18 results on '"Sathesh-Prabu C"'
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2. Biolarvicidal activity of extracellular metabolites of the keratinophilic fungus Trichophyton mentagrophytes against arvae of Aedes aegypti — a major vector for Chikungunya and dengue
- Author
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Murugesan, A. G., Sathesh Prabu, C., and Selvakumar, C.
- Published
- 2009
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3. Biodegradation of acrylamide employing free and immobilized cells of Pseudomonas aeruginosa
- Author
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Sathesh Prabu, C. and Thatheyus, A.J.
- Published
- 2007
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4. Oligo- and dsDNA-mediated genome editing using a tetA dual selection system in Escherichia coli.
- Author
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Young Shin Ryu, Sathesh-Prabu Chandran, Kyungchul Kim, and Sung Kuk Lee
- Subjects
Medicine ,Science - Abstract
The ability to precisely and seamlessly modify a target genome is needed for metabolic engineering and synthetic biology techniques aimed at creating potent biosystems. Herein, we report on a promising method in Escherichia coli that relies on the insertion of an optimized tetA dual selection cassette followed by replacement of the same cassette with short, single-stranded DNA (oligos) or long, double-stranded DNA and the isolation of recombinant strains by negative selection using NiCl2. This method could be rapidly and successfully used for genome engineering, including deletions, insertions, replacements, and point mutations, without inactivation of the methyl-directed mismatch repair (MMR) system and plasmid cloning. The method we describe here facilitates positive genome-edited recombinants with selection efficiencies ranging from 57 to 92%. Using our method, we increased lycopene production (3.4-fold) by replacing the ribosome binding site (RBS) of the rate-limiting gene (dxs) in the 1-deoxy-D-xylulose-5-phosphate (DXP) biosynthesis pathway with a strong RBS. Thus, this method could be used to achieve scarless, proficient, and targeted genome editing for engineering E. coli strains. more...
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- 2017
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5. Simultaneous utilization of glucose and xylose by metabolically engineered Pseudomonas putida for the production of 3-hydroxypropionic acid.
- Author
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Tiwari R, Sathesh-Prabu C, Kim Y, and Kuk Lee S
- Subjects
- Glucose metabolism, Fermentation, Metabolic Engineering methods, Xylose metabolism, Pseudomonas putida genetics, Pseudomonas putida metabolism, Lactic Acid analogs & derivatives
- Abstract
Pseudomonas putida,a robust candidate for lignocellulosicbiomass-based biorefineries, encounters challenges in metabolizing xylose. In this study, Weimberg pathway was introduced intoP. putidaEM42 under a xylose-inducible promoter, resulting in slow cell growth (0.05 h
-1 ) on xylose.Through adaptive laboratory evolution, an evolved strain exhibited highly enhanced growth on xylose (0.36 h-1 ), comparable to that on glucose (0.39 h-1 ). Whole genome sequencing identified four mutations, with two key mutations located inPP3380andPP2219. Reverse-engineered strain 8EM42_Xyl, harboring these two mutations, showed enhanced growth on xylose but co-utilizing glucose and xylose at a rate of 0.3 g/L/h. Furthermore, 8EM42_Xyl was employed for 3-hydroxypropionic acid (3HP) production from glucose and xylose by expressing malonyl-CoA reductase and acetyl-CoA carboxylase, yielding 29 g/L in fed-batch fermentation. Moreover, the engineered strain exhibited promising performance in 3HP production from empty palm fruit bunch hydrolysate, demonstrating its potential as a promising cell factory forbiorefineries., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.) more...- Published
- 2024
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6. Comparative Genomic Analysis and BTEX Degradation Pathways of a Thermotolerant Cupriavidus cauae PHS1.
