scholarly journals Impact of acetolactate synthase inactivation on 1,3-propanediol fermentation byKlebsiella pneumoniae

2018 ◽  
Author(s):  
Sheng Zhou ◽  
Youhua Huang ◽  
Xinliang Mao ◽  
Lili Li ◽  
Chuanyu Guo ◽  
...  
Keyword(s):  
Metabolic Flux ◽  
Wild Type Strain ◽  
Lactate Production ◽  
Acetolactate Synthase ◽  
Gene Replacement ◽  
Wild Type ◽  
Fed Batch ◽  
Formation Pathway ◽  
Fed Batch Fermentation ◽  
The Impact

Abstract1,3-Propanediol (1,3-PDO) is an important compound that is mainly used in industry for polymer production. Fermentation of 1,3-PDO from glycerol by marineKlebsiella pneumoniaeis accompanied by formation of 2,3-butanediol (2,3-BDO) as one of the main byproduct. The first step in the formation of 2,3-BDO from pyruvate is catalyzed by acetolactate synthase (ALS), an enzyme that competes with 1,3-PDO oxidoreductase for the cofactor NADH. This study aimed to analyze the impact of engineering the 2,3-BDO formation pathway via inactivation of ALS on 1,3-PDO fermentation by marineK. pneumoniaeHSL4. An ALS mutant was generated using Red recombinase assisted gene replacement. The ALS specific activities ofK. pneumoniaeΔALS were notably lower than that of the wild-type strain. Fed-batch fermentation of the mutant strain resulted in a 1,3-PDO concentration, productivity and conversion of 72.04 g L−1, 2.25 g L−1h−1, and 0.41 g g−1, a slightly increase compared with the parent strain. Moreover, inactivation of ALS decreasedmeso-2,3-BDO formation to trace amounts, significantly increased 2S,3S-BDO and lactate production, and a pronounced redistribution of intracellular metabolic flux was apparent.

scholarly journals Impact of Inorganic Carbon Availability on Microcystin Production by Microcystis aeruginosa PCC 7806

2007 ◽  
Vol 73 (21) ◽  
pp. 6994-7002 ◽  
Author(s):  
Sabine J�hnichen ◽  
Tilo Ihle ◽  
Thomas Petzoldt ◽  
J�rgen Benndorf
Keyword(s):  
Microcystis Aeruginosa ◽  
Type Strain ◽  
Inorganic Carbon ◽  
Wild Type Strain ◽  
Light Cycle ◽  
Variable Fluorescence ◽  
Wild Type ◽  
Dark Light ◽  
Sodium Dependent ◽  
The Impact

ABSTRACT Batch culture experiments with the cyanobacterium Microcystis aeruginosa PCC 7806 were performed in order to test the hypothesis that microcystins (MCYSTs) are produced in response to a relative deficiency of intracellular inorganic carbon (Ci,i). In the first experiment, MCYST production was studied under increased Ci,i deficiency conditions, achieved by restricting sodium-dependent bicarbonate uptake through replacement of sodium bicarbonate in the medium with its potassium analog. The same experimental approach was used in a second experiment to compare the response of the wild-type strain M. aeruginosa PCC 7806 with its mcyB mutant, which lacks the ability to produce MCYSTs. In a third experiment, the impact of varying the Ci,i status on MCYST production was examined without suppressing the sodium-dependent bicarbonate transporter; instead, a detailed investigation of a dark-light cycle was performed. In all experiments, a relative Ci,i deficiency was indicated by an elevated variable fluorescence signal and led to enhanced phycocyanin cell quotas. Higher MCYST cell quotas (in the first and third experiments) and increased total (intracellular plus extracellular) MCYST production (in the first experiment) were detected with increased Ci,i deficiency. Furthermore, the MCYST-producing wild-type strain and its mcyB mutant showed basically the same response to restrained inorganic carbon uptake, with elevated variable fluorescence and phycocyanin cell quotas with increased Ci,i deficiency. The response of the wild type, however, was distinctly stronger and also included elevated chlorophyll a cell quotas. These differences indicate the limited ability of the mutant to adapt to low-Ci,i conditions. We concluded that MCYSTs may be involved in enhancing the efficiency of the adaptation of the photosynthetic apparatus to fluctuating inorganic carbon conditions in cyanobacterial cells.

