the local level). The local administration of the nanoparticles by bullostomy provides an adequate dose of drug without systemic interference with the chemotherapeutic effect of cisplatin. 100% of the cancer patients receiving ultra-high doses of CDDP (16mg/kg) suffer severe hearing loss, being a limiting factor in antineoplastic treatments. In this paper we describe the application of polymeric nanoparticles loaded with dexamethasone or α-tocopheryl succinate to palliate the cisplatin ototoxicity derived from chemotherapy treatment. These new nanoparticles, that encapsulate, transport, and deliver dexamethasone or α-tocopheryl succinate in the middle ear, are able to partially prevent ototoxicity derived from high doses of CDDP. This is an interdisciplinary study in which in vitro and in vivo experiments are described and extensively discussed. The importance of the results opens an excellent opportunity to the translation to the clinic. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  10. Investigation of griseofulvin and hydroxypropylmethyl cellulose acetate succinate miscibility in ball milled solid dispersions.

    Science.gov (United States)

    Al-Obaidi, Hisham; Lawrence, M Jayne; Al-Saden, Noor; Ke, Peng

    2013-02-25

    Solid dispersions of varying weight ratios compositions of the nonionic drug, griseofulvin and the hydrophilic, anionic polymer, hydroxylpropylmethyl cellulose acetate succinate, have been prepared by ball milling and the resulting samples characterized using a combination of Fourier transform infra-red spectroscopy, X-ray powder diffraction and differential scanning calorimetry. The results suggest that griseofulvin forms hydrogen bonds with the hydroxylpropylmethyl cellulose acetate succinate polymer when prepared in the form of a solid dispersion but not when prepared in a physical mixture of the same composition. As anticipated, the actual measured glass transition temperature of the solid dispersions displayed a linear relationship between that predicted using the Gordon-Taylor and Fox equations assuming ideal mixing, but interestingly only at griseofulvin contents less than 50 wt%. At griseofulvin concentrations greater than this, the measured glass transition temperature of the solid dispersions was almost constant. Furthermore, the crystalline content of the solid dispersions, as determined by differential scanning calorimetry and X-ray powder diffraction followed a similar trend in that the crystalline content significantly decreased at ratios less than 50 wt% of griseofulvin. When the physical mixtures of griseofulvin and the hydroxylpropylmethyl cellulose acetate succinate polymer were analyzed using the Flory-Huggins model, a negative free energy of mixing with an interaction parameter of -0.23 were obtained. Taken together these results suggest that anionic hydrophilic hydroxylpropylmethyl cellulose acetate succinate polymer is a good solvent for crystalline nonionic griseofulvin with the solubility of griseofulvin in the solid dispersion being was estimated to be within the range 40-50 wt%. Below this solubility limit, the amorphous drug exists as amorphous glassy solution while above these values the system is supersaturated and glassy suspension and

  11. Effect of vitamin E succinate on inflammatory cytokines induced by high-intensity interval training.

    Science.gov (United States)

    Sarir, Hadi; Emdadifard, Ghodsieh; Farhangfar, Homayoun; TaheriChadorneshin, Hossein

    2015-12-01

    The anti-inflammatory effect of vitamin E under moderate exercises has been evaluated. However, the effect of vitamin E succinate, which has more potent anti-inflammatory effect than other isomers of vitamin E has not been evaluated. Therefore, the aim of the present study was to evaluate the effects of vitamin E succinate on tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) production induced by high-intensity interval training (HIIT). In the present study, 24 rats were randomly divided into control (C), supplementation (S), HIIT, and HIIT + supplementation (HIIT+S) groups. HIIT training protocol on a treadmill (at a speed of 40-54 m/min) and vitamin E succinate supplementation (60 mg/kg/day) was conducted for 6 weeks. Serum IL-6 in the HIIT group significantly increased compared with the C group (350.42 ± 123.31 pg/mL vs 158.60 ± 41.96 pg/mL; P = 0.002). Also, serum TNF-α concentrations significantly enhanced (718.15 ± 133.42 pg/mL vs 350.87 ± 64.93 pg/mL; P = 0.001) in the HIIT group compared with the C group. Treatment of the training group with vitamin E numerically reduced IL-6 and TNF-α when compared with the HIIT group (217.31 ± 29.21 and 510.23 ± 217.88, respectively, P > 0.05). However, no significant changes were observed in serum TNF-α (P = 0.31) and IL-6 (P = 0.52) concentrations in the HIIT + S group compared with the C group. HIIT-induced IL-6 and TNF-α decreased by administration of Vitamin E succinate.

  12. Efficient and repeated production of succinic acid by turning sugarcane bagasse into sugar and support.

    Science.gov (United States)

    Chen, Pengcheng; Tao, Shengtao; Zheng, Pu

    2016-07-01

    Here we reported an endeavor in making full use of sugarcane bagasse for biological production of succinic acid. Through NaOH pre-treatment and multi-enzyme hydrolysis, a reducing sugar solution mainly composed of glucose and xylose was obtained from the sugarcane bagasse. By optimizing portions of cellulase, xylanase, β-glucanase and pectinase in the multi-enzyme "cocktail", the hydrolysis percentage of the total cellulose in pre-treated sugarcane bagasse can be as high as 88.5%. A. succinogenes CCTCC M2012036 was used for converting reducing sugars into succinic acid in a 3-L bioreactor with a sugar-fed strategy to prevent cell growth limitation. Importantly, cells were found to be adaptive on the sugarcane bagasse residue, offering possibilities of repeated batch fermentation and replacement for MgCO3 with soluble NaHCO3 in pH modulation. Three cycles of fermentation without activity loss were realized with the average succinic acid yield and productivity to be 80.5% and 1.65g·L(-1)·h(-1). Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Co-Consumption of Methanol and Succinate by Methylobacterium extorquens AM1

    Science.gov (United States)

    Peyraud, Rémi; Kiefer, Patrick; Christen, Philipp; Portais, Jean-Charles; Vorholt, Julia A.

    2012-01-01

    Methylobacterium extorquens AM1 is a facultative methylotrophic Alphaproteobacterium and has been subject to intense study under pure methylotrophic as well as pure heterotrophic growth conditions in the past. Here, we investigated the metabolism of M. extorquens AM1 under mixed substrate conditions, i.e., in the presence of methanol plus succinate. We found that both substrates were co-consumed, and the carbon conversion was two-thirds from succinate and one-third from methanol relative to mol carbon. 13C-methanol labeling and liquid chromatography mass spectrometry analyses revealed the different fates of the carbon from the two substrates. Methanol was primarily oxidized to CO2 for energy generation. However, a portion of the methanol entered biosynthetic reactions via reactions specific to the one-carbon carrier tetrahydrofolate. In contrast, succinate was primarily used to provide precursor metabolites for bulk biomass production. This work opens new perspectives on the role of methylotrophy when substrates are simultaneously available, a situation prevailing under environmental conditions. PMID:23133625

  14. Integrated production of cellulosic bioethanol and succinic acid from industrial hemp in a biorefinery concept.

    Science.gov (United States)

    Kuglarz, Mariusz; Alvarado-Morales, Merlin; Karakashev, Dimitar; Angelidaki, Irini

    2016-01-01

    The aim of this study was to develop integrated biofuel (cellulosic bioethanol) and biochemical (succinic acid) production from industrial hemp (Cannabis sativa L.) in a biorefinery concept. Two types of pretreatments were studied (dilute-acid and alkaline oxidative method). High cellulose recovery (>95%) as well as significant hemicelluloses solubilization (49-59%) after acid-based method and lignin solubilization (35-41%) after alkaline H2O2 method were registered. Alkaline pretreatment showed to be superior over the acid-based method with respect to the rate of enzymatic hydrolysis and ethanol productivity. With respect to succinic acid production, the highest productivity was obtained after liquid fraction fermentation originated from steam treatment with 1.5% of acid. The mass balance calculations clearly showed that 149kg of EtOH and 115kg of succinic acid can be obtained per 1ton of dry hemp. Results obtained in this study clearly document the potential of industrial hemp for a biorefinery. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Plant Glyoxylate/Succinic Semialdehyde Reductases: Comparative Biochemical Properties, Function during Chilling Stress, and Subcellular Localization