- Author
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Sathesh-Prabu C, Woo J, Kim Y, Kim SM, Lee SB, Jeon CO, Kim D, and Lee SK
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- Toluene, Xylenes metabolism, Biodegradation, Environmental, Benzene Derivatives metabolism, Genomics, Benzene metabolism, Cupriavidus genetics, Cupriavidus metabolism
- Abstract
Volatile organic compounds such as benzene, toluene, ethylbenzene, and isomers of xylenes (BTEX) constitute a group of monoaromatic compounds that are found in petroleum and have been classified as priority pollutants. In this study, based on its newly sequenced genome, we reclassified the previously identified BTEX-degrading thermotolerant strain Ralstonia sp. PHS1 as Cupriavidus cauae PHS1. Also presented are the complete genome sequence of C. cauae PHS1, its annotation, species delineation, and a comparative analysis of the BTEX-degrading gene cluster. Moreover, we cloned and characterized the BTEX-degrading pathway genes in C. cauae PHS1, the BTEX-degrading gene cluster of which consists of two monooxygenases and meta-cleavage genes. A genome-wide investigation of the PHS1 coding sequence and the experimentally confirmed regioselectivity of the toluene monooxygenases and catechol 2,3-dioxygenase allowed us to reconstruct the BTEX degradation pathway. The degradation of BTEX begins with aromatic ring hydroxylation, followed by ring cleavage, and eventually enters the core carbon metabolism. The information provided here on the genome and BTEX-degrading pathway of the thermotolerant strain C. cauae PHS1 could be useful in constructing an efficient production host. more...
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- 2023
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7. Combinatorial Metabolic Engineering Strategies for the Enhanced Production of Free Fatty Acids in Escherichia coli .
- Author
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Park WS, Shin KS, Jung HW, Lee Y, Sathesh-Prabu C, and Lee SK
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- Fatty Acids, Nonesterified metabolism, Fatty Acids chemistry, Malonyl Coenzyme A metabolism, Metabolic Engineering, Escherichia coli genetics
- Abstract
In this study, we evaluated the effects of several metabolic engineering strategies in a systematic and combinatorial manner to enhance the free fatty acid (FFA) production in Escherichia coli . The strategies included (i) overexpression of mutant thioesterase I ('TesA
R64C ) to efficiently release the FFAs from fatty acyl-ACP; (ii) coexpression of global regulatory protein FadR; (iii) heterologous expression of methylmalonyl-CoA carboxyltransferase and phosphoenolpyruvate carboxylase to synthesize fatty acid precursor molecule malonyl-CoA; and (iv) disruption of genes associated with membrane proteins (GusC, MdlA, and EnvR) to improve the cellular state and export the FFAs outside the cell. The synergistic effects of these genetic modifications in strain SBF50 yielded 7.2 ± 0.11 g/L FFAs at the shake flask level. In fed-batch cultivation under nitrogen-limiting conditions, strain SBF50 produced 33.6 ± 0.02 g/L FFAs with a productivity of 0.7 g/L/h from glucose, which is the maximum titer reported in E. coli to date. Combinatorial metabolic engineering approaches can prove to be highly useful for the large-scale production of FA-derived chemicals and fuels. more...- Published
- 2022
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8. Bioproduction of propionic acid using levulinic acid by engineered Pseudomonas putida .
- Author
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Tiwari R, Sathesh-Prabu C, and Lee SK
- Abstract
The present study elaborates on the propionic acid (PA) production by the well-known microbial cell factory Pseudomonas putida EM42 and its capacity to utilize biomass-derived levulinic acid (LA). Primarily, the P. putida EM42 strain was engineered to produce PA by deleting the methylcitrate synthase (PrpC) and propionyl-CoA synthase (PrpE) genes. Subsequently, a LA-inducible expression system was employed to express yciA (encoding thioesterase) from Haemophilus influenzae and ygfH (encoding propionyl-CoA: succinate CoA transferase) from Escherichia coli to improve the PA production by up to 10-fold under flask scale cultivation. The engineered P. putida EM42:ΔCE: yciA:ygfH was used to optimize the bioprocess to further improve the PA production titer. Moreover, the fed-batch fermentation performed under optimized conditions in a 5 L bioreactor resulted in the titer, productivity, and molar yield for PA production of 26.8 g/L, 0.3 g/L/h, and 83%, respectively. This study, thus, successfully explored the LA catabolic pathway of P. putida as an alternative route for the sustainable and industrial production of PA from LA., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tiwari, Sathesh-Prabu and Lee.) more...
