Metabolic engineering for improving anthranilate synthesis from glucose in Escherichia coli

Balderas‐Hernández, Víctor E.; Sabido-Ramos, Andrea; Silva, Patrı́cia; Cabrera-Valladares, Natividad; Hernández-Chávez, Georgina; Báez-Viveros, José Luis; Martı́nez, Alfredo; Bolívar, Francisco; Gosset, Guillermo

doi:10.1186/1475-2859-8-19

Cited by 91 publications

(130 citation statements)

References 30 publications

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“…Biosynthesis of tryptophan starts with the condensation of D-erythrose 4-phosphate (E4P) and phosphoenolpyruvate (PEP) to form 3-deoxy-D-arabinoheptulosonate 7-phosphate (DAHP). After several more steps, chorismate is synthesized, from which the shikimate pathway branches to the biosynthesis of tryptophan, tyrosine, and phenylalanine (39). Anthranilate is the first dedicated intermediate in the tryptophan branch pathway.…”

Section: Resultsmentioning

confidence: 99%

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A Novel Muconic Acid Biosynthesis Approach by Shunting Tryptophan Biosynthesis via Anthranilate

Sun

Lin

Huang

et al. 2013

Appl Environ Microbiol

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cMuconic acid is the synthetic precursor of adipic acid, and the latter is an important platform chemical that can be used for the production of nylon-6,6 and polyurethane. Currently, the production of adipic acid relies mainly on chemical processes utilizing petrochemicals, such as benzene, which are generally considered environmentally unfriendly and nonrenewable, as starting materials. Microbial synthesis from renewable carbon sources provides a promising alternative under the circumstance of petroleum depletion and environment deterioration. Here we devised a novel artificial pathway in Escherichia coli for the biosynthesis of muconic acid, in which anthranilate, the first intermediate in the tryptophan biosynthetic branch, was converted to catechol and muconic acid by anthranilate 1,2-dioxygenase (ADO) and catechol 1,2-dioxygenase (CDO), sequentially and respectively. First, screening for efficient ADO and CDO from different microbial species enabled the production of gram-per-liter level muconic acid from supplemented anthranilate in 5 h. To further achieve the biosynthesis of muconic acid from simple carbon sources, anthranilate overproducers were constructed by overexpressing the key enzymes in the shikimate pathway and blocking tryptophan biosynthesis. In addition, we found that introduction of a strengthened glutamine regeneration system by overexpressing glutamine synthase significantly improved anthranilate production. Finally, the engineered E. coli strain carrying the full pathway produced 389.96 ؎ 12.46 mg/liter muconic acid from simple carbon sources in shake flask experiments, a result which demonstrates scale-up potential for microbial production of muconic acid.

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Section: Resultsmentioning

confidence: 99%

“…Previous approaches to enhance the production of anthranilate and tryptophan focused mainly on the key enzymes in the pathway itself (6,8,39). The role of the auxiliary pathway, such as the glutamine regeneration system, had been ignored.…”

Section: Discussionmentioning

confidence: 99%

A Novel Muconic Acid Biosynthesis Approach by Shunting Tryptophan Biosynthesis via Anthranilate

Sun

Lin

Huang

et al. 2013

Appl Environ Microbiol

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“…Strains W3110 tyrR and VH33 tyrR are tyrR ¡ derivatives of W3110 and VH33, respectively. Plasmid pJLBaroG fbr tktA has a replication origin from pACYC 184, it carries gene aroG fbr encoding a feedback resistant version of DAHP synthase and gene tktA encoding transketolase, both from E. coli [3]. The aroG fbr gene is transcribed from the lacUV5 promoter that is regulated by the LacI repressor and can be induced by addition of isopropyl--D-thiogalactopyranoside (IPTG).…”

Section: Methodsmentioning

confidence: 99%

Metabolic engineering of Escherichia coli for improving l-3,4-dihydroxyphenylalanine (l-DOPA) synthesis from glucose

Muñoz

Hernández-Chávez

Anda

et al. 2011

J Ind Microbiol Biotechnol

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L-3,4-dihydroxyphenylalanine (L-DOPA) is an aromatic compound employed for the treatment of Parkinson's disease. Metabolic engineering was applied to generate Escherichia coli strains for the production of L-DOPA from glucose by modifying the phosphoenolpyruvate:sugar phosphotransferase system (PTS) and aromatic biosynthetic pathways. Carbon flow was directed to the biosynthesis of L-tyrosine (L-Tyr), an L-DOPA precursor, by transforming strains with compatible plasmids carrying genes encoding a feedback-inhibition resistant version of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase, transketolase, the chorismate mutase domain from chorismate mutase-prephenate dehydratase from E. coli and cyclohexadienyl dehydrogenase from Zymomonas mobilis. The effects on L-Tyr production of PTS inactivation (PTS(-) gluc(+) phenotype), as well as inactivation of the regulatory protein TyrR, were evaluated. PTS inactivation caused a threefold increase in the specific rate of L-Tyr production (q( L-Tyr)), whereas inactivation of TyrR caused 1.7- and 1.9-fold increases in q( L-Tyr) in the PTS(+) and the PTS(-) gluc(+) strains, respectively. An 8.6-fold increase in L-Tyr yield from glucose was observed in the PTS(-) gluc(+) tyrR (-) strain. Expression of hpaBC genes encoding the enzyme 4-hydroxyphenylacetate 3-hydroxylase from E. coli W in the strains modified for L-Tyr production caused the synthesis of L-DOPA. One of such strains, having the PTS(-) gluc(+) tyrR (-) phenotype, displayed the best production parameters in minimal medium, with a specific rate of L-DOPA production of 13.6 mg/g/h, L-DOPA yield from glucose of 51.7 mg/g and a final L-DOPA titer of 320 mg/l. In a batch fermentor culture in rich medium this strain produced 1.51 g/l of L-DOPA in 50 h.

