| 1. |
- Bhattacharya, Abhishek, et al.
(författare)
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Cross-Feeding and Enzymatic Catabolism for Mannan-Oligosaccharide Utilization by the Butyrate-Producing Gut Bacterium Roseburia hominis A2-183
- 2022
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Ingår i: Microorganisms. - : MDPI AG. - 2076-2607. ; 10:12
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Tidskriftsartikel (refereegranskat)abstract
- β-Mannan is abundant in the human diet and in hemicellulose derived from softwood. Linear or galactose-substituted β-mannan-oligosaccharides (MOS/GMOSs) derived from β-mannan are considered emerging prebiotics that could stimulate health-associated gut microbiota. However, the underlying mechanisms are not yet resolved. Therefore, this study investigated the cross-feeding and metabolic interactions between Bifidobacterium adolescentis ATCC 15703, an acetate producer, and Roseburia hominis A2-183 DSMZ 16839, a butyrate producer, during utilization of MOS/GMOSs. Cocultivation studies suggest that both strains coexist due to differential MOS/GMOS utilization, along with the cross-feeding of acetate from B. adolescentis E194a to R. hominis A2-183. The data suggest that R. hominis A2-183 efficiently utilizes MOS/GMOS in mono- and cocultivation. Notably, we observed the transcriptional upregulation of certain genes within a dedicated MOS/GMOS utilization locus (RhMosUL), and an exo-oligomannosidase (RhMan113A) gene located distally in the R. hominis A2-183 genome. Significantly, biochemical analysis of β-1,4 mannan-oligosaccharide phosphorylase (RhMOP130A), α-galactosidase (RhGal36A), and exo-oligomannosidase (RhMan113A) suggested their potential synergistic role in the initial utilization of MOS/GMOSs. Thus, our results enhance the understanding of MOS/GMOS utilization by potential health-promoting human gut microbiota and highlight the role of cross-feeding and metabolic interactions between two secondary mannan degraders inhabiting the same ecological niche in the gut.
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| 2. |
- Boshagh, Fatemeh, et al.
(författare)
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Application of kinetic models in dark fermentative hydrogen production–A critical review
- 2022
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Ingår i: International Journal of Hydrogen Energy. - : Elsevier BV. - 0360-3199. ; 47:52, s. 21952-21968
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Forskningsöversikt (refereegranskat)abstract
- Kinetic modeling could be viewed as an important step in developing a bioprocess, since models can be used in process control, reducing costs and optimizing processes. In the present study, the application of kinetic models in dark fermentative hydrogen production has been investigated. A wide variety of kinetic models are addressed and compared regarding their accuracy to fit the data. This literature survey indicates that the modified Gompertz was extensively used to describe the production of hydrogen, organic acids and alcohols, substrate degradation, and biomass growth. The development of kinetic models can assist researchers to identify the most important variables, facilitate future research, and maximize hydrogen production.
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| 3. |
- Byrne, Eoin, et al.
(författare)
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Characterization and adaptation of Caldicellulosiruptor strains to higher sugar concentrations, targeting enhanced hydrogen production from lignocellulosic hydrolysates
- 2021
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Ingår i: Biotechnology for Biofuels. - : BioMed Central Ltd. - 1754-6834. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: The members of the genus Caldicellulosiruptor have the potential for future integration into a biorefinery system due to their capacity to generate hydrogen close to the theoretical limit of 4 mol H2/mol hexose, use a wide range of sugars and can grow on numerous lignocellulose hydrolysates. However, members of this genus are unable to survive in high sugar concentrations, limiting their ability to grow on more concentrated hydrolysates, thus impeding their industrial applicability. In this study five members of this genus, C.owensensis, C. kronotskyensis, C.bescii, C.acetigenus and C.kristjanssonii, were developed to tolerate higher sugar concentrations through an adaptive laboratory evolution (ALE) process. The developed mixed population C.owensensis CO80 was further studied and accompanied by the development of a kinetic model based on Monod kinetics to quantitatively compare it with the parental strain. Results: Mixed populations of Caldicellulosiruptor tolerant to higher glucose concentrations were obtained with C.owensensis adapted to grow up to 80 g/L glucose; other strains in particular C. kristjanssonii demonstrated a greater restriction to adaptation. The C.owensensis CO80 mixed population was further studied and demonstrated the ability to grow in glucose concentrations up to 80 g/L glucose, but