Fusion of Pyruvate Decarboxylase and Alcohol Dehydrogenase Increases Ethanol Production inEscherichia coli

2014 ◽  
Vol 3 (12) ◽  
pp. 976-978 ◽  
Author(s):  
Aleksandra J. Lewicka ◽  
Jan J. Lyczakowski ◽  
Gavin Blackhurst ◽  
Christiana Pashkuleva ◽  
Kyle Rothschild-Mancinelli ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Pyruvate Decarboxylase ◽  
Inescherichia Coli

Effects of manipulation of pyruvate decarboxylase and alcohol dehydrogenase levels on the submergence tolerance of rice

2001 ◽  
Vol 28 (12) ◽  
pp. 1231 ◽  
Author(s):  
Musrur Rahman ◽  
Anil Grover ◽  
W. James Peacock ◽  
Elizabeth S. Dennis ◽  
Marc H. Ellis
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Rice Variety ◽  
Pyruvate Decarboxylase ◽  
Inducible Promoter ◽  
Moderate Increase ◽  
Sense Orientation ◽  
Transgenic Lines ◽  
Hybrid Lines ◽  
Adh Activity

A transgenic approach was taken to manipulate the levels of pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) in rice, in order to investigate whether alteration of ethanol fermentation can affect anaerobic tolerance. A line transformed with an antisense Adh1 construct had only 4–8% of the ADH activity of untransformed plants. This line showed reduced ethanol production and coleoptile growth under anoxia. Mature plants had reduced survival when submerged in water and exposed to anoxia, suggesting that ADH plays an essential role in seed germination and plant survival in the absence of O2. A transgenic line transformed with a cotton Adh2 cDNA in the sense orientation relative to a constitutive promoter, showed 3–4-fold more ADH activity than either untransformed controls, or a flooding-tolerant rice variety (FR13A), both in air and under hypoxia. However, ethanol production by this line was only slightly higher than that of untransformed controls, and there was no increase in survival following anoxia treatments. Three independent transgenic lines containing the ricePdc1 cDNA driven by an anaerobically-inducible promoter (6XARE) showed an increase in PDC1 polypeptide in shoots, but not in roots or endosperm. A moderate increase in PDC activity and ethanol production was observed in shoots of these lines under anaerobic conditions, as well as decreased survival of shoots when submerged and exposed to anoxia. F1 plants containing both the PDC and ADH constructs showed levels of anoxia-tolerance similar to those of untransformed plants. These results suggest that over-production of PDC may be toxic to rice plants because of increased acetaldehyde. Consistent with this view, acetaldehyde levels were appreciably higher in plants over-producing PDC, compared with untransformed plants, or hybrid lines containing both the PDC and ADH constructs.

scholarly journals Zymobacter palmae pyruvate decarboxylase is less efficient than that of Zymomonas mobilis for ethanol production in metabolically engineered Synechocystis sp PCC6803

2019 ◽  
Author(s):  
Lorraine Quinn ◽  
Patricia Armshaw ◽  
Tewfik Soulimane ◽  
Con Sheehan ◽  
Michael P Ryan ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Zymomonas Mobilis ◽  
Pyruvate Decarboxylase ◽  
Inducible Promoter ◽  
Control Strain ◽  
Zymobacter Palmae ◽  
Synechocystis Sp ◽  
Pcc 6803

AbstractPyruvate decarboxylase (PDC) from Zymobacter palmae (ZpPDC) has been reported to have a lower Km the Zymomonas mobilis PDC (ZmPDC). ZpPDC was combined with native slr1192 alcohol dehydrogenase (adh) in an attempt to increase ethanol production in the photoautotrophic cyanobacterium Synechocystis sp. PCC 6803 over constructs created with Zmpdc. Native (Zppdc) and codon optimised (ZpOpdc) versions of the ZpPDC were cloned into a construct where the pdc expression was controlled via the psbA2 light inducible promoter from Synechocystis PCC 6803. These constructs were transformed into wildtype Synechocystis PCC 6803. Ethanol levels were then compared with identical constructs containing the Zmpdc. While strains with the Zppdc (UL071) and ZpOpdc (UL072) constructs did produce ethanol, levels were lower compared to a control strain (UL004) expressing the pdc from Zymomonas mobilis. The utilisation of a PDC with a lower Km from Zymobacter palmae did not result in enhanced ethanol production in Synechocystis PCC 6803.