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- 2022
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9. Substrate-inducible and antibiotic-free high-level 4-hydroxyvaleric acid production in engineered Escherichia coli .
- Author
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Sathesh-Prabu C, Tiwari R, and Lee SK
- Abstract
In this study, we developed a levulinic acid (LA)-inducible and antibiotic-free plasmid system mediated by HpdR/P
hpdH and infA -complementation to produce 4-hydroxyvaleric acid (4-HV) from LA in an engineered Escherichia coli strain. The system was efficiently induced by the addition of the LA substrate and resulted in tight dose-dependent control and fine-tuning of gene expression. By engineering the 5' untranslated region (UTR) of hpdR mRNA, the gene expression of green fluorescent protein (GFP) increased by at least two-fold under the hpdH promoter. Furthermore, by evaluating the robustness and plasmid stability of the proposed system, the engineered strain, IRV750f , expressing the engineered 3-hydroxybutyrate dehydrogenase (3HBDH∗ ) and formate dehydrogenase ( Cb FDH), produced 82 g/L of 4-HV from LA, with a productivity of 3.4 g/L/h and molar conversion of 92% in the fed-batch cultivation (5 L fermenter) without the addition of antibiotics or external inducers. Overall, the reported system was highly beneficial for the large-scale and cost-effective microbial production of value-added products and bulk chemicals from the renewable substrate, LA., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Sathesh-Prabu, Tiwari and Lee.) more...- Published
- 2022
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10. Enhanced production of nonanedioic acid from nonanoic acid by engineered Escherichia coli.
- Author
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Lee Y, Sathesh-Prabu C, Kwak GH, Bang I, Jung HW, Kim D, and Lee SK
- Subjects
- Dicarboxylic Acids metabolism, Escherichia coli genetics, Escherichia coli metabolism, Fatty Acids
- Abstract
In this study, whole-cell biotransformation was conducted to produce nonanedioic acid from nonanoic acid by expressing the alkane hydroxylating system (AlkBGT) from Pseudomonas putida GPo1 in Escherichia coli. Following adaptive laboratory evolution, an efficient E. coli mutant strain, designated as MRE, was successfully obtained, demonstrating the fastest growth (27-fold higher) on nonanoic acid as the sole carbon source compared to the wild-type strain. Additionally, the MRE strain was engineered to block nonanoic acid degradation by deleting fadE. The resulting strain exhibited a 12.8-fold increase in nonanedioic acid production compared to the wild-type strain. Six mutations in acrR, P
crp , dppA, PfadD , e14, and yeaR were identified in the mutant MRE strain, which was characterized using genomic modifications and RNA-sequencing. The acquired mutations were found to be beneficial for rapid growth and nonanedioic acid production., (© 2021 Wiley-VCH GmbH.) more...- Published
- 2022
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11. Levulinic Acid-Inducible and Tunable Gene Expression System for Methylorubrum extorquens .