A dynamic metabolic flux balance based model of fed-batch fermentation ofbordetella pertussis

2013 ◽  
Vol 29 (2) ◽  
pp. 520-531 ◽  
Author(s):  
Hector Budman ◽  
Nilesh Patel ◽  
Melih Tamer ◽  
Walid Al-Gherwi
Keyword(s):  
Batch Fermentation ◽  
Metabolic Flux ◽  
Fed Batch ◽  
Flux Balance ◽  
Fed Batch Fermentation ◽  
Ofbordetella Pertussis

scholarly journals Side-by-Side Comparison of Clean and Biomass-Derived, Impurity-Containing Syngas as Substrate for Acetogenic Fermentation with Clostridium ljungdahlii

Fermentation ◽  
2020 ◽  
Vol 6 (3) ◽  
pp. 84
Author(s):  
Alba Infantes ◽  
Michaela Kugel ◽  
Klaus Raffelt ◽  
Anke Neumann
Keyword(s):  
Biomass Gasification ◽  
Product Formation ◽  
Inhibitory Effects ◽  
Fed Batch ◽  
Gaseous Mixtures ◽  
Clostridium Ljungdahlii ◽  
Acetogenic Bacteria ◽  
Fed Batch Fermentation ◽  
Total Productivity ◽  
The Impact

Syngas, the product of biomass gasification, can play an important role in moving towards the production of renewable chemical commodities, by using acetogenic bacteria to ferment those gaseous mixtures. Due to the complex and changing nature of biomass, the composition and the impurities present in the final biomass-derived syngas will vary. Because of this, it is important to assess the impact of these factors on the fermentation outcome, in terms of yields, productivity, and product formation and ratio. In this study, Clostridium ljungdahlii was used in a fed-batch fermentation system to analyze the effect of three different biomass-derived syngases, and to compare them to equivalent, clean syngas mixtures. Additionally, four other clean syngas mixtures were used, and the effects on product ratio, productivity, yield, and growth were documented. All biomass-derived syngases were suitable to be used as substrates, without experiencing any complete inhibitory effects. From the obtained results, it is clear that the type of syngas, biomass-derived or clean, had the greatest impact on product formation ratios, with all biomass-derived syngases producing more ethanol, albeit with lesser total productivity.

scholarly journals C-5(6) Sterol Desaturase from Tetrahymena thermophila: Gene Identification and Knockout, Sequence Analysis, and Comparison to Other C-5(6) Sterol Desaturases

2009 ◽  
Vol 8 (8) ◽  
pp. 1287-1297 ◽  
Author(s):  
Alejandro D. Nusblat ◽  
Sebastián R. Najle ◽  
Mariela L. Tomazic ◽  
Antonio D. Uttaro ◽  
Clara B. Nudel
Keyword(s):  
Type Strain ◽  
Deletion Mutant ◽  
Wild Type Strain ◽  
Tetrahymena Thermophila ◽  
Gene Replacement ◽  
Gene Identification ◽  
Wild Type ◽  
Reverse Transcription Pcr ◽  
Gene Coding ◽  
Transmembrane Regions

ABSTRACT The gene coding for a C-5(6) sterol desaturase in Tetrahymena thermophila, DES5A, has been identified by the knockout of the TTHERM_01194720 sequence. Macronucleus transformation was achieved by biolistic bombardment and gene replacement through phenotypic assortment, using paromomycin as the selective agent. A knockout cell line (KO270) showed a phenotype consistent with that of the DES5A deletion mutant. KO270 converted only 6% of the added sterol into the C-5 unsaturated derivative, while the wild type accumulated 10-fold larger amounts under similar conditions. The decreased desaturation activity is specific for the C-5(6) position of lathosterol and cholestanol; other desaturations, namely C-7(8) and C-22(23), were not affected. Analysis by reverse transcription-PCR reveals that DES5A is transcribed both in the presence and absence of cholestanol in wild-type cells, whereas the transcribed gene was not detected in KO270. The growth of KO270 was undistinguishable from that of the wild-type strain. Des5Ap resembles known C-5(6) sterol desaturases, displaying the three typical histidine motifs, four hydrophobic transmembrane regions, and two other highly conserved domains of unknown function. A phylogenetic analysis placed T. thermophila's enzyme and Paramecium orthologues in a cluster together with functionally characterized C-5 sterol desaturases from vertebrates, fungi, and plants, although in a different branch.