    Directory of Open Access Journals (Sweden)

    Adel Zarei

    2017-08-01

    Full Text Available Plant NADPH-dependent glyoxylate/succinic semialdehyde reductases 1 and 2 (cytosolic GLYR1 and plastidial/mitochondrial GLYR2 are considered to be of particular importance under abiotic stress conditions. Here, the apple (Malus × domestica Borkh. and rice (Oryza sativa L. GLYR1s and GLYR2s were characterized and their kinetic properties were compared to those of previously characterized GLYRs from Arabidopsis thaliana [L.] Heynh. The purified recombinant GLYRs had an affinity for glyoxylate and succinic semialdehyde, respectively, in the low micromolar and millimolar ranges, and were inhibited by NADP+. Comparison of the GLYR activity in cell-free extracts from wild-type Arabidopsis and a glyr1 knockout mutant revealed that approximately 85 and 15% of the cellular GLYR activity is cytosolic and plastidial/mitochondrial, respectively. Recovery of GLYR activity in purified mitochondria from the Arabidopsis glyr1 mutant, free from cytosolic GLYR1 or plastidial GLYR2 contamination, provided additional support for the targeting of GLYR2 to mitochondria, as well as plastids. The growth of plantlets or roots of various Arabidopsis lines with altered GLYR activity responded differentially to succinic semialdehyde or glyoxylate under chilling conditions. Taken together, these findings highlight the potential regulation of highly conserved plant GLYRs by NADPH/NADP+ ratios in planta, and their roles in the reduction of toxic aldehydes in plants subjected to chilling stress.

  16. Genetics Home Reference: glucose-6-phosphate dehydrogenase deficiency

    Science.gov (United States)

    ... deficiency Encyclopedia: Glucose-6-phosphate dehydrogenase test Encyclopedia: Hemolytic anemia Encyclopedia: Newborn jaundice Health Topic: Anemia Health Topic: G6PD Deficiency Health Topic: Newborn Screening Genetic and Rare Diseases Information Center (1 link) Glucose-6-phosphate dehydrogenase ...

  17. 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

  18. Coenzyme and effector binding to glutamate dehydrogenase

    NARCIS (Netherlands)

    Zantema, Alt

    1979-01-01

    Glutamaat-dehydrogenase is een enzym dat de reactie katalyseert van 2-oxoglutaraat (substraat), NAD(P)H (co-enzym) en ammonia naar L-glutaminezuur en NAD(P)+. Het enzym is opgebouwd uit 6 identieke subeenheden. Dit proefschrift beschrijft de bestudering van twee aspecten van dit enzym, nl. 1. de

  19. Optimization of Adsorptive Immobilization of Alcohol Dehydrogenases

    NARCIS (Netherlands)

    Trivedi, Archana; Heinemann, Matthias; Spiess, Antje C.; Daussmann, Thomas; Büchs, Jochen

    2005-01-01

    In this work, a systematic examination of various parameters of adsorptive immobilization of alcohol dehydrogenases (ADHs) on solid support is performed and the impact of these parameters on immobilization efficiency is studied. Depending on the source of the enzymes, these parameters differently

  20. 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.

  1. NAD-dependent isocitrate dehydrogenase as a novel target of tributyltin in human embryonic carcinoma cells

    Science.gov (United States)

    Yamada, Shigeru; Kotake, Yaichiro; Demizu, Yosuke; Kurihara, Masaaki; Sekino, Yuko; Kanda, Yasunari

    2014-08-01

    Tributyltin (TBT) is known to cause developmental defects as endocrine disruptive chemicals (EDCs). At nanomoler concentrations, TBT actions were mediated by genomic pathways via PPAR/RXR. However, non-genomic target of TBT has not been elucidated. To investigate non-genomic TBT targets, we performed comprehensive metabolomic analyses using human embryonic carcinoma NT2/D1 cells. We found that 100 nM TBT reduced the amounts of α-ketoglutarate, succinate and malate. We further found that TBT decreased the activity of NAD-dependent isocitrate dehydrogenase (NAD-IDH), which catalyzes the conversion of isocitrate to α-ketoglutarate in the TCA cycle. In addition, TBT inhibited cell growth and enhanced neuronal differentiation through NAD-IDH inhibition. Furthermore, studies using bacterially expressed human NAD-IDH and in silico simulations suggest that TBT inhibits NAD-IDH due to a possible interaction. These results suggest that NAD-IDH is a novel non-genomic target of TBT at nanomolar levels. Thus, a metabolomic approach may provide new insights into the mechanism of EDC action.

  2. Gene

    Data.gov (United States)

    U.S. Department of Health & Human Services — Gene integrates information from a wide range of species. A record may include nomenclature, Reference Sequences (RefSeqs), maps, pathways, variations, phenotypes,...

  3. Increasing anaerobic acetate consumption and ethanol yields in Saccharomyces cerevisiae with NADPH-specific alcohol dehydrogenase.

    Science.gov (United States)

    Henningsen, Brooks M; Hon, Shuen; Covalla, Sean F; Sonu, Carolina; Argyros, D Aaron; Barrett, Trisha F; Wiswall, Erin; Froehlich, Allan C; Zelle, Rintze M

    2015-12-01

    Saccharomyces cerevisiae has recently been engineered to use acetate, a primary inhibitor in lignocellulosic hydrolysates, as a cosubstrate during anaerobic ethanolic fermentation. However, the original metabolic pathway devised to convert acetate to ethanol uses NADH-specific acetylating acetaldehyde dehydrogenase and alcohol dehydrogenase and quickly becomes constrained by limited NADH availability, even when glycerol formation is abolished. We present alcohol dehydrogenase as a novel target for anaerobic redox engineering of S. cerevisiae. Introduction of an NADPH-specific alcohol dehydrogenase (NADPH-ADH) not only reduces the NADH demand of the acetate-to-ethanol pathway but also allows the cell to effectively exchange NADPH for NADH during sugar fermentation. Unlike NADH, NADPH can be freely generated under anoxic conditions, via the oxidative pentose phosphate pathway. We show that an industrial bioethanol strain engineered with the original pathway (expressing acetylating acetaldehyde dehydrogenase from Bifidobacterium adolescentis and with deletions of glycerol-3-phosphate dehydrogenase genes GPD1 and GPD2) consumed 1.9 g liter(-1) acetate during fermentation of 114 g liter(-1) glucose. Combined with a decrease in glycerol production from 4.0 to 0.1 g liter(-1), this increased the ethanol yield by 4% over that for the wild type. We provide evidence that acetate consumption in this strain is indeed limited by NADH availability. By introducing an NADPH-ADH from Entamoeba histolytica and with overexpression of ACS2 and ZWF1, we increased acetate consumption to 5.3 g liter(-1) and raised the ethanol yield to 7% above the wild-type level. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  4. Phosphoglycerate dehydrogenase (PHGDH) deficiency without epilepsy mimicking primary microcephaly.