with reduced volumetric hydrogen productivities (QH2) and incomplete sugar conversion at elevated glucose concentrations. In addition, the carbon yield decreased with elevated concentrations of glucose. The ability of the mixed population C.owensensis CO80 to grow in high glucose concentrations was further described with a kinetic growth model, which revealed that the critical sugar concentration of the cells increased fourfold when cultivated at higher concentrations. When co-cultured with the adapted C.saccharolyticus G5 mixed culture at a hydraulic retention time (HRT) of 20 h, C.owensensis constituted only 0.09–1.58% of the population in suspension. Conclusions: The adaptation of members of the Caldicellulosiruptor genus to higher sugar concentrations established that the ability to develop improved strains via ALE is species dependent, with C.owensensis adapted to grow on 80 g/L, whereas C.kristjanssonii could only be adapted to 30 g/L glucose. Although C.owensensis CO80 was adapted to a higher sugar concentration, this mixed population demonstrated reduced QH2 with elevated glucose concentrations. This would indicate that while ALE permits adaptation to elevated sugar concentrations, this approach does not result in improved fermentation performances at these higher sugar concentrations. Moreover, the observation that planktonic mixed culture of CO80 was outcompeted by an adapted C.saccharolyticus, when co-cultivated in continuous mode, indicates that the robustness of CO80 mixed culture should be improved for industrial application. © 2021, The Author(s).
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| 4. |
- Kristjansdottir, Thordis, et al.
(författare)
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Engineering the carotenoid biosynthetic pathway in Rhodothermus marinus for lycopene production
- 2020
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Ingår i: Metabolic Engineering Communications. - : Elsevier BV. - 2214-0301. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Rhodothermus marinus has the potential to be well suited for biorefineries, as an aerobic thermophile that produces thermostable enzymes and is able to utilize polysaccharides from different 2nd and 3rd generation biomass. The bacterium produces valuable chemicals such as carotenoids. However, the native carotenoids are not established for industrial production and R. marinus needs to be genetically modified to produce higher value carotenoids. Here we genetically modified the carotenoid biosynthetic gene cluster resulting in three different mutants, most importantly the lycopene producing mutant TK-3 (ΔtrpBΔpurAΔcruFcrtB::trpBcrtBT.thermophilus). The genetic modifications and subsequent structural analysis of carotenoids helped clarify the carotenoid biosynthetic pathway in R. marinus. The nucleotide sequences encoding the enzymes phytoene synthase (CrtB) and the previously unidentified 1′,2′-hydratase (CruF) were found fused together and encoded by a single gene in R. marinus. Deleting only the cruF part of the gene did not result in an active CrtB enzyme. However, by deleting the entire gene and inserting the crtB gene from Thermus thermophilus, a mutant strain was obtained, producing lycopene as the sole carotenoid. The lycopene produced by TK-3 was quantified as 0.49 g/kg CDW (cell dry weight).
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| 5. |
- Ron, Emanuel Y C, et al.
(författare)
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Cultivation technology development of Rhodothermus marinus DSM 16675
- 2019
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Ingår i: Extremophiles : life under extreme conditions. - : Springer Science and Business Media LLC. - 1433-4909. ; 23:6, s. 735-745
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Tidskriftsartikel (refereegranskat)abstract
- This work presents an evaluation of batch, fed-batch, and sequential batch cultivation techniques for production of R. marinus DSM 16675 and its exopolysaccharides (EPSs) and carotenoids in a bioreactor, using lysogeny broth (LB) and marine broth (MB), respectively, in both cases supplemented with 10 g/L maltose. Batch cultivation using LB supplemented with maltose (LBmalt) resulted in higher cell density (OD620 = 6.6) than use of MBmalt (OD620 = 1.7). Sequential batch cultivation increased the cell density threefold (OD620 = 20) in LBmalt and eightfold (OD620 = 14) in MBmalt. In both single and sequential batches, the production of carotenoids and EPSs using LBmalt was detected in the exponential phase and stationary phase, respectively, while in MBmalt formation of both products was detectable in both the exponential and stationary phases of the culture. Heteropolymeric EPSs were produced with an overall volumetric productivity (QE) of 0.67 (mg/L h) in MBmalt and the polymer contained xylose. In LB, QE was lower (0.1 mg/L h) and xylose could not be detected in the composition of the produced EPSs. In conclusion, this study showed the importance of a process design and medium source for production of R. marinus DSM 16675 and its metabolites.