- Author
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Sathesh-Prabu C, Ryu YS, and Lee SK
- Abstract
Methylorubrum extorquens AM1 is an efficient platform strain possessing biotechnological potential in formate- and methanol-based single carbon (C1) bioeconomy. Constitutive expression or costly chemical-inducible expression systems are not always desirable. Here, several glucose-, xylose-, and levulinic acid (LA)-inducible promoter systems were assessed for the induction of green fluorescent protein (GFP) as a reporter protein. Among them, the LA-inducible gene expression system (HpdR/P
hpdH ) showed a strong expression of GFP (51-fold) compared to the control. The system was induced even at a low concentration of LA (0.1 mM). The fluorescence intensity increased with increasing concentrations of LA up to 20 mM. The system was tunable and tightly controlled with meager basal expression. The maximum GFP yield obtained using the system was 42 mg/g biomass, representing 10% of the total protein content. The efficiency of the proposed system was nearly equivalent (90%-100%) to that of the widely used strong promoters such as PmxaF and PL/O4 . The HpdR/PhpdH system worked equally efficiently in five different strains of M. extorquens . LA is a low-cost, renewable, and sustainable platform chemical that can be used to generate a wide range of products. Hence, the reported system in potent strains of M. extorquens is highly beneficial in the C1-biorefinery industry to produce value-added products and bulk chemicals ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Sathesh-Prabu, Ryu and Lee.) more...- Published
- 2021
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12. Inducible and tunable gene expression systems for Pseudomonas putida KT2440.
- Author
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Sathesh-Prabu C, Tiwari R, Kim D, and Lee SK
- Subjects
- Flow Cytometry, Gene Expression, Genes, Reporter, Glucose metabolism, Glucose pharmacology, Lactic Acid analogs & derivatives, Lactic Acid metabolism, Lactic Acid pharmacology, Promoter Regions, Genetic, Pseudomonas putida metabolism, Recombinant Proteins genetics, Gene Expression Regulation, Bacterial drug effects, Genetic Engineering, Genetic Vectors genetics, Pseudomonas putida genetics
- Abstract
Inducible and tunable expression systems are essential for the microbial production of biochemicals. Five different carbon source- and substrate-inducible promoter systems were developed and further evaluated in Pseudomonas putida KT2440 by analyzing the expression of green fluorescent protein (GFP) as a reporter protein. These systems can be induced by low-cost compounds such as glucose, 3-hydroxypropionic acid (3HP), levulinic acid (LA), and xylose. 3HP-inducible HpdR/P
hpdH was also efficiently induced by LA. LvaR/PlvaA and XutR/PxutA systems were induced even at low concentrations of LA (0.1 mM) and xylose (0.5 mM), respectively. Glucose-inducible HexR/Pzwf1 showed weak GFP expression. These inducer agents can be used as potent starting materials for both cell growth and the production of a wide range of biochemicals. The efficiency of the reported systems was comparable to that of conventional chemical-inducible systems. Hence, the newly investigated promoter systems are highly useful for the expression of target genes in the widely used synthetic biology chassis P. putida KT2440 for industrial and medical applications., (© 2021. The Author(s).) more...- Published
- 2021
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13. Metabolic engineering of Escherichia coli for 2,3-butanediol production from cellulosic biomass by using glucose-inducible gene expression system.
- Author
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Sathesh-Prabu C, Kim D, and Lee SK
- Subjects
- Biomass, Butylene Glycols, Fermentation, Glucose, Escherichia coli, Metabolic Engineering
- Abstract
A glucose-inducible gene expression system has been developed using HexR-P
zwf1 of Pseudomonas putida to induce the metabolic pathways. Since the system is controlled by an Entner-Doudoroff pathway (EDP) intermediate, the EDP of Escherichia coli was activated by deleting pfkA and gntR genes. Growth experiment with green fluorescent protein as a reporter indicated that the induction of this system was tightly controlled over a wide range of glucose in E. coli without adding any inducer. 2,3-butanediol (BDO) synthetic pathway genes were expressed by this system in the pfkA-gntR-deleted strain. The resultant engineered strain harbouring this system efficiently produced BDO with a 71% increased titer than the control strain. The strain was also able to produce BDO from a mixture of glucose and xylose which is comparable to glucose alone. Further, the strain produced 11 g/L of BDO at a yield of 0.48 g/g from the hydrolysate of empty palm fruit bunches. This system can also be applied in many other bio-production processes from lignocellulosic biomass., (Copyright © 2020 Elsevier Ltd. All rights reserved.) more...- Published
- 2020
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14. High-Level Production of 4-Hydroxyvalerate from Levulinic Acid via Whole-Cell Biotransformation Decoupled from Cell Metabolism.