    Science.gov (United States)

    Poli, Antoine; Vial, Yoann; Haye, Damien; Passemard, Sandrine; Schiff, Manuel; Nasser, Hala; Delanoe, Catherine; Cuadro, Emma; Kom, Rémi; Elanga, Narcisse; Favre, Anne; Drunat, Séverine; Verloes, Alain

    2017-04-25

    Phosphoglycerate dehydrogenase (PHGDH) deficiency (OMIM 256520) is a rare autosomal recessive disorder of serine synthesis, with mostly severe congenital microcephaly, caused by mutations in the PHGDH gene. Fourteen patients reported to date show severe, early onset, drug resistant epilepsy. In a cohort of patients referred for primary microcephaly, compound heterozygosity for two unreported variants in PHGDG was identified by exome sequencing in a pair of sibs who died aged 4.5 months and 4.5 years. They had severe neurological involvement with congenital microcephaly, disorganized EEG, and progressive spasticity, but never had seizures. Exome usage in clinical practice is likely to lead to an expansion of the clinical spectrum of known disorders. © 2017 Wiley Periodicals, Inc.

  5. Physiological Regulation of Isocitrate Dehydrogenase and the Role of 2-Oxoglutarate in Prochlorococcus sp. Strain PCC 9511

    Science.gov (United States)

    Diez, Jesús; Gómez-Baena, Guadalupe; Rangel-Zúñiga, Oriol Alberto; García-Fernández, José Manuel

    2014-01-01

    The enzyme isocitrate dehydrogenase (ICDH; EC 1.1.1.42) catalyzes the oxidative decarboxylation of isocitrate, to produce 2-oxoglutarate. The incompleteness of the tricarboxylic acids cycle in marine cyanobacteria confers a special importance to isocitrate dehydrogenase in the C/N balance, since 2-oxoglutarate can only be metabolized through the glutamine synthetase/glutamate synthase pathway. The physiological regulation of isocitrate dehydrogenase was studied in cultures of Prochlorococcus sp. strain PCC 9511, by measuring enzyme activity and concentration using the NADPH production assay and Western blotting, respectively. The enzyme activity showed little changes under nitrogen or phosphorus starvation, or upon addition of the inhibitors DCMU, DBMIB and MSX. Azaserine, an inhibitor of glutamate synthase, induced clear increases in the isocitrate dehydrogenase activity and icd gene expression after 24 h, and also in the 2-oxoglutarate concentration. Iron starvation had the most significant effect, inducing a complete loss of isocitrate dehydrogenase activity, possibly mediated by a process of oxidative inactivation, while its concentration was unaffected. Our results suggest that isocitrate dehydrogenase responds to changes in the intracellular concentration of 2-oxoglutarate and to the redox status of the cells in Prochlorococcus. PMID:25061751

  6. 3-Keto-5 alpha-steroid Delta'-dehydrogenase from Rhodococcus erythropolis SQ1 and its orthologue in Mycobacterium tuberculosis H37Rv are highly specific enzymes that function in cholesterol catabolism

    NARCIS (Netherlands)

    Knol, Jan; Bodewits, Karin; Hessels, Gerda I.; Dijkhuizen, Lubbert; Van der Geize, Robert

    2008-01-01

    The Rhodococcus erythropolis SQ1 kstD3 gene was cloned, heterologously expressed and biochemically characterized as a KSTD3 (3-keto-5 alpha-steroid Delta'-dehydrogenase). Upstream of kstD3, an ORF (open reading frame) with similarity to Delta(4) KSTD (3-keto-5 alpha-steroid Delta(4)-dehydrogenase)

  7. Genes involved in lactose catabolism and organic acid production during growth of Lactobacillus delbrueckii UFV H2b20 in skimmed milk.

    Science.gov (United States)

    Do Carmo, A P; De Oliveira, M N V; Da Silva, D F; Castro, S B; Borges, A C; De Carvalho, A F; De Moraes, C A

    2012-03-01

    There are three main reasons for using lactic acid bacteria (LAB) as starter cultures in industrial food fermentation processes: food preservation due to lactic acid production; flavour formation due to a range of organic molecules derived from sugar, lipid and protein catabolism; and probiotic properties attributed to some strains of LAB, mainly of lactobacilli. The aim of this study was to identify some genes involved in lactose metabolism of the probiotic Lactobacillus delbrueckii UFV H2b20, and analyse its organic acid production during growth in skimmed milk. The following genes were identified, encoding the respective enzymes: ldh - lactate dehydrogenase, adhE - Ldb1707 acetaldehyde dehydrogenase, and ccpA-pepR1 - catabolite control protein A. It was observed that L. delbrueckii UFV H2b20 cultivated in different media has the unexpected ability to catabolyse galactose, and to produce high amounts of succinic acid, which was absent in the beginning, raising doubts about the subspecies in question. The phylogenetic analyses showed that this strain can be compared physiologically to L. delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis, which are able to degrade lactose and can grow in milk. L. delbrueckii UFV H2b20 sequences have grouped with L. delbrueckii subsp. bulgaricus ATCC 11842 and L. delbrueckii subsp. bulgaricus ATCC BAA-365, strengthening the classification of this probiotic strain in the NCFM group proposed by a previous study. Additionally, L. delbrueckii UFV H2b20 presented an evolutionary pattern closer to that of probiotic Lactobacillus acidophilus NCFM, corroborating the suggestion that this strain might be considered as a new and unusual subspecies among L. delbrueckii subspecies, the first one identified as a probiotic. In addition, its unusual ability to metabolise galactose, which was significantly consumed in the fermentation medium, might be exploited to produce low-browning probiotic Mozzarella cheeses, a desirable property

  8. Molecular cloning and characterization of a novel Dehydrogenase/reductase (SDR family) member 1 genea from human fetal brain.

    Science.gov (United States)

    Wu, Q; Xu, M; Cheng, C; Zhou, Z; Huang, Y; Zhao, W; Zeng, L; Xu, J; Fu, X; Ying, K; Xie, Y; Mao, Y

    2001-01-01

    Short-chain dehydrogenases/reductases (SDR) constitute a large protein family of NAD(P)(H)-dependent oxidoreductase. They are defined by distinct, common sequence motifs and show a wide range of substrate specialisms. By large-scale sequencing analysis of a human fetal brain cDNA library, we isolated a novel human SDR-type dehydrogenase/reductase gene named Dehydrogenase/reductase (SDR family) member 1 (DHRS1). The DHRS1 cDNA is 1411 base pair in length, encoding a 314-amino-acid polypeptide which has a SDR motif. Northern blot reveals two bands, of about 0.9 and 1.4 kb in size. These two forms are expressed in many tissues. The DHRS1 gene is localized on chromosome 14q21.3. It has 9 exons and spans 9.2 kb of the genomic DNA.

  9. Monoterpene Metabolism. Cloning, Expression, and Characterization of (−)-Isopiperitenol/(−)-Carveol Dehydrogenase of Peppermint and Spearmint1

    Science.gov (United States)

    Ringer, Kerry L.; Davis, Edward M.; Croteau, Rodney

    2005-01-01

    The essential oils of peppermint (Mentha x piperita) and spearmint (Mentha spicata) are distinguished by the oxygenation position on the p-menthane ring of the constitutive monoterpenes that is conferred by two regiospecific cytochrome P450 limonene-3- and limonene-6-hydroxylases. Following hydroxylation of limonene, an apparently similar dehydrogenase oxidizes (−)-trans-isopiperitenol to (−)-isopiperitenone in peppermint and (−)-trans-carveol to (−)-carvone in spearmint. Random sequencing of a peppermint oil gland secretory cell cDNA library revealed a large number of clones that specified redox-type enzymes, including dehydrogenases. Full-length dehydrogenase clones were screened by functional expression in Escherichia coli using a recently developed in situ assay. A single full-length acquisition encoding (−)-trans-isopiperitenol dehydrogenase (ISPD) was isolated. The (−)-ISPD cDNA has an open reading frame of 795 bp that encodes a 265-residue enzyme with a calculated molecular mass of 27,191. Nondegenerate primers were designed based on the (−)-trans-ISPD cDNA sequence and employed to screen a spearmint oil gland secretory cell cDNA library from which a 5′-truncated cDNA encoding the spearmint homolog, (−)-trans-carveol-dehydrogenase, was isolated. Reverse transcription-PCR amplification and RACE were used to acquire the remaining 5′-sequence from RNA isolated from oil gland secretory cells of spearmint leaf. The full-length spearmint dehydrogenase shares >99% amino acid identity with its peppermint homolog and both dehydrogenases are capable of utilizing (−)-trans-isopiperitenol and (−)-trans-carveol. These isopiperitenol/carveol dehydrogenases are members of the short-chain dehydrogenase/reductase superfamily and are related to other plant short-chain dehydrogenases/reductases involved in secondary metabolism (lignan biosynthesis), stress responses, and phytosteroid biosynthesis, but they are quite dissimilar (approximately 13