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| 6. |
- Seshagiri Rao, Nikhil, et al.
(författare)
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Non-inhibitory levels of oxygen during cultivation increase freeze-drying stress tolerance in Limosilactobacillus reuteri DSM 17938
- 2023
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Ingår i: Frontiers in Microbiology. - 1664-302X. ; 14
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Tidskriftsartikel (refereegranskat)abstract
- The physiological effects of oxygen on Limosilactobacillus reuteri DSM 17938 during cultivation and the ensuing properties of the freeze-dried probiotic product was investigated. On-line flow cytometry and k-means clustering gating was used to follow growth and viability in real time during cultivation. The bacterium tolerated aeration at 500 ml/min, with a growth rate of 0.74 ± 0.13 h-1 which demonstrated that low levels of oxygen did not influence the growth kinetics of the bacterium. Modulation of the redox metabolism was, however, seen already at non-inhibitory oxygen levels by 1.5-fold higher production of acetate and 1.5-fold lower ethanol production. A significantly higher survival rate in the freeze-dried product was observed for cells cultivated in presence of oxygen compared to absence of oxygen (61.8 ± 2.4 % vs 11.5 ± 4.3 %), coinciding with a higher degree of unsaturated fatty acids (UFA:SFA ratio of 10 for air sparged vs 3.59 for N2 sparged conditions.). Oxygen also resulted in improved bile tolerance and boosted 5’nucleotidase activity (370 U/L vs 240 U/L in N2 sparged conditions) but lower tolerance to acidic conditions compared bacteria grown under complete anaerobic conditions which survived up to 90 min of exposure at pH 2. Overall, our results indicate the controlled supply of oxygen during production may be used as means for probiotic activity optimisation of L. reuteri DSM 17938.
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| 7. |
- Soto, Luis Romero, et al.
(författare)
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Hydrogen and polyhydroxybutyrate production from wheat straw hydrolysate using Caldicellulosiruptor species and Ralstonia eutropha in a coupled process
- 2019
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Ingår i: Bioresource Technology. - : Elsevier BV. - 0960-8524. ; 272, s. 259-266
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Tidskriftsartikel (refereegranskat)abstract
- This report presents an integrated biorefinery concept in which wheat straw hydrolysate was treated with co-cultures of osmotolerant thermophilic bacterial strains, Caldicellulosiruptor saccharolyticus and C. owensensis to obtain hydrogen, while the liquid effluent containing acetate and residual glucose was used as feed for polyhydroxybutyrate (PHB) production by Ralstonia eutropha. The Caldicellulosiruptor spp. co-culture consumed 10.8 g/L of pretreated straw sugars, glucose and xylose, producing 134 mmol H2/L. PHB accumulation by R. eutropha was first studied in minimal salts medium using acetate with/without glucose as carbon source. Addition of salts promoted cell growth and PHB production in the effluent. Fed-batch cultivation in a nitrogen limited medium with 40% (v/v) aeration resulted in a cell density of 15.1 g/L with PHB content of 80.1% w/w and PHB concentration of 12.1 g/L, while 20% aeration gave a cell density of 11.3 g/L with 83.4% w/w PHB content and 9.4 g/L PHB concentration.
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| 8. |
- Sreenivas, Krishnan, et al.