scholarly journals METABOLIC CHANGES ASSOCIATED WITH OXYGEN STRESS IN PEAR FRUITS

HortScience ◽  
1990 ◽  
Vol 25 (11) ◽  
pp. 1355F-1356
Author(s):  
George D. Nanos ◽  
Roger J. Romani ◽  
Adel A. Kader
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Ethylene Production ◽  
Pyruvate Decarboxylase ◽  
Insect Control ◽  
Pear Fruit ◽  
Oxygen Stress ◽  
Fruit Tissue ◽  
Subsequent Recovery ◽  
Alcohol Dehydrogenase Activity

The response of pear fruits and suspension-cultured pear fruit cells to 0% or 0.25% O2 is being examined to evaluate the feasibility of using such atmospheres for postharvest insect control. These treatments inhibited ethylene production, had no effect on acetaldehyde content, and increased ethanol production in pears kept at 20C for 10 days. The blossom end area of pear fruits was more prone to anaerobiosis, as indicated by increased alcohol dehydrogenase activity and ethanol content. Pear fruit plugs showed increased respiration and ethylene production rates when skin was present compared to plugs without skin. Methods for measuring activity of alcohol dehydrogenase, pyruvate decarboxylase, and pyruvate kinase have been modified and optimized and will be used to determine changes in pear fruit tissue during low O2 treatment and subsequent recovery in air.

scholarly journals Zymobacter palmae Pyruvate Decarboxylase is Less Effective Than That of Zymomonas mobilis for Ethanol Production in Metabolically Engineered Synechocystis sp. PCC6803

2019 ◽  
Vol 7 (11) ◽  
pp. 494 ◽  
Author(s):  
Lorraine Quinn ◽  
Patricia Armshaw ◽  
Tewfik Soulimane ◽  
Con Sheehan ◽  
Michael P. Ryan ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Zymomonas Mobilis ◽  
Pyruvate Decarboxylase ◽  
Biomass Productivity ◽  
Gene Cassette ◽  
Major Effect ◽  
Zymobacter Palmae ◽  
Synechocystis Sp ◽  
Pcc 6803

To produce bioethanol from model cyanobacteria such as Synechocystis, a two gene cassette consisting of genes encoding pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) are required to transform pyruvate first to acetaldehyde and then to ethanol. However the partition of pyruvate to ethanol comes at a cost, a reduction in biomass and pyruvate availability for other metabolic processes. Hence strategies to divert flux to ethanol as a biofuel in Synechocystis are of interest. PDC from Zymobacter palmae (ZpPDC) has been reported to have a lower Km then the Zymomonas mobilis PDC (ZmPDC), which has traditionally been used in metabolic engineering constructs. The Zppdc gene was combined with the native slr1192 alcohol dehydrogenase gene (adhA) in an attempt to increase ethanol production in the photoautotrophic cyanobacterium Synechocystis sp. PCC 6803 over constructs created with the traditional Zmpdc. Native (Zppdc) and codon optimized (ZpOpdc) versions of the ZpPDC were cloned into a construct where pdc expression was controlled via the psbA2 light inducible promoter from Synechocystis sp. PCC 6803. These constructs were transformed into wildtype Synechocystis sp. PCC 6803 for expression and ethanol production. Ethanol levels were then compared with identical constructs containing the Zmpdc. While strains with the Zppdc (UL071) and ZpOpdc (UL072) constructs did produce ethanol, levels were lower compared to a control strain (UL070) expressing the pdc from Zymomonas mobilis. All constructs demonstrated lower biomass productivity illustrating that the flux from pyruvate to ethanol has a major effect on biomass and ultimately overall biofuel productivity. Thus the utilization of a PDC with a lower Km from Zymobacter palmae unusually did not result in enhanced ethanol production in Synechocystis sp. PCC 6803.