  10. Precipitation of lysozyme with sodium succinate, sodium tartrate and sodium citrate: Solubility and osmotic second virial coefficient data

    International Nuclear Information System (INIS)

    López Vélez, José Sebastián; Azzoni, Adriano Rodrigues; Pessoa Filho, Pedro de Alcantara

    2017-01-01

    Highlights: • The solubility of lysozyme in biodegradable salt solutions was investigated. • Studied biodegradable salts: sodium succinate, sodium tartrate and sodium citrate. • Sodium succinate and sodium tartrate are promising salting-out agents. • Osmotic second virial coefficient (B 22 ) of lysozyme was also investigated. • Values of B 22 indicate that crystallization may be achieved using these salts. - Abstract: Precipitation and crystallization are unit operations widely used to concentrate and purify proteins in biotechnology industry. In this work, the potential use of the biodegradable salts sodium succinate, sodium tartrate and sodium citrate to reduce the solubility of proteins is investigated. Lysozyme was studied as a model protein. The solubility of lysozyme in aqueous solutions of sodium succinate, sodium tartrate and sodium citrate was experimentally determined as a function of the ionic strength and pH at 298.2 K. All these salts induce the precipitation of lysozyme, being sodium succinate the most effective one to reduce the solubility of this protein. The osmotic second virial coefficient of lysozyme as a function of the ionic strength was also determined using self-interaction chromatography at 298.2 K and pH 8.5. The experimental data show that the value of the second virial coefficient is negative at the investigated conditions and lies mostly within or close to the crystallization slot, i.e., the range of values of this coefficient for which the formation of crystals is favored.

  11. Identification of Glyceraldehyde 3-Phosphate Dehydrogenase Sequence and Expression Profiles in Tree Shrew (Tupaia belangeri)

    OpenAIRE

    Zheng, Yu; Wang, Qihui; Yun, Chenxia; Wang, Yingjun; Smith, Wanli W.; Leng, Jing

    2014-01-01

    The tree shrews (Tupaia belangeri) diverged from the primate order (Primates) and are classified as Scandentia, a separate taxonomic group of mammals. The tree shrew has been suggested to use an animal model to study human disease but the genomic sequences of tree shrew is largely unidentified. Here we identified the full-length cDNA sequence of a housekeeping gene, Glyceraldehyde 3-phosphate Dehydrogenase (GAPDH), in tree shrew. We further constructed a phylogenetic family tree base on GAPDH...

  12. Alcohol drinking habits, alcohol dehydrogenase genotypes and risk of acute coronary syndrome

    DEFF Research Database (Denmark)

    Tolstrup, J.S.; Hansen, J.L.; Gronbaek, M.

    2010-01-01

    Aims: The risk of myocardial infarction is lower among light-to-moderate drinkers compared with abstainers. Results from some previous studies, but not all, suggest that this association is modified by variations in genes coding for alcohol dehydrogenase (ADH). We aimed to test this hypothesis, i...... for the association between alcohol drinking habits and the risk of developing acute coronary syndrome, if any, is very limited....

  13. Multiple soluble malate dehydrogenase of Geophagus brasiliensis (Cichlidae, Perciformes

    Directory of Open Access Journals (Sweden)

    Aquino-Silva Maria Regina de

    1998-01-01

    Full Text Available A recent locus duplication hypothesis for sMDH-B* was proposed to explain the complex electrophoretic pattern of six bands detected for the soluble form of malate dehydrogenase (MDH, EC 1.1.1.37 in 84% of the Geophagus brasiliensis (Cichlidae, Perciformes analyzed (AB1B2 individuals. Klebe's serial dilutions were carried out in skeletal muscle extracts. B1 and B2 subunits had the same visual end-points, reflecting a nondivergent pattern for these B-duplicated genes. Since there is no evidence of polyploidy in the Cichlidae family, MDH-B* loci must have evolved from regional gene duplication. Tissue specificities, thermostability and kinetic tests resulted in similar responses from both B-isoforms, in both sMDH phenotypes, suggesting that these more recently duplicated loci underwent the same regulatory gene action. Similar results obtained with the two sMDH phenotypes did not show any indication of a six-banded specimen adaptive advantage in subtropical regions.

  14. Mitochondrially Targeted Vitamin E Succinate Modulates Expression of Mitochondrial DNA Transcripts and Mitochondrial Biogenesis

    Czech Academy of Sciences Publication Activity Database

    Truksa, Jaroslav; Dong, L.F.; Rohlena, Jakub; Stursa, J.; Vondrusová, Magdaléna; Goodwin, J.; Nguyen, M.; Klučková, Katarína; Rychtarčíková, Zuzana; Lettlová, Sandra; Spáčilová, Jana; Stapelberg, M.; Zoratti, M.; Neužil, Jiří

    2015-01-01

    Roč. 22, č. 11 (2015), s. 883-900 ISSN 1523-0864 R&D Projects: GA ČR GAP301/10/1937; GA AV ČR KAN200520703; GA ČR GAP305/12/1708; GA ČR GAP301/12/1851; GA MŠk(CZ) ED1.1.00/02.0109 Institutional research plan: CEZ:AV0Z50520701 Institutional support: RVO:86652036 Keywords : ALPHA -TOCOPHERYL SUCCINATE * ELECTRON-TRANSPORT CHAIN * RESPIRATORY COMPLEX-II Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.093, year: 2015

  15. Heterogeneous catalytic hydrogenation of biobased levulinic and succinic acids in aqueous solutions.

    Science.gov (United States)

    Corbel-Demailly, Louis; Ly, Bao-Khanh; Minh, Doan-Pham; Tapin, Benoit; Especel, Catherine; Epron, Florence; Cabiac, Amandine; Guillon, Emmanuelle; Besson, Michèle; Pinel, Catherine

    2013-12-01

    Supported noble-metal catalysts (Ru, Pd or Pt) and the corresponding Re-promoted catalysts exhibit a high activity for the hydrogenation of biobased carboxylic acids. Levulinic acid and succinic acid are converted into the lactones or the diols depending on the nature of the catalyst and the reaction conditions. The highest selectivity to 1,4-pentanediol of 82 % is achieved at 140 °C in the presence of the 1.9 % Ru-3.6 % Re/C catalyst. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. O-Alkyl Hydroxamates as Metaphors of Enzyme-Bound Enolate Intermediates in Hydroxy Acid Dehydrogenases. Inhibitors of Isopropylmalate Dehydrogenase, Isocitrate Dehydrogenase, and Tartrate Dehydrogenase(1).