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“…No hints for enhanced carbon flux via PEP carboxylase to oxaloacetate have been shown during PTS-independent growth and L-lysine production, since PEP carboxylase activity was comparable to that of the parental strain. In E. coli, strains engineered for PTSindependent glucose uptake showed the improved production of aromatic compounds such as phenylalanine, shikimate, and anthranilate (3,7,15,17,18,26,35,70) as a consequence of reduced pyruvate and increased PEP levels in the cells. For example, the expression of the genes for galactose permease and glucokinase in PTS-negative E. coli strains bypassed PEP usage for glucose phosphorylation and resulted in a higher yield of 3-deoxy-D-arabinoheptulosonate-7-phosphate, which was assumed to be a consequence of an increased level of PEP, the immediate precursor of 3-deoxy-D-arabinoheptulosonate-7-phosphate (18,21).…”

Section: Discussionmentioning

confidence: 99%

Phosphotransferase System-Independent Glucose Utilization in Corynebacterium glutamicum by Inositol Permeases and Glucokinases

Lindner

Seibold

Henrich

et al. 2011

Appl Environ Microbiol

101

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Phosphoenolpyruvate-dependent glucose phosphorylation via the phosphotransferase system (PTS) is the major path of glucose uptake in Corynebacterium glutamicum, but some growth from glucose is retained in the absence of the PTS. The growth defect of a deletion mutant lacking the general PTS component HPr in glucose medium could be overcome by suppressor mutations leading to the high expression of inositol utilization genes or by the addition of inositol to the growth medium if a glucokinase is overproduced simultaneously. PTSindependent glucose uptake was shown to require at least one of the inositol transporters IolT1 and IolT2 as a mutant lacking IolT1, IolT2, and the PTS component HPr could not grow with glucose as the sole carbon source. Efficient glucose utilization in the absence of the PTS necessitated the overexpression of a glucokinase gene in addition to either iolT1 or iolT2. IolT1 and IolT2 are low-affinity glucose permeases with K s values of 2.8 and 1.9 mM, respectively. As glucose uptake and phosphorylation via the PTS differs from glucose uptake via IolT1 or IolT2 and phosphorylation via glucokinase by the requirement for phosphoenolpyruvate, the roles of the two pathways for L-lysine production were tested. The L-lysine yield by C. glutamicum DM1729, a rationally engineered L-lysine-producing strain, was lower than that by its PTS-deficient derivate DM1729⌬hpr, which, however, showed low production rates. The combined overexpression of iolT1 or iolT2 with ppgK, the gene for PolyP/ATP-dependent glucokinase, in DM1729⌬hpr enabled L-lysine production as fast as that by the parent strain DM1729 but with 10 to 20% higher L-lysine yield.The Gram-positive soil bacterium Corynebacterium glutamicum is used industrially for the production of more than 2,160,000 tons of L-glutamate and more than 1,330,000 tons of L-lysine per year (Ajinomoto, Tokyo, Japan). The amino acid production with C. glutamicum is focused on glucose, fructose, and sucrose as carbon sources, with a preference for glucose (30,31). However, C. glutamicum also can use sugars like ribose and maltose, the alcohols inositol and ethanol, and organic acids like acetate, propionate, pyruvate, L-lactate, citrate, and L-glutamate as carbon and energy sources (2, 6).At the phosphoenolpyruvate (PEP)-pyruvate-oxaloacetate node (PPON) (Fig. 1), the distribution of the carbon flux within the metabolism takes place and is especially important for amino acid synthesis (59). To improve L-lysine yields, the optimization of the PPON toward increased oxaloacetate supply is crucial (10, 46). Oxaloacetate utilized for L-lysine synthesis can be regenerated by PEP carboxylase (encoded by ppc) (13) or pyruvate carboxylase (pyc) (48). In terms of gluconeogenesis, oxaloacetate can be converted to either pyruvate by oxaloacetate decarboxylase (odx) (32) or to PEP by PEP carboxykinase (pck) (28). The irreversible conversion of PEP to pyruvate is catalyzed either by pyruvate kinase (pyk) with the formation of ATP (24) or by enzyme I (EI) of the PEP-dependent phosph...

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Metabolic engineering for improving anthranilate synthesis from glucose in Escherichia coli

Cited by 91 publications

References 30 publications

A Novel Muconic Acid Biosynthesis Approach by Shunting Tryptophan Biosynthesis via Anthranilate

A Novel Muconic Acid Biosynthesis Approach by Shunting Tryptophan Biosynthesis via Anthranilate

Metabolic engineering of Escherichia coli for improving l-3,4-dihydroxyphenylalanine (l-DOPA) synthesis from glucose

Phosphotransferase System-Independent Glucose Utilization in Corynebacterium glutamicum by Inositol Permeases and Glucokinases

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