- Author
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Kim D, Sathesh-Prabu C, JooYeon Y, and Lee SK
- Subjects
- Adenosine Triphosphate metabolism, Biotransformation, Escherichia coli genetics, Fermentation, Metabolic Engineering, Escherichia coli metabolism, Levulinic Acids metabolism, Valerates metabolism
- Abstract
γ-Hydroxyvalerate (4HV) is an important monomer used to produce various valuable polymers and products. In this study, an engineered 3-hydroxybutyrate dehydrogenase that can convert levulinic acid (LA) into 4HV was co-expressed with a cofactor (NADH) regeneration system mediated by an NAD
+ -dependent formate dehydrogenase ( Cb FDH) in the Escherichia coli strain, MG1655. The resulting strain produced 23-fold more 4HV in a shake flask. The 4HV production was not dependent on ATP and required low aeration; all of these are considered beneficial characteristics for the production of target compounds, especially at an industrial scale. Under optimized conditions in a 5 L fermenter, the titer, productivity, and molar conversion efficiency for 4HV reached 100 g/L, 4.2 g/L/h, and 92%, respectively. Our system could prove to be a promising method for the large-scale production of 4HV from LA at low-cost and using a renewable biomass source. more...- Published
- 2019
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15. Engineering the lva operon and Optimization of Culture Conditions for Enhanced Production of 4-Hydroxyvalerate from Levulinic Acid in Pseudomonas putida KT2440.
- Author
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Sathesh-Prabu C and Lee SK
- Subjects
- Coenzyme A metabolism, Escherichia coli genetics, Gene Expression, Pseudomonas putida metabolism, Recombinant Proteins, Thiolester Hydrolases genetics, Genetic Engineering methods, Levulinic Acids metabolism, Operon genetics, Pseudomonas putida genetics, Valerates metabolism
- Abstract
Monomeric 4-hydroxyvalerate is a versatile chemical used to produce various commodities and fine chemicals. In the present study, the lvaAB gene was deleted from the lva operon in Pseudomonas putida KT2440 and tesB, obtained from Escherichia coli, was overexpressed under the control of the lva operon system, which is induced by the substrate levulinic acid and the product 4-hydroxyvalerate to produce 4-hydroxyvalerate from levulinic acid. The lvaAB-deleted strain showed almost complete conversion of levulinic acid to 4-hydroxyvalerate, compared with 24% conversion in the wild-type strain. In addition, under optimized culture conditions, the final engineered strain produced a maximum of 50 g/L 4-hydroxyvalerate with 97% conversion from levulinic acid. The system presented here could be applied to produce high titers of 4-hydroxyvalerate in a cost-effective manner at a large scale from renewable cellulosic biomass. more...
- Published
- 2019
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16. Enhancement of α,ω-Dicarboxylic Acid Production by the Expression of Xylose Reductase for Refactoring Redox Cofactor Regeneration.