    Science.gov (United States)

    Pirrung, Michael C.; Han, Hyunsoo; Chen, Jrlung

    1996-07-12

    The inhibition of Thermus thermophilus isopropylmalate dehydrogenase by O-methyl oxalohydroxamate was studied for comparison to earlier results of Schloss with the Salmonella enzyme. It is a fairly potent (1.2 &mgr;M), slow-binding, uncompetitive inhibitor against isopropylmalate and is far superior to an oxamide (25 mM K(i) competitive) that is isosteric with the ketoisocaproate product of the enzyme. This improvement in inhibition was attributed to its increased NH acidity, which presumably is due to the inductive effect of the hydroxylamine oxygen. This principle was extended to the structurally homologous enzyme isocitrate dehydrogenase from E. coli, for which the compound O-(carboxymethyl) oxalohydroxamate is a 30 nM inhibitor, uncompetitive against isocitrate. The pH dependence of its inhibition supports the idea that it is bound to the enzyme in the anionic form. Another recently discovered homologous enzyme, tartrate dehydrogenase from Pseudomonas putida, was studied with oxalylhydroxamate. It has a relatively low affinity for the enzyme, though it is superior to tartrate. On the basis of these leads, squaric hydroxamates with increased acidity compared to squaric amides directed toward two of these enzymes were prepared, and they also show increased inhibitory potency, though not approaching the nanomolar levels of the oxalylhydroxamates.

  17. Toxic response caused by a misfolding variant of the mitochondrial protein short-chain acyl-CoA dehydrogenase

    DEFF Research Database (Denmark)

    Schmidt, Stinne P; Corydon, Thomas J; Pedersen, Christina B

    2011-01-01

    BACKGROUND: Variations in the gene ACADS, encoding the mitochondrial protein short-chain acyl CoA-dehydrogenase (SCAD), have been observed in individuals with clinical symptoms. The phenotype of SCAD deficiency (SCADD) is very heterogeneous, ranging from asymptomatic to severe, without a clear ge...

  18. Toxic response caused by a misfolding variant of the mitochondrial protein short-chain acyl-CoA dehydrogenase

    NARCIS (Netherlands)

    Schmidt, S.P.; Corydon, T.J.; Pedersen, C.B.; Vang, S.; Palmfeldt, J.; Stenbroen, V.; Wanders, R.J.A.; Ruiter, J.P.N.; Gregersen, N.

    2011-01-01

    Variations in the gene ACADS, encoding the mitochondrial protein short-chain acyl CoA-dehydrogenase (SCAD), have been observed in individuals with clinical symptoms. The phenotype of SCAD deficiency (SCADD) is very heterogeneous, ranging from asymptomatic to severe, without a clear

  19. In vitro and in vivo consequences of variant medium-chain acyl-CoA dehydrogenase genotypes

    NARCIS (Netherlands)

    Touw, Catharina M. L.; Smit, G. Peter A.; Niezen-Koning, Klary E.; Bosgraaf-de Boer, Conny; Gerding, Albert; Reijngoud, Dirk-Jan; Derks, Terry

    2013-01-01

    Background: Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most common inherited disorder of the mitochondrial fatty acid oxidation, caused by mutations in the ACADM gene. Since the introduction of neonatal screening for MCAD deficiency, a subgroup of newborns have been identified with

  20. Changing flux of xylose metabolites by altering expression of xylose reductase and xylitol dehydrogenase in recombinant Saccharomyces cerevisiae

    Science.gov (United States)

    Yong-Su Jin; Thomas W. Jeffries

    2003-01-01

    We changed the fluxes of xylose metabolites in recombinant Saccharomyces cerevisiae by manipulating expression of Pichia stipitis genes(XYL1 and XYL2) coding for xylose reductase (XR) and xylitol dehydrogenase (XDH), respectively. XYL1 copy number was kept constant by integrating it into the chromosome. Copy numbers of XYL2 were varied either by integrating XYL2 into...

  1. The validation of housekeeping genes as a reference in quantitative Real Time PCR analysis: application in the milk somatic cells and frozen whole blood of goats infected with caprine arthritis encephalitis virus.

    Science.gov (United States)

    Jarczak, Justyna; Kaba, Jarosław; Bagnicka, Emilia

    2014-10-10

    The validation of housekeeping genes (HKGs) for normalization of RNA expression in Real-Time PCR is crucial to obtain the most reliable results. There is limited information on reference genes used in the study of gene expression in milk somatic cells and the frozen whole blood of goats. Thus, the aim of this study was to propose the most stable housekeeping genes that can be used as a reference in Real-Time PCR analysis of milk somatic cells and whole blood of goats infected with caprine arthritis encephalitis virus (CAEV). Animals were divided into two groups: non-infected (N=13) and infected with CAEV (N=13). Biological material (milk somatic cells and whole blood) was collected 4 times during the lactation period (7, 30, 100 and 240days post-partum). The expression levels of candidate reference genes were analyzed using geNorm and NormFinder software. The stability of candidates for reference gene expression was analyzed for CAEV-free (control) and CAEV-infected groups, and also for both groups together (combined group). The stability of expression of β-actin (ACTB), glyceraldehyde-3P-dehydrogenase (GAPDH), cyclophilin A (PPIA), RNA18S1, ubiquilin (UBQLN1) and ribosomal protein large subunit P0 (RPLP0) was determined in milk somatic cells, while ACTB, PPIA, RPLP0, succinate dehydrogenase complex subunit A (SDHA), zeta polypeptide (YWHAZ), battenin (CLN3), eukaryotic translation initiation factor 3K (EIF3K) and TATA box-binding protein (TBP) were measured in frozen whole blood of goats. PPIA and RPLP0 were considered as the most suitable internal controls as they were stably expressed in milk somatic cells regardless of disease status, according to NormFinder software. Furthermore, geNorm results indicated the expression of PPIA/RPLP0 genes as the best combination under these experimental conditions. The results of frozen whole blood analysis using NormFinder software revealed that the most stable reference gene in control, CAEV-infected and combined groups is

  2. Nb-Based Zeolites: Efficient bi-Functional Catalysts for the One-Pot Synthesis of Succinic Acid from Glucose

    Directory of Open Access Journals (Sweden)

    Magdi El Fergani

    2017-12-01

    Full Text Available The one-pot production of succinic acid from glucose was investigated in pure hot water as solvent using Nb (0.02 and 0.05 moles%-Beta zeolites obtained by a post-synthesis methodology. Structurally, they are comprised of residual framework Al-acid sites, extra-framework isolated Nb (V and Nb2O5 pore-encapsulated clusters. The Nb-modified Beta-zeolites acted as bi-functional catalysts in which glucose is dehydrated to levulinic acid (LA which, further, suffers an oxidation process to succinic acid (SA. After the optimization of the reaction conditions, that is, at 180 °C, 18 bar O2, and 12 h reaction time, the oxidation of glucose occurred with a selectivity to succinic acid as high as 84% for a total conversion.

  3. Extraction and separation of mercury(II from succinate media with high molecular weight amine as an extractant

    Directory of Open Access Journals (Sweden)

    Chandrashekhar P. Mane

    2015-01-01

    Full Text Available The extraction of mercury(II from succinate solutions by 2-octylaminopyridine (2-OAP has been studied by metal distribution measurements. The optimum extraction conditions were determined from a critical study of effects of pH, sodium succinate concentration and 2-octylaminopyridine concentration. The probable composition of the species has been deduced from log–log plots. The extraction reaction proceeds with ion-pair formation and the stoichiometry of extracted species was found to be [(2OAPH+2 Hg(Succinate22−](org. The method has been used to separate mercury(II from commonly associated elements. The method has been further extended to separate and to estimate mercury in synthetic mixtures and real samples.

