Metabolic engineering of Escherichia coli for the production of phenylalanine and related compounds

Doroshenko, V. G.; Livshits, Vitaliy A.; Airich, L. G.; Шмагина, И.С.; Savrasova, Ekaterina A.; Ovsienko, M. V.; Mashko, Sergey V.

doi:10.1134/s0003683815070017

Cited by 7 publications

(3 citation statements)

References 88 publications

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“…Various aromatic compounds can be produced from glucose by use of the shikimate pathway (30)(31)(32)(33)(34)(35)(36). For example, introduction of genes encoding the PP decarboxylase gene from Azospirillum brasilense enabled Phe-producing E. coli to become 2-phenylethanol-and phenylacetic acid-producing strains, and tyrosol-and 4-hydroxyphenylacetic acid-producing strains could be obtained by introduction of these genes into a Tyr-producing strain (36).…”

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confidence: 99%

Chromosome Engineering To Generate Plasmid-Free Phenylalanine- and Tyrosine-Overproducing Escherichia coli Strains That Can Be Applied in the Generation of Aromatic-Compound-Producing Bacteria

Koma

Kishida

Yoshida

et al. 2020

Appl Environ Microbiol

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ABSTRACT Many phenylalanine- and tyrosine-producing strains have used plasmid-based overexpression of pathway genes. The resulting strains achieved high titers and yields of phenylalanine and tyrosine. Chromosomally engineered, plasmid-free producers have shown lower titers and yields than plasmid-based strains, but the former are advantageous in terms of cultivation cost and public health/environmental risk. Therefore, we engineered here the Escherichia coli chromosome to create superior phenylalanine- and tyrosine-overproducing strains that did not depend on plasmid-based expression. Integration into the E. coli chromosome of two central metabolic pathway genes (ppsA and tktA) and eight shikimate pathway genes (aroA, aroB, aroC, aroD, aroE, aroGfbr, aroL, and pheAfbr), controlled by the T7lac promoter, resulted in excellent titers and yields of phenylalanine; the superscript “fbr” indicates that the enzyme encoded by the gene was feedback resistant. The generated strain could be changed to be a superior tyrosine-producing strain by replacing pheAfbr with tyrAfbr. A rational approach revealed that integration of seven genes (ppsA, tktA, aroA, aroB, aroC, aroGfbr, and pheAfbr) was necessary as the minimum gene set for high-yield phenylalanine production in E. coli MG1655 (tyrR, adhE, ldhA, pykF, pflDC, and ascF deletant). The phenylalanine- and tyrosine-producing strains were further applied to generate phenyllactic acid-, 4-hydroxyphenyllactic acid-, tyramine-, and tyrosol-producing strains; yield of these aromatic compounds increased proportionally to the increase in phenylalanine and tyrosine yields. IMPORTANCE Plasmid-free strains for aromatic compound production are desired in the aspect of industrial application. However, the yields of phenylalanine and tyrosine have been considerably lower in plasmid-free strains than in plasmid-based strains. The significance of this research is that we succeeded in generating superior plasmid-free phenylalanine- and tyrosine-producing strains by engineering the E. coli chromosome, which was comparable to that in plasmid-based strains. The generated strains have a potential to generate superior strains for the production of aromatic compounds. Actually, we demonstrated that four kinds of aromatic compounds could be produced from glucose with high yields (e.g., 0.28 g tyrosol/g glucose).

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confidence: 99%

Chromosome Engineering To Generate Plasmid-Free Phenylalanine- and Tyrosine-Overproducing Escherichia coli Strains That Can Be Applied in the Generation of Aromatic-Compound-Producing Bacteria

Koma

Kishida

Yoshida

et al. 2020

Appl Environ Microbiol

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“…Furthermore, the transcriptional repressor tyrR was deleted as its inactivation is known to enhance the expression of aromatic biosynthesis genes, like aroG, tyrB, aroP, tyrA , and aroL (Pittard, 1996; Bongaerts et al, 2001; Pittard et al, 2005; Salgado et al, 2006). The beneficial effect of a tyrR deletion had been already shown for the biosynthesis of aromatic amino acids like l -Tyr (Lutke-Eversloh and Stephanopoulos, 2007), l -Phe (Doroshenko et al, 2015), l -DOPA (Munoz et al, 2011; Das et al, 2018), and recently l -PAPA (Mohammadi Nargesi et al, 2018). The cultivation of FUS4BCR/pC53BCA or/pC53BCAF increased the PAPE or 4-APA titer up to 27 or 17% than FUS4BC/pC53BCA or/pC53BCAF, respectively (Figure 3).…”

Section: Discussionmentioning

confidence: 88%

Metabolic Engineering of Escherichia coli for para-Amino-Phenylethanol and para-Amino-Phenylacetic Acid Biosynthesis

Nargesi

Sprenger

Youn

2019

Front. Bioeng. Biotechnol.