- Author
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Sathesh-Prabu C and Lee SK
- Subjects
- Alcohol Dehydrogenase genetics, Alcohol Dehydrogenase metabolism, Aldehyde Reductase metabolism, Escherichia coli enzymology, Escherichia coli genetics, Glucose metabolism, Mixed Function Oxygenases chemistry, Mixed Function Oxygenases genetics, Mixed Function Oxygenases metabolism, NAD metabolism, NADP metabolism, Oxidation-Reduction, Aldehyde Reductase genetics, Dicarboxylic Acids metabolism, Escherichia coli metabolism, Xylose metabolism
- Abstract
The production of α,ω-dicarboxylic acids (DCAs) by whole-cell biocatalysis is often limited by cofactor regeneration. Here, ω-oxidation pathway genes (monooxygenase, alcohol dehydrogenase, and aldehyde dehydrogenase) were coexpressed with a xylose reductase (XR) gene to regenerate cofactors in an engineered Escherichia coli strain that cometabolizes glucose and xylose. The resulting strain exhibited a 180% increase in DCA production compared with the control strain without XR, and produced xylitol in the presence of xylose. Expression of monooxygenase and XR without other ω-oxidation pathway genes resulted in an additional increase in tetradecanedioic acid concentration and a substrate conversion of 95%, which was 198% higher than that associated with the control strain. The expression of XR helped the system to regenerate and balance the cofactors thereby achieving maximum substrate conversion efficiency. It could serve as an efficient platform for the industrial production of α,ω-DCAs. more...
- Published
- 2018
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17. Genetic Variability and Proteome Profiling of a Radiation Induced Cellulase Mutant Mushroom Pleurotus florida.
- Author
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Sathesh-Prabu C and Lee YK
- Subjects
- Amplified Fragment Length Polymorphism Analysis, Fungal Proteins genetics, Pleurotus metabolism, Polymorphism, Restriction Fragment Length, Genetic Variation, Pleurotus genetics
- Abstract
We report the genetic similarity changes between a mutant mushroom (Pleurotus florida, designated as PfCM4) having increased cellulolytic activity developed through radiation mutagenesis and its wild type by amplified fragment length polymorphism (AFLP). On average, 23 AFLP fragments were amplified per primer combination, and a total of 286 polymorphic fragments (78.57% polymorphism) with maximal fragment length of 1365 base pairs (bp) were obtained. The genetic similarity between wild type and PfCM4 was found to be 22.30%. In addition, mycelial and secreted protein profiling by 2D-PAGE showed at least three and five different protein spots in the range of 25 kD to 100 kD, respectively, in PfCM4. It seems that the variation in genetic similarity and different expression of both mycelial and secreted proteins in PfCM4 in comparison to the wild type could likely be correlated with its increased cellulolytic activity effected by the irradiation. more...
- Published
- 2016
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18. Production of Long-Chain α,ω-Dicarboxylic Acids by Engineered Escherichia coli from Renewable Fatty Acids and Plant Oils.
- Author
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Sathesh-Prabu C and Lee SK
- Subjects
- Aminolevulinic Acid pharmacology, Cloning, Molecular, Coconut Oil, Cytochrome P-450 Enzyme System genetics, Escherichia coli genetics, Escherichia coli metabolism, Gene Expression, Hydrogen Peroxide metabolism, Oxidation-Reduction, Recombinant Proteins genetics, Recombinant Proteins metabolism, Thiourea pharmacology, Cytochrome P-450 Enzyme System metabolism, Dicarboxylic Acids metabolism, Escherichia coli enzymology, Fatty Acids metabolism, Plant Oils metabolism
- Abstract
Long-chain α,ω-dicarboxylic acids (LDCAs, ≥ C12) are widely used as a raw material for preparing various commodities and polymers. In this study, a CYP450-monooxygenase-mediated ω-oxidation pathway system with high ω-regioselectivity was heterologously expressed in Escherichia coli to produce DCAs from fatty acids. The resulting engineered E. coli produced a maximum of 41 mg/L of C12 DCA and 163 mg/L of C14 DCA from fatty acids (1 g/L), following 20 h of whole cell biotransformation. Addition of a heme precursor and the hydroxyl radical scavenger, thiourea, increased product concentration (159 mg/L of C12 DCA and 410 mg/L of C14 DCA) in a shorter culture duration than that of the corresponding controls. DCAs of various chain lengths were synthesized from coconut oil hydrolysate using the engineered E. coli. This novel synthetic biocatalytic system could be applied to produce high value DCAs in a cost-effective manner from renewable plant oils. more...
- Published
- 2015
- Full Text
- View/download PDF
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