(författare)
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Evaluation of Pyrophosphate‐Driven Proton Pumps in Saccharomyces cerevisiae under Stress Conditions
- 2024
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Ingår i: Microorganisms. - 2076-2607. ; 12:3
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Tidskriftsartikel (refereegranskat)abstract
- In Saccharomyces cerevisiae, pH homeostasis is reliant on ATP due to the use of proton-translocating ATPase (H+-ATPase) which constitutes a major drain within cellular ATP supply. Here, an exogenous proton-translocating pyrophosphatase (H+-PPase) from Arabidopsis thaliana, which uses inorganic pyrophosphate (PPi) rather than ATP, was evaluated for its effect on reducing the ATP burden. The H+-Ppase was localized to the vacuolar membrane or to the cell membrane, and their impact was studied under acetate stress at a low pH. Biosensors (pHluorin and mQueen-2m) were used to observe changes in intracellular pH (pHi) and ATP levels during growth on either glucose or xylose. A significant improvement of 35% in the growth rate at a pH of 3.7 and 6 g·L−1 acetic acid stress was observed in the vacuolar membrane H+-PPase strain compared to the parent strain. ATP levels were elevated in the same strain during anaerobic glucose and xylose fermentations. During anaerobic xylose fermentations, co-expression of pHluorin and a vacuolar membrane H+-PPase improved the growth characteristics by means of an improved growth rate (11.4%) and elongated logarithmic growth duration. Our study identified a potential method for improving productivity in the use of S. cerevisiae as a cell factory under the harsh conditions present in industry.
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| 9. |
- Vongkampang, Thitiwut, et al.
(författare)
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Characterization of simultaneous uptake of xylose and glucose in Caldicellulosiruptor kronotskyensis for optimal hydrogen production
- 2021
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Ingår i: Biotechnology for Biofuels. - : Springer Science and Business Media LLC. - 1754-6834. ; 14
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Tidskriftsartikel (refereegranskat)abstract
- Background: Caldicellulosiruptor kronotskyensis has gained interest for its ability to grow on various lignocellulosic biomass. The aim of this study was to investigate the growth profles of C. kronotskyensis in the presence of mixtures of glucose–xylose. Recently, we characterized a diauxic-like pattern for C. saccharolyticus on lignocellulosic sugar mixtures. In this study, we aimed to investigate further whether C. kronotskyensis has adapted to uptake glucose in the disaccharide form (cellobiose) rather than the monosaccharide (glucose).Results: Interestingly, growth of C. kronotskyensis on glucose and xylose mixtures did not display diauxic-like growth patterns. Closer investigation revealed that, in contrast to C. saccharolyticus, C. kronotskyensis does not possess a second uptake system for glucose. Both C. saccharolyticus and C. kronotskyensis share the characteristics of preferring xylose over glucose. Growth on xylose was twice as fast (μmax=0.57 h−1) as on glucose (μmax=0.28 h−1). A study of the sugar uptake was made with diferent glucose–xylose ratios to fnd a kinetic relationship between the two sugars for transport into the cell. High concentrations of glucose inhibited xylose uptake and vice versa. The inhibition constants were estimated to be KI,glu=0.01 cmol L−1 and KI,xyl=0.001 cmol L−1, hence glucose uptake was more severely inhibited by xylose uptake. Bioinformatics analysis could not exclude that C. kronotskyensis possesses more than one transporter for glucose. As a next step it was investigated whether glucose uptake by C. kronotskyensis improved in the form of cellobiose. Indeed, cellobiose is taken up faster than glucose; nevertheless, the growth rate on each sugar remained similar.Conclusions: C. kronotskyensis possesses a xylose transporter that might take up glucose at an inferior rate even in the absence of xylose. Alternatively, glucose can be taken up in the form of cellobiose, but growth performance is still inferior to growth on xylose. Therefore, we propose that the catabolism of C. kronotskyensis has adapted more strongly to pentose rather than hexose, thereby having obtained a specifc survival edge in thermophilic lignocellulosic degradation communities.
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| 10. |
- Vongkampang, Thitiwut, et al.
(författare)
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Chitosan flocculation associated with biofilms of C. saccharolyticus and C. owensensis enhances biomass retention in a CSTR
- 2021
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Ingår i: International Journal of Hydrogen Energy. - : Elsevier BV. - 1879-3487 .- 0360-3199. ; 46:41, s. 21338-21347
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Tidskriftsartikel (refereegranskat)abstract
- Cell immobilization and co-culture techniques have gained attention due to its potential to obtain high volumetric hydrogen productivities (QH2). Chitosan retained biomass in the fermentation of co-cultures of Caldicellulosiruptor saccharolyticus and C. owensensis efficiently, up to a maximum dilution rate (D) of 0.9 h−1. Without chitosan, wash out of the co-culture occurred earlier, accompanied with approximately 50% drop in QH2 (D > 0.4 h−1). However, butyl rubber did not show as much potential as carrier material; it did neither improve QH2 nor biomass retention in continuous culture. The population dynamics revealed that C. owensensis was the dominant species (95%) in the presence of chitosan, whereas C. saccharolyticus was the predominant (99%) during cultivation without chitosan. In contrast, the co-culture with rubber as carrier maintained the relative population ratios around 1:1. This study highlighted chitosan as an effective potential carrier for immobilization, thereby paving the way for cost – effective hydrogen production.