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Aromatic amines are an important class of chemicals which are used as building blocks for the synthesis of polymers and pharmaceuticals. In this study we establish a de novo pathway for the biosynthesis of the aromatic amines para-amino-phenylethanol (PAPE) and para-amino-phenylacetic acid (4-APA) in Escherichia coli. We combined a synthetic para-amino-l-phenylalanine pathway with the fungal Ehrlich pathway. Therefore, we overexpressed the heterologous genes encoding 4-amino-4-deoxychorismate synthase (pabAB from Corynebacterium glutamicum), 4-amino-4-deoxychorismate mutase and 4-amino-4-deoxyprephenate dehydrogenase (papB and papC from Streptomyces venezuelae) and ThDP-dependent keto-acid decarboxylase (aro10 from Saccharomyces cerevisiae) in E. coli. The resulting para-amino-phenylacetaldehyde either was reduced to PAPE or oxidized to 4-APA. The wild type strain E. coli LJ110 with a plasmid carrying these four genes produced (in shake flask cultures) 11 ± 1.5 mg l−1 of PAPE from glucose (4.5 g l−1). By the additional cloning and expression of feaB (phenylacetaldehyde dehydrogenase from E. coli) 36 ± 5 mg l−1 of 4-APA were obtained from 4.5 g l−1 glucose. Competing reactions, such as the genes for aminotransferases (aspC and tyrB) or for biosynthesis of L-phenylalanine and L-tyrosine (pheA, tyrA) and for the regulator TyrR were removed. Additionally, the E. coli genes aroFBL were cloned and expressed from a second plasmid. The best producer strains of E. coli showed improved formation of PAPE and 4-APA, respectively. Plasmid-borne expression of an aldehyde reductase (yahK from E. coli) gave best values for PAPE production, whereas feaB-overexpression led to best values for 4-APA. In fed-batch cultivation, the best producer strains achieved 2.5 ± 0.15 g l−1 of PAPE from glucose (11% C mol mol-1 glucose) and 3.4 ± 0.3 g l−1 of 4-APA (17% C mol mol−1 glucose), respectively which are the highest values for recombinant strains reported so far.

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“…Entre los diversos microorganismos utilizados en la producción de L-Fe por la ruta fermentativa se encentran el Brevibacterium lactofermentum (Ito et al, 1990), Corynebacterium glutamicum (Liu & Liao, 1994), Bacillus subtilis (McEvoy & Joyce, 1974), y la bacteria Escherichia coli recombinante (Doroshenko et al, 2015), la cual ha atraído un gran interés debido a su elevada tasa de crecimiento y sus características fisiológicas bien conocidas (Yuan et al, 2015). Varios estudios se han llevado a cabo con respecto a la ingeniería metabólica y control del proceso de fermentación utilizando E. coli recombinante para la producción de L-Fe ) (Báez-Viveros et al, 2007 (Yuan et al, 2015) (Liu et al, 2018) (Wu et al, 2019).…”

Section: Introductionunclassified

Simulación del proceso de producción de L-fenilalanina por la ruta fermentativa utilizando el simulador SuperPro Designer®

Sánchez

Ranero-González

Pérez-Sánchez³

et al. 2021

reveia

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La L-fenilalanina (L-Fe) es uno de los ocho aminoácidos esenciales para el cuerpo humano. En el presente trabajo se efectuó la simulación del proceso de producción de la L-Fe por la ruta fermentativa mediante el simulador SuperPro Designerâ, con el fin de conocer sus indicadores de rentabilidad más importantes bajo las condiciones económicas actuales de Cuba. También se efectuó un estudio de sensibilidad con el objetivo de saber a partir de qué valor del parámetro precio de venta unitario del frasco de L-Fe se comienza a obtener un valor positivo del indicador Valor Actual Neto (VAN). Se obtuvo un margen bruto de 70,15 %, un costo unitario de producción de USD $ 66,75 por frasco y un retorno de la inversión de 38,92 %. A partir de un valor del precio de venta unitario del frasco de L-Fe de USD $ 115,3 empieza a ser rentable la planta de producción. El proceso de producción de L-Fe puede considerarse de rentable y factible desde el punto de vista técnico-económico atendiendo a los resultados obtenidos de VAN (USD $ 14 040 000), Tasa Interna de Retorno (49,14 %) y Período de Recuperación de la Inversión (2,57 años).

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Metabolic engineering of Escherichia coli for the production of phenylalanine and related compounds

Cited by 7 publications

References 88 publications

Chromosome Engineering To Generate Plasmid-Free Phenylalanine- and Tyrosine-Overproducing Escherichia coli Strains That Can Be Applied in the Generation of Aromatic-Compound-Producing Bacteria

Chromosome Engineering To Generate Plasmid-Free Phenylalanine- and Tyrosine-Overproducing Escherichia coli Strains That Can Be Applied in the Generation of Aromatic-Compound-Producing Bacteria

Metabolic Engineering of Escherichia coli for para-Amino-Phenylethanol and para-Amino-Phenylacetic Acid Biosynthesis

Simulación del proceso de producción de L-fenilalanina por la ruta fermentativa utilizando el simulador SuperPro Designer®

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