Consolidated Bioprocessing for Bioethanol Production by Metabolically Engineered Bacillus Subtilis Strains

2021 ◽  
Author(s):  
Fatemeh Maleki ◽  
Mohammad Changizian ◽  
Narges Zolfaghari ◽  
Sarah Rajaei ◽  
Kambiz Akbari Noghabi ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Plant Biomass ◽  
Fusion Gene ◽  
Consolidated Bioprocessing ◽  
Gene Dosage ◽  
Pyruvate Decarboxylase ◽  
Single Step ◽  
Bioethanol Production ◽  
Genes Encoding

Abstract Background: Bioethanol produced by fermentative microorganisms is regarded as an alternative to fossil fuel. Bioethanol to be used as a viable energy source must be produced cost-effectively by removing expense-intensive steps such as the enzymatic hydrolysis of substrate. Consolidated bioprocessing (CBP) is believed to be a practical solution combining saccharification and fermentation in a single step catalyzed by a microorganism. Bacillus subtills with innate ability to grow on a diversity of carbohydrates seems promising for affordable CBP bioethanol production using renewable plant biomass and wastes. Results: In this study, the genes encoding alcohol dehydrogenase from Z. mobilis (adhZ) and S. cerevisiae (adhS) were each used with Z. mobilis pyruvate decarboxylase gene (pdcZ) to create ethanologenic operons in a lactate-deficient (Δldh) B. subtilis resulting in NZ and NZS strains, respectively. The S. cerevisiae adhS caused significantly more ethanol production by NZS and therefore was used to make two other operons including one with double copies of both pdcZ and adhS and the other with a single pdcZ but double adhS genes expressed in N(ZS)2 and NZS2 strains, respectively. In addition, two fusion genes were constructed with pdcZ and adhS in alternate orientations and used for ethanol production by the harboring strains namely NZ:S and NS:Z, respectively. While the increase of gene dosage was not associated with elevated carbon flow for ethanol production, the fusion gene adhS:pdcZ resulted in more than two times increase of productivity by strain NS:Z as compared with NZS during 48 h fermentation. The CBP ethanol production by NZS and NS:Z using potatoes resulted in 16.3 g/L and 21.5 g/L ethanol during 96 h fermentation, respectively. Conclusion: In this study for the first time, Bacillus subtilis was successfully used for CBP ethanol production with S. cerevisiae alcohol dehydrogenase. The results of the study provide insights on the potentials of B. subtilis for affordable bioethanol production from inexpensive plant biomass and wastes. However, the potentials need to be improved by metabolic and process engineering for higher yields of ethanol production and plant biomass utilization.

scholarly journals Pyruvate-to-ethanol pathway in Entamoeba histolytica

1978 ◽  
Vol 171 (1) ◽  
pp. 225-230 ◽  
Author(s):  
H S Lo ◽  
R E Reeves
Keyword(s):  
Enzyme Activity ◽  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Entamoeba Histolytica ◽  
Aldehyde Dehydrogenase ◽  
Pyruvate Decarboxylase ◽  
Acetyl Coa ◽  
Key Enzyme ◽  
Pyruvate Synthase ◽  
Soluble Enzymes

The pyruvate-to-ethanol pathway in Entamoeba histolytica is unusual when compared with most investigated organisms. Pyruvate decarboxylase (EC 4.1.1.1), a key enzyme for ethanol production, is not found. Pyruvate is converted into acetyl-CoA and CO2 by the enzyme pyruvate synthase (EC 1.2.7.1), which has been demonstrated previously in this parasitic amoeba. Acetyl-CoA is reduced to acetaldehyde and CoA by the enzyme aldehyde dehydrogenase (acylating) (EC 1.2.1.10) at an enzyme activity of 9 units per g of fresh cells with NADH as a reductant. Acetaldehyde is further reduced by either a previously identified NADP+-linked alcohol dehydrogenase or by a newly found NAD+-linked alcohol dehydrogenase at an enzyme activity of 136 units per g of fresh cells. Ethanol is identified as the product of soluble enzymes of amoeba acting on pyruvate or acetyl-CoA. This result is confirmed by radioactive isotopic, spectrophotometric and gas-chromatographic methods.