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| 11. |
- Vongkampang, Thitiwut, et al.
(författare)
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Immobilization techniques improve volumetric hydrogen productivity of Caldicellulosiruptor species in a modified continuous stirred tank reactor
- 2023
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Ingår i: Biotechnology for Biofuels and Bioproducts. - : Springer Science and Business Media LLC. - 2731-3654. ; 16:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Co-cultures and cell immobilization have been used for retaining biomass in a bioreactor, with the aim to improve the volumetric hydrogen productivity (QH2). Caldicellulosiruptor kronotskyensis is a strong cellulolytic species that possesses tāpirin proteins for attaching on lignocellulosic materials. C. owensensis has its reputation as a biofilm former. It was investigated whether continuous co-cultures of these two species with different types of carriers can improve the QH2. Results: QH2 up to 30 ± 0.2 mmol L−1 h−1 was obtained during pure culture of C. kronotskyensis with combined acrylic fibres and chitosan. In addition, the yield of hydrogen was 2.95 ± 0.1 mol H2 mol−1 sugars at a dilution rate (D) of 0.3 h−1. However, the second-best QH2 26.4 ± 1.9 mmol L−1 h−1 and 25.4 ± 0.6 mmol L−1 h−1 were obtained with a co-culture of C. kronotskyensis and C. owensensis with acrylic fibres only and a pure culture of C. kronotskyensis with acrylic fibres, respectively. Interestingly, the population dynamics revealed that C. kronotskyensis was the dominant species in the biofilm fraction, whereas C. owensensis was the dominant species in the planktonic phase. The highest amount of c-di-GMP (260 ± 27.3 µM at a D of 0.2 h−1) were found with the co-culture of C. kronotskyensis and C. owensensis without a carrier. This could be due to Caldicellulosiruptor producing c-di-GMP as a second messenger for regulation of the biofilms under the high dilution rate (D) to prevent washout. Conclusions: The cell immobilization strategy using a combination of carriers exhibited a promising approach to enhance the QH2. The QH2 obtained during the continuous culture of C. kronotskyensis with combined acrylic fibres and chitosan gave the highest QH2 among the pure culture and mixed cultures of Caldicellulosiruptor in the current study. Moreover, it was the highest QH2 among all cultures of Caldicellulosiruptor species studied so far.
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| 12. |
- Örn, Oliver Englund, et al.
(författare)
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Enhanced Protocatechuic Acid Production From Glucose Using Pseudomonas putida 3-Dehydroshikimate Dehydratase Expressed in a Phenylalanine-Overproducing Mutant of Escherichia coli
- 2021
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Ingår i: Frontiers in Bioengineering and Biotechnology. - : Frontiers Media SA. - 2296-4185. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- Protocatechuic acid (PCA) is a strong antioxidant and is also a potential platform for polymer building blocks like vanillic acid, vanillin, muconic acid, and adipic acid. This report presents a study on PCA production from glucose via the shikimate pathway precursor 3-dehydroshikimate by heterologous expression of a gene encoding 3-dehydroshikimate dehydratase in Escherichia coli. The phenylalanine overproducing E. coli strain, engineered to relieve the allosteric inhibition of 3-deoxy-7-phosphoheptulonate synthase by the aromatic amino acids, was shown to give a higher yield of PCA than the unmodified strain under aerobic conditions. Highest PCA yield of 18 mol% per mol glucose and concentration of 4.2 g/L was obtained at a productivity of 0.079 g/L/h during cultivation in fed-batch mode using a feed of glucose and ammonium salt. Acetate was formed as a major side-product indicating a shift to catabolic metabolism as a result of feedback inhibition of the enzymes including 3-dehydroshikimate dehydratase by PCA when reaching a critical concentration. Indirect measurement of proton motive force by flow cytometry revealed no membrane damage of the cells by PCA, which was thus ruled out as a cause for affecting PCA formation.
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