High-yield and plasmid-free biocatalytic production of 5-methylpyrazine-2-carboxylic acid by combinatorial genetic elements engineering and genome engineering of Escherichia coli

Gu, Liuyan; Yuan, Haibo; Li, Guang-Sheng; Cong, Rigang; Li, Jianghua; Du, Guocheng; Liu, Long

doi:10.1016/j.enzmictec.2019.109488

Cited by 18 publications

(6 citation statements)

References 32 publications

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“…As the stress from multiple expression and plasmid duplication occurred, the integration of exogenous genes is considered as a stable and feasible approach for recombinant expression without using antibiotics. , Therefore, RcALAS with or without GroELS was integrated onto the BD chromosome to form Rc7GI and RcI, respectively. As a result, the biomass increased to OD 600 6.8–7.3 in RcI and Rc7GI (Figure A), wherein ALAS was integrated to the chromosome and even increased ALA production (Figure B).…”

Section: Resultsmentioning

confidence: 99%

Low-Carbon-Footprint Production of High-End 5-Aminolevulinic Acid via Integrative Strain Engineering and RuBisCo-Equipped Escherichia coli

Xue

2021

ACS Sustainable Chem. Eng.

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5-Aminolevulinic acid (ALA) is an imperative compound that has been widely applied in the agricultural and medical fields. The ALA synthesis in Escherichia coli via the C4 pathway depends on ALA synthase (ALAS) and releases CO2 in a single-step reaction. However, enhancement of the ALA production by increasing the protein’s quality and quantity with simultaneous CO2 reduction has never been reported. In this study, the expression and activity of ALAS were improved by employing chaperone and rare tRNAs. Moreover, the pdxY gene was introduced for pyridoxal phosphate (PLP) regeneration, a cofactor for ALAS, and significantly increased ALA production to 14.3 g/L through stepwise strain engineering with an optimal feeding strategy. The leading excess CO2 emission was further reused as a second carbon source for cell growth and ALA synthesis by cloning the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCo) and phosphoribulokinase (Prk) in E. coli. As a result, the co-application of RuBisCo, Prk, and ALAS reduced CO2 emission by 53.8% in combination with pdxY and pRARE. Finally, the crude ALA was demonstrated and applied in the antibacterial photodynamic therapy (aPDT) against three different pathogens. The robust strain, RSSA-BY with a lower carbon footprint, achieved the highest cell growth and efficient ALA production for aPDT for the first time, providing a vital and green bioprocess toward an advanced and sustainable future.

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Section: Resultsmentioning

confidence: 99%

Low-Carbon-Footprint Production of High-End 5-Aminolevulinic