  4. Over-Expression, Purification and Crystallization of Human Dihydrolipoamide Dehydrogenase

    Science.gov (United States)

    Hong, Y. S.; Ciszak, Ewa; Patel, Mulchand

    2000-01-01

    Dehydrolipoamide dehydrogenase (E3; dihydrolipoan-tide:NAD+ oxidoreductase, EC 1.8.1.4) is a common catalytic component found in pyruvate dehydrogenase complex, alpha-ketoglutarate dehydrogenase complex, and branched-chain cc-keto acid dehydrogenase complex. E3 is also a component (referred to as L protein) of the glycine cleavage system in bacterial metabolism (2). Active E3 forms a homodimer with four distinctive subdomain structures (FAD binding, NAD+ binding, central and interface domains) with non-covalently but tightly bound FAD in the holoenzyme. Deduced amino acids from cloned full-length human E3 gene showed a total of 509 amino acids with a leader sequence (N-terminal 35 amino acids) that is excised (mature form) during transportation of expressed E3 into mitochondria membrane. So far, three-dimensional structure of human E3 has not been reported. Our effort to achieve the elucidation of the X-ray crystal structure of human E3 will be presented. Recombinant pPROEX-1 expression vector (from GIBCO BRL Life Technologies) having the human E3 gene without leader sequence was constructed by Polymerase Chain Reaction (PCR) and subsequent ligation, and cloned in E.coli XL1-Blue by transformation. Since pPROEX-1 vector has an internal His-tag (six histidine peptide) located at the upstream region of a multicloning site, one-step affinity purification of E3 using nickelnitriloacetic acid (Ni-NTA) agarose resin, which has a strong affinity to His-tag, was feasible. Also a seven-amino-acid spacer peptide and a recombinant tobacco etch virus protease recognition site (seven amino acids peptide) found between His-tag and first amino acid of expressed E3 facilitated the cleavage of His-tag from E3 after the affinity purification. By IPTG induction, ca. 15 mg of human E3 (mature form) was obtained from 1L LB culture with overnight incubation at 25C. Over 98% of purity of E3 from one-step Ni-NTA agarose affinity purification was confirmed by SDS-PAGE analysis. For

  5. Dynamic changes in the gene expression profile during rat oral carcinogenesis induced by 4-nitroquinoline 1-oxide.

    Science.gov (United States)

    Ge, Shuyun; Zhang, Ji; Du, Yanzhi; Hu, Bin; Zhou, Zengtong; Lou, Jianing

    2016-03-01

    The typical progression of oral cancer is from hyperplastic epithelial lesions through dysplasia to invasive carcinoma. It is important to investigate malignant oral cancer progression and development in order to determine useful approaches of prevention of dysplastic lesions. The present study aimed to gain insights into the underlying molecular mechanism of oral carcinogenesis by establishing a rat model of oral carcinogenesis using 4‑nitroquinoline 1‑oxide. Subsequently, transcription profile analysis using an integrating microarray was performed. The dynamic gene expression changes of the six stages of rat oral carcinogenesis (normal, mild epithelial dysplasia, moderate dysplasia, severe dysplasia, carcinoma in situ and oral squamous cell carcinomas) were analyzed using component plane presentations (CPP)‑self‑organizing map (SOM). Six genes were verified by quantitative polymerase chain reaction, immunohistochemistry and succinate dehydrogenase (SDH) activity assay kit. Numerous differentially expressed genes (DEGs) were identified during rat oral carcinogenesis. CPP‑SOM determined that these DEGs were primarily enriched during cell cycle, apoptosis, inflammatory response and tricarboxylic acid cycle, indicating the coordinated regulation of molecular networks. In addition, the expression of specific DEGs, such as janus kinase 3, cyclin‑dependent kinase A‑1, B‑cell chronic lymphocytic leukaemia/lymphoma 2‑like 2, nuclear factor‑κB, tumor necrosis factor receptor superfamily member 1A, cyclin D1 and SDH were identified to have high concordance with the results from microarray data. The current study demonstrated that oral carcinogenesis is a multi‑step and multi‑gene process, with a distinct pattern alteration along a continuum of malignant transformation. In addition, this comprehensive investigation provided a theoretical basis for the understanding of the molecular alterations associated with oral carcinogenesis.

  6. Study on fluorescence properties of carbogenic nanoparticles and their application for the determination of ferrous succinate

    Energy Technology Data Exchange (ETDEWEB)

    Sun Wen [Department of Analytical Chemistry, China Pharmaceutical University, Nanjing 210009 (China); Du Yingxiang, E-mail: du_yingxiang@126.co [Department of Analytical Chemistry, China Pharmaceutical University, Nanjing 210009 (China) and Key Laboratory of Drug Quality Control and Pharmacovigilance, Ministry of Education, China Pharmaceutical University, Nanjing 210009 (China) and Key Laboratory of Modern Chinese Medicines, Ministry of Education, China Pharmaceutical University, Nanjing 210009 (China); Wang Yunqing [Yantai Institute of Coastal Zone Research, Chinese Academy of Sciences, Yantai 264003 (China)

    2010-08-15

    A new type of fluorescent nanomaterial named carbogenic nanoparticles (NPs) has drawn considerable attention recently. In this study, we adopted a direct and simple synthetic method to produce the carbogenic NPs and investigated the fluorescence properties of the as-prepared carbogenic NPs in detail. It was found that the fluorescence of carbogenic NPs was stable with the variance of environmental conditions such as pH, temperature and UV irradiation. More interestingly, we found carbogenic NPs exhibited high selectivity and sensitivity towards ferric ions. Under optimum conditions, a good linear relationship could be obtained between the fluorescence intensity and concentration of ferric ions in the range of 5.0x10{sup -5}-5.0x10{sup -4} mol L{sup -1}, and the limit of detection is 11.2 {mu}mol L{sup -1}. Based on the fluorescence quenching of carbogenic NPs, a rapid and specific quantitative method was proposed for the determination of ferrous succinate. The content of ferrous succinate in commercial tablets determined by the present method was agreed with the spectrophotometric method results and the reproducibility and the recovery of the proposed method were satisfactory.

  7. Crystallization and melting behavior of poly (ethylene succinate) in presence of graphene nanoplatelets

    Energy Technology Data Exchange (ETDEWEB)

    Asadinezhad, Ahmad [Department of Chemical Engineering, Isfahan University of Technology, 84156-83111 Isfahan (Iran, Islamic Republic of); Khonakdar, Hossein Ali, E-mail: hakhonakdar@gmail.com [Iran Polymer and Petrochemical Institute, P.O. Box 14965/115, Tehran (Iran, Islamic Republic of); Leibniz-Institut für Polymerforschung Dresden e.V., D-01069 Dresden (Germany); Häußler, Liane; Wagenknecht, Udo [Leibniz-Institut für Polymerforschung Dresden e.V., D-01069 Dresden (Germany); Heinrich, Gert [Leibniz-Institut für Polymerforschung Dresden e.V., D-01069 Dresden (Germany); Technische Universität Dresden, Institut für Werkstoffwissenschaft, D-01069 Dresden (Germany)

    2014-06-01

    Highlights: • Poly (ethylene succinate)/graphene nanocomposites were characterized in this work. • Dynamic and isothermal crystallization rates were enhanced upon graphene addition. • Overall crystallinity and melting remained almost unchanged in presence of graphene. • Graphene changed crystal perfection and its effect was found concentration-dependent. - Abstract: In this study, poly (ethylene succinate)/graphene nanoplatelets composites were prepared via solution casting method in different compositions and analyzed via differential scanning calorimetry. Crystallization and melting characteristics in both dynamic (non-isothermal) and isothermal modes were investigated at various cooling rates and isothermal temperatures. It was confirmed that graphene, while being incompatible with the matrix polymer, could act as nucleating agent so that hot (melt) crystallization rates, intensities, and positions were increased in the dynamic mode. However, the overall crystallinity remained almost unchanged. Cold crystallization was also decreased to lower temperatures on graphene addition; however its enthalpy was also reduced. Similar findings were observed for the isothermal mode, but to a lesser extent. The presence of graphene changed the perfection of crystals as their thermodynamic stability against heating and recrystallization behavior varied.