Co-Expression of Threonine Dehydratase and Acetolactate Synthase in Escherichia Coli for L-Isoleucine Production

2014 ◽  
Vol 989-994 ◽  
pp. 997-1002 ◽  
Author(s):  
Jian Wang ◽  
Jia Kai Sun ◽  
Qing Yang Xu
Keyword(s):  
Escherichia Coli ◽  
Corynebacterium Glutamicum ◽  
Carbon Flux ◽  
Batch Fermentation ◽  
Acetolactate Synthase ◽  
Fed Batch ◽  
Threonine Dehydratase ◽  
E Coli ◽  
Fed Batch Fermentation ◽  
Fermentation Period

Metabolic engineering ofCorynebacterium glutamicumhas sought to divert carbon into L-isoleucine. However, the fermentation period of this strain is long. TheC.glutamicumYILW strain (LeuL, AHVr, SGr, Leu-MEr) was previously derived by repeated compound mutagenesis which could accumulate 20.2 g/L L-isoleucine in a 5-L jar fermentor. Overexpression of the threonine dehydratase gene (ilvA) fromCorynebacterium glutamicumYILW and coexpression of threonine dehydratase and acetolactate synthase (ilvBN) from it were employed to divert carbon flux toward L-isoleucine. The strainE. coliTRFC with the expression ofilvA could accumulate L-isoleucine of 6.8 g/L without accumulation of any L-threonine by fed-batch fermentation in a 5-L jar fermentor. However, the production of L-isoleucine by the strainE.coliTRFC with the co-expression ofilvA andilvBN was decreased by 19.1%, and the production of L-valine was increased by 40% compared with that ofE. coliTRFC with the expression ofilvA.

scholarly journals Metabolic Engineering of Lactobacillus helveticus CNRZ32 for Production of Purel-(+)-Lactic Acid

2000 ◽  
Vol 66 (9) ◽  
pp. 3835-3841 ◽  
Author(s):  
Kari Kyl�-Nikkil� ◽  
Mervi Hujanen ◽  
Matti Leisola ◽  
Airi Palva
Keyword(s):  
Lactic Acid ◽  
Structural Gene ◽  
Acid Production ◽  
Wild Type Strain ◽  
Lactic Acid Production ◽  
Gene Replacement ◽  
Lactobacillus Helveticus ◽  
Statistical Experimental Design ◽  
Wild Type ◽  
In The Wild

ABSTRACT Expression of d-(−)-lactate dehydrogenase (d-LDH) and l-(+)-LDH genes (ldhDand ldhL, respectively) and production ofd-(−)- and l-(+)-lactic acid were studied inLactobacillus helveticus CNRZ32. In order to develop a host for production of pure l-(+)-isomer of lactic acid, twoldhD-negative L. helveticus CNRZ32 strains were constructed using gene replacement. One of the strains was constructed by deleting the promoter region of the ldhD gene, and the other was constructed by replacing the structural gene ofldhD with an additional copy of the structural gene (ldhL) of l-LDH of the same species. The resulting strains were designated GRL86 and GRL89, respectively. In strain GRL89, the second copy of the ldhL structural gene was expressed under the ldhD promoter. The twod-LDH-negative strains produced onlyl-(+)-lactic acid in an amount equal to the total lactate produced by the wild type. The maximum l-LDH activity was found to be 53 and 93% higher in GRL86 and GRL89, respectively, than in the wild-type strain. Furthermore, process variables forl-(+)-lactic acid production by GRL89 were optimized using statistical experimental design and response surface methodology. The temperature and pH optima were 41�C and pH 5.9. At low pH, when the growth and lactic acid production are uncoupled, strain GRL89 produced approximately 20% more lactic acid than GRL86.

scholarly journals Global Analysis of Cellular Factors and Responses Involved in Pseudomonas aeruginosa Resistance to Arsenite

2005 ◽  
Vol 187 (14) ◽  
pp. 4853-4864 ◽  
Author(s):  
Kislay Parvatiyar ◽  
Eyad M. Alsabbagh ◽  
Urs A. Ochsner ◽  
Michelle A. Stegemeyer ◽  
Alan G. Smulian ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Pseudomonas Aeruginosa ◽  
Glutathione Reductase ◽  
Type Strain ◽  
Wild Type Strain ◽  
Storage Proteins ◽  
Global Analysis ◽  
Wild Type ◽  
Isogenic Mutants ◽  
The Impact