scholarly journals Rewiring Lactococcus lactis for Ethanol Production

2013 ◽  
Vol 79 (8) ◽  
pp. 2512-2518 ◽  
Author(s):  
Christian Solem ◽  
Tore Dehli ◽  
Peter Ruhdal Jensen
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Lactococcus Lactis ◽  
Model Organism ◽  
Pyruvate Decarboxylase ◽  
Major Product ◽  
High Yield ◽  
Content Type ◽  
Synthetic Promoters ◽  
In The Wild

ABSTRACTLactic acid bacteria (LAB) are known for their high tolerance toward organic acids and alcohols (R. S. Gold, M. M. Meagher, R. Hutkins, and T. Conway, J. Ind. Microbiol.10:45–54, 1992) and could potentially serve as platform organisms for production of these compounds. In this study, we attempted to redirect the metabolism of LAB model organismLactococcus lactistoward ethanol production. Codon-optimizedZymomonas mobilispyruvate decarboxylase (PDC) was introduced and expressed from synthetic promoters in different strain backgrounds. In the wild-typeL. lactisstrain MG1363 growing on glucose, only small amounts of ethanol were obtained after introducing PDC, probably due to a low native alcohol dehydrogenase activity. When the same strains were grown on maltose, ethanol was the major product and lesser amounts of lactate, formate, and acetate were formed. Inactivating the lactate dehydrogenase genesldhX,ldhB, andldhand introducing codon-optimizedZ. mobilisalcohol dehydrogenase (ADHB) in addition to PDC resulted in high-yield ethanol formation when strains were grown on glucose, with only minor amounts of by-products formed. Finally, a strain with ethanol as the sole observed fermentation product was obtained by further inactivating the phosphotransacetylase (PTA) and the native alcohol dehydrogenase (ADHE).

scholarly journals Physiology, Genomics, and Pathway Engineering of an Ethanol-Tolerant Strain of Clostridium phytofermentans

2015 ◽  
Vol 81 (16) ◽  
pp. 5440-5448 ◽  
Author(s):  
Andrew C. Tolonen ◽  
Trevor R. Zuroff ◽  
Mohandass Ramya ◽  
Magali Boutard ◽  
Tristan Cerisy ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Ethanol Production ◽  
Ethanol Yield ◽  
Complex Cation ◽  
Pyruvate Decarboxylase ◽  
Pathway Engineering ◽  
Solvent Tolerance ◽  
Potential Cost ◽  
Content Type ◽  
Clostridium Phytofermentans

ABSTRACTNovel processing strategies for hydrolysis and fermentation of lignocellulosic biomass in a single reactor offer large potential cost savings for production of biocommodities and biofuels. One critical challenge is retaining high enzyme production in the presence of elevated product titers. Toward this goal, the cellulolytic, ethanol-producing bacteriumClostridium phytofermentanswas adapted to increased ethanol concentrations. The resulting ethanol-tolerant (ET) strain has nearly doubled ethanol tolerance relative to the wild-type level but also reduced ethanol yield and growth at low ethanol concentrations. The genome of the ET strain has coding changes in proteins involved in membrane biosynthesis, the Rnf complex, cation homeostasis, gene regulation, and ethanol production. In particular, purification of the mutant bifunctional acetaldehyde coenzyme A (CoA)/alcohol dehydrogenase showed that a G609D variant abolished its activities, including ethanol formation. Heterologous expression ofZymomonas mobilispyruvate decarboxylase and alcohol dehydrogenase in the ET strain increased cellulose consumption and restored ethanol production, demonstrating how metabolic engineering can be used to overcome disadvantageous mutations incurred during adaptation to ethanol. We discuss how genetic changes in the ET strain reveal novel potential strategies for improving microbial solvent tolerance.

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