  8. Maleic acid and succinic acid in fermented alcoholic beverages are the stimulants of gastric acid secretion.

    Science.gov (United States)

    Teyssen, S; González-Calero, G; Schimiczek, M; Singer, M V

    1999-03-01

    Alcoholic beverages produced by fermentation (e.g., beer and wine) are powerful stimulants of gastric acid output and gastrin release in humans. The aim of this study was to separate and specify the gastric acid stimulatory ingredients in alcoholic beverages produced by fermentation. Yeast-fermented glucose was used as a simple model of fermented alcoholic beverages; it was stepwise separated by different methods of liquid chromatography, and each separated solution was tested in human volunteers for its stimulatory action on gastric acid output and gastrin release. Five substances were detected by high-performance liquid chromatography and were analyzed by mass spectrometry and 1H-13C nuclear magnetic resonance spectroscopy. At the end of the separation process of the five identified substances, only the two dicarboxylic acids, maleic acid and succinic acid, had a significant (P fermented glucose, respectively), but not on gastrin release. When given together, they increased gastric acid output by 100% of fermented glucose and by 95% of maximal acid output. We therefore conclude that maleic acid and succinic acid are the powerful stimulants of gastric acid output in fermented glucose and alcoholic beverages produced by fermentation, and that gastrin is not their mediator of action.

  9. Succinate Functionalization of Hyperbranched Polyglycerol-Coated Magnetic Nanoparticles as a Draw Solute During Forward Osmosis.

    Science.gov (United States)

    Yang, Hee-Man; Choi, Hye Min; Jang, Sung-Chan; Han, Myeong Jin; Seo, Bum-Kyoung; Moon, Jei-Kwon; Lee, Kune-Woo

    2015-10-01

    Hyperbranched polyglycerol-coated magnetic nanoparticles (SHPG-MNPs) were functionalized with succinate groups to form a draw solute for use in a forward osmosis (FO). After the one-step synthesis of hyperbranched polyglycerol-coated magnetic nanoparticles (HPG-MNPs), the polyglycerol groups on the surfaces of the HPG-MNPs were functionalized with succinic anhydride moieties. The resulting SHPG-MNPs showed no change of size and magnetic property compared with HPG-MNPs and displayed excellent dispersibility in water up to the concentration of 400 g/L. SHPG-MNPs solution showed higher osmotic pressure than that of HPG-MNPs solution due to the presence of surface carboxyl groups in SHPG-MNPs and could draw water from a feed solution across an FO membrane without any reverse draw solute leakage during FO process. Moreover, the water flux remained nearly constant over several SHPG-MNP darw solute regeneration cycles applied to the ultrafiltration (UF) process. The SHPG-MNPs demonstrate strong potential for use as a draw solute in FO processes.

  10. Crystallization and degradation behaviors of poly(butylene succinate)/poly(Z-L-lysine) composites

    International Nuclear Information System (INIS)

    Tan, Licheng; Hu, Jun; Ye, Suwen; Wei, Junchao; Chen, Yiwang

    2014-01-01

    Highlights: • A new biodegradable poly(butylene succinate) (PBS)/poly(Z-L-lysine) (PZlys) composites were successfully prepared through physical blend. • PZlys may greatly affected the crystallization behaviors of PBS without changing its crystalline structure. • The degradation speed of PBS may be greatly accelerated by introduction of PZlys in PBS matrix. - Abstract: A new type of biodegradable poly(butylene succinate) (PBS)/poly(Z-L-lysine) (PZlys) composites were prepared. The crystallization behaviors were investigated by differential scanning calorimetry (DSC), wide angle X-ray diffraction (WAXD) and polarizing optical microscopy (POM) and the results showed that PZlys can restrict the crystallization of PBS, the crystallization speed of PBS/PZlys were slower than that of PBS, and the crystallization degree of the composites were smaller than that of PBS. However, the WAXD results showed that the incorporation of PZlys did not change the crystalline structure of PBS. The in vitro degradation experiments demonstrated that the degradation speed of the composites were faster than that of PBS. Moreover, the mechanical properties of the composites showed that the composites with a proper composition (for example, 80/20) can keep the mechanical properties of PBS without evident difference, which implied that the composites might be potentially useful as biodegradable materials

  11. Variation in gastric alcohol dehydrogenase and the risk of alcohol dependence

    Directory of Open Access Journals (Sweden)

    Paulina Całka

    2017-03-01

    Full Text Available Alcohol dependence is both a medical and socioeconomic problem. The disease is multifactorial, i.e. its development is attributable to gene-gene and gene-environment interactions. Multi-centre studies investigating the genetic background of alcoholism stress the role of genes encoding enzymes of the ethanol decomposition pathway in the human body, particularly alcohol dehydrogenase (ADH, in the development of alcohol dependence. Among five classes of alcohol dehydrogenases, class I and IV isoenzymes have been found to be associated with alcohol dependence. Class IV is of particular interest due to its occurrence in the upper gastrointestinal tract, mainly in the stomach. No activity of the enzyme has been demonstrated in the liver. Single nucleotide polymorphism (SNP of the gene encoding ADH class IV (ADH7 affects its ethanol-oxidizing activity in the gastric lumen, thereby influencing the first-pass metabolism (FPM of the substance. The findings published by various research centres have demonstrated that specific SNP changes in the ADH7 gene are of different significance for the risk of alcohol dependence according to the population studied.

  12. Rapid screening for glucose-6-phosphate dehydrogenase deficiency and haemoglobin polymorphisms in Africa by a simple high-throughput SSOP-ELISA method

    DEFF Research Database (Denmark)

    Enevold, Anders; Vestergaard, Lasse S; Lusingu, John

    2005-01-01

    BACKGROUND: Mutations in the haemoglobin beta-globin (HbB) and glucose-6-phosphate dehydrogenase (G6PD) genes cause widespread human genetic disorders such as sickle cell diseases and G6PD deficiency. In sub-Saharan Africa, a few predominant polymorphic variants of each gene account for a majority...

  13. Flavin Adenine Dinucleotide Status and the Effects of High-Dose Riboflavin Treatment in Short-Chain Acyl-CoA Dehydrogenase Deficiency

    NARCIS (Netherlands)

    van Maldegem, Bianca T.; Duran, Marinus; Wanders, Ronald J. A.; Waterham, Hans R.; Wijburg, Frits A.

    2010-01-01

    Short-chain acyl-CoA dehydrogenase deficiency (SCADD) is an inborn error, biochemically characterized by increased plasma butyrylcarnitine (C4-C) concentration and increased ethylmalonic acid (EMA) excretion and caused by rare mutations and/or common gene variants in the SCAD encoding gene. Although

  14. The mitochondrial FAD-dependent glycerol-3-phosphate dehydrogenase of Trypanosomatidae and the glycocomal redox balance of insect stages of Trypanosoma brucei and Leishmania spp.

    NARCIS (Netherlands)

    Guerra, D.G.; Decottignies, A.; Bakker, B.M.; Michels, P.A.M.

    2006-01-01

    The genes for the mitochondrial FAD-dependent glycerol-3-phosphate dehydrogenase were identified in Trypanosoma brucei and Leishmania major genomes. We have expressed the L. major gene in Saccharomyces cerevisiae and confirmed the subcellular localization and activity of the produced enzyme. Using

  15. Hexose-6-phosphate dehydrogenase contributes to skeletal muscle homeostasis independent of 11β-hydroxysteroid dehydrogenase type 1.