ABSTRACT The impact of arsenite [As(III)] on several levels of cellular metabolism and gene regulation was examined in Pseudomonas aeruginosa. P. aeruginosa isogenic mutants devoid of antioxidant enzymes or defective in various metabolic pathways, DNA repair systems, metal storage proteins, global regulators, or quorum sensing circuitry were examined for their sensitivity to As(III). Mutants lacking the As(III) translocator (ArsB), superoxide dismutase (SOD), catabolite repression control protein (Crc), or glutathione reductase (Gor) were more sensitive to As(III) than wild-type bacteria. The MICs of As(III) under aerobic conditions were 0.2, 0.3, 0.8, and 1.9 mM for arsB, sodA sodB, crc, and gor mutants, respectively, and were 1.5- to 13-fold less than the MIC for the wild-type strain. A two-dimensional gel/matrix-assisted laser desorption ionization-time of flight analysis of As(III)-treated wild-type bacteria showed significantly (>40-fold) increased levels of a heat shock protein (IbpA) and a putative allo-threonine aldolase (GlyI). Smaller increases (up to 3.1-fold) in expression were observed for acetyl-coenzyme A acetyltransferase (AtoB), a probable aldehyde dehydrogenase (KauB), ribosomal protein L25 (RplY), and the probable DNA-binding stress protein (PA0962). In contrast, decreased levels of a heme oxygenase (HemO/PigA) were found upon As(III) treatment. Isogenic mutants were successfully constructed for six of the eight genes encoding the aforementioned proteins. When treated with sublethal concentrations of As(III), each mutant revealed a marginal to significant lag period prior to resumption of apparent normal growth compared to that observed in the wild-type strain. Our results suggest that As(III) exposure results in an oxidative stress-like response in P. aeruginosa, although activities of classic oxidative stress enzymes are not increased. Instead, relief from As(III)-based oxidative stress is accomplished from the collective activities of ArsB, glutathione reductase, and the global regulator Crc. SOD appears to be involved, but its function may be in the protection of superoxide-sensitive sulfhydryl groups.

scholarly journals A general framework for modelling the impact of co-infections on pathogen evolution

2019 ◽  
Vol 16 (155) ◽  
pp. 20190165 ◽  
Author(s):  
Mary Bushman ◽  
Rustom Antia
Keyword(s):  
Mutant Strain ◽  
Type Strain ◽  
General Framework ◽  
Wild Type Strain ◽  
Theoretical Models ◽  
Epidemiological Model ◽  
Wild Type ◽  
Pathogen Evolution ◽  
Evolutionary Emergence ◽  
The Impact

Theoretical models suggest that mixed-strain infections, or co-infections, are an important driver of pathogen evolution. However, the within-host dynamics of co-infections vary enormously, which complicates efforts to develop a general understanding of how co-infections affect evolution. Here, we develop a general framework which condenses the within-host dynamics of co-infections into a few key outcomes, the most important of which is the overall R 0 of the co-infection. Similar to how fitness is determined by two different alleles in a heterozygote, the R 0 of a co-infection is a product of the R 0 values of the co-infecting strains, shaped by the interaction of those strains at the within-host level. Extending the analogy, we propose that the overall R 0 reflects the dominance of the co-infecting strains, and that the ability of a mutant strain to invade a population is a function of its dominance in co-infections. To illustrate the utility of these concepts, we use a within-host model to show how dominance arises from the within-host dynamics of a co-infection, and then use an epidemiological model to demonstrate that dominance is a robust predictor of the ability of a mutant strain to save a maladapted wild-type strain from extinction (evolutionary emergence).

scholarly journals The transcriptomic response of a wine strain of Lachancea thermotolerans to oxygen deprivation

2020 ◽  
Vol 20 (7) ◽  
Author(s):  
Kirti Shekhawat ◽  
Florian F Bauer ◽  
Mathabatha E Setati
Keyword(s):  
Metabolic Flux ◽  
Anaerobic Fermentation ◽  
Lactate Production ◽  
Starter Cultures ◽  
Wine Fermentation ◽  
Limiting Factor ◽  
Transcriptomic Response ◽  
Fermentation Conditions ◽  
The Impact ◽  
Lachancea Thermotolerans

ABSTRACT The yeast Lachancea thermotolerans is of significant biotechnological interest, and selected strains of this species have become commonly used starter cultures in wine fermentation. However, the impact of this species on wine is frequently limited by the rapid dominance of Saccharomyces cerevisiae strains which are better adapted to wine alcoholic fermentation conditions. Previous studies have shown that the major limiting factor for L. thermotolerans competitive performance in the wine ecosystem is oxygen availability, and not ethanol levels as had been previously suggested. Here we investigated the transcriptional response of L. thermotolerans to anaerobiosis in wine fermentation conditions. The data show that L. thermotolerans broadly redirects gene expression towards genes involved in central carbon metabolism, lipid metabolism, remodeling of the cell wall as well as autophagy. Furthermore, the induction of genes that are likely involved in the generation of lactate indicates a redirection of metabolic flux towards this metabolite. The data provide the first insight into the oxygen-dependent response of L. thermotolerans and suggest potential genetic targets to improve lactate production and/or anaerobic fermentation performance of this yeast.

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