    LENUS (Irish Health Repository)

    Semjonous, Nina M

    2011-01-01

    Glucose-6-phosphate (G6P) metabolism by the enzyme hexose-6-phosphate dehydrogenase (H6PDH) within the sarcoplasmic reticulum lumen generates nicotinamide adenine dinucleotide phosphate (reduced) to provide the redox potential for the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) to activate glucocorticoid (GC). H6PDH knockout (KO) mice have a switch in 11β-HSD1 activity, resulting in GC inactivation and hypothalamic-pituitary-adrenal axis activation. Importantly, H6PDHKO mice develop a type II fiber myopathy with abnormalities in glucose metabolism and activation of the unfolded protein response (UPR). GCs play important roles in muscle physiology, and therefore, we have examined the importance of 11β-HSD1 and GC metabolism in mediating aspects of the H6PDHKO myopathy. To achieve this, we examined 11β-HSD1\\/H6PDH double-KO (DKO) mice, in which 11β-HSD1 mediated GC inactivation is negated. In contrast to H6PDHKO mice, DKO mice GC metabolism and hypothalamic-pituitary-adrenal axis set point is similar to that observed in 11β-HSD1KO mice. Critically, in contrast to 11β-HSD1KO mice, DKO mice phenocopy the salient features of the H6PDHKO, displaying reduced body mass, muscle atrophy, and vacuolation of type II fiber-rich muscle, fasting hypoglycemia, increased muscle glycogen deposition, and elevated expression of UPR genes. We propose that muscle G6P metabolism through H6PDH may be as important as changes in the redox environment when considering the mechanism underlying the activation of the UPR and the ensuing myopathy in H6PDHKO and DKO mice. These data are consistent with an 11β-HSD1-independent function for H6PDH in which sarcoplasmic reticulum G6P metabolism and nicotinamide adenine dinucleotide phosphate-(oxidized)\\/nicotinamide adenine dinucleotide phosphate (reduced) redox status are important for maintaining muscle homeostasis.

  16. NADPH-dependent glutamate dehydrogenase in Penicillium chrysogenum is involved in regulation of beta-lactam production

    DEFF Research Database (Denmark)

    Thykær, Jette; Kildegaard, Kanchana Rueksomtawin; Noorman, H.

    2008-01-01

    The interactions between the ammonium assimilatory pathways and beta-lactam production were investigated by disruption of the NADPH-dependent glutamate dehydrogenase gene (gdhA) in two industrial beta-lactam-producing strains of Penicillium chrysogenum. The strains used were an adipoyl-7-ADCA...... continued beta-lactam production for the reference strains whereas the Delta gdhA strains remained non-productive under all conditions. By overexpressing the NAD-dependent glutamate dehydrogenase, the specific growth rate could be restored, but still no beta-lactam production was detected. The results...... indicate that the NADPH-dependent glutamate dehydrogenase may be directly or indirectly involved in the regulation of beta-lactann production in industrial strains of P. chrysogenum....

  17. Purification, crystallization and preliminary X-ray analysis of isocitrate dehydrogenase kinase/phosphatase from Escherichia coli

    International Nuclear Information System (INIS)

    Zheng, Jimin; Lee, Daniel C.; Jia, Zongchao

    2009-01-01

    Isocitrate dehydrogenase kinase/phosphatase has been crystallized in three different crystal forms. Data were collected from each crystal form for structure determination. The Escherichia coli aceK gene encodes isocitrate dehydrogenase kinase/phosphatase (EC 2.7.11.5), a bifunctional protein that phosphorylates and dephosphorylates isocitrate dehydrogenase (IDH), resulting in its inactivation and activation, respectively. This reversible (de)phosphorylation directs isocitrate, an intermediate of the citric acid cycle, to either go through the full cycle or to enter the glyoxylate bypass. In the present study, the AceK protein from E. coli has been purified and crystallized. Three crystal forms were obtained from very similar crystallization conditions. The crystals belong to space groups P4 1 2 1 2, P3 2 21 and P2 1 2 1 2 1 and diffracted X-rays to resolutions of 2.9, 3.0 and 2.7 Å, respectively

  18. XoxF Is Required for Expression of Methanol Dehydrogenase in Methylobacterium extorquens AM1 ▿

    Science.gov (United States)

    Skovran, Elizabeth; Palmer, Alexander D.; Rountree, Austin M.; Good, Nathan M.; Lidstrom, Mary E.

    2011-01-01

    In Gram-negative methylotrophic bacteria, the first step in methylotrophic growth is the oxidation of methanol to formaldehyde in the periplasm by methanol dehydrogenase. In most organisms studied to date, this enzyme consists of the MxaF and MxaI proteins, which make up the large and small subunits of this heterotetrameric enzyme. The Methylobacterium extorquens AM1 genome contains two homologs of MxaF, XoxF1 and XoxF2, which are ∼50% identical to MxaF and ∼90% identical to each other. It was previously reported that xoxF is not required for methanol growth in M. extorquens AM1, but here we show that when both xoxF homologs are absent, strains are unable to grow in methanol medium and lack methanol dehydrogenase activity. We demonstrate that these defects result from the loss of gene expression from the mxa promoter and suggest that XoxF is part of a complex regulatory cascade involving the 2-component systems MxcQE and MxbDM, which are required for the expression of the methanol dehydrogenase genes. PMID:21873495

  19. Development of a solid-phase extraction method for simultaneous extraction of adipic acid, succinic acid and 1,4-butanediol formed during hydrolysis of poly(butylene adipate) and poly(butylene succinate).

    Science.gov (United States)

    Lindström, Annika; Albertsson, Ann-Christine; Hakkarainen, Minna

    2004-01-02

    A solid-phase extraction (SPE) method was developed for the simultaneous extraction of dicarboxylic acids and diols formed during hydrolysis of poly(butylene succinate), PBS, and poly(butylene adipate), PBA. Four commercial non-polar SPE columns, three silica based: C8, C18, C18 (EC), and one resin based: ENV+, were tested for the extraction of succinic acid, adipic acid and 1,4-butanediol, the expected final hydrolysis products of PBS and PBA. ENV+ resin was chosen as a solid-phase, because it displayed the best extraction efficiency for 1,4-butanediol and succinic acid. Linear range for the extracted analytes was 1-500 ng/microl for adipic acid and 2-500 ng/microl for 1,4-butanediol and succinic acid. Detection and quantification limits for the analytes were between 1-2 and 2-7 ng/microl, respectively, and relative standard deviations were between 3 and 7%. Good repeatability and low detection limits made the developed SPE method and subsequent gas chromatography-mass spectrometry (GC-MS) analysis a sensitive tool for identification and quantification of hydrolysis products at early stages of degradation.

  20. Alcohol drinking habits, alcohol dehydrogenase genotypes and risk of acute coronary syndrome.

    Science.gov (United States)

    Tolstrup, Janne S; Hansen, Jane L; Grønbaek, Morten; Vogel, Ulla; Tjønneland, Anne; Joensen, Albert Marni; Overvad, Kim

    2010-07-01

    The risk of myocardial infarction is lower among light-to-moderate drinkers compared with abstainers. Results from some previous studies, but not all, suggest that this association is modified by variations in genes coding for alcohol dehydrogenase (ADH). We aimed to test this hypothesis, including alcohol as both the amount of alcohol and the frequency of drinking. we conducted a nested case-cohort study within the Danish Diet, Cancer and Health study, including 1,645 men (770 incident cases of acute coronary syndrome from 1993-1997 through 2004 and 875 randomly selected controls). Higher alcohol intake (measured as amount or drinking frequency) was associated with lower risk of acute coronary syndrome; however, there was no evidence that these finding were modified by ADH1B or ADH1C genotypes. The importance of functional variation in alcohol dehydrogenase for the association between alcohol drinking habits and the risk of developing acute coronary syndrome, if any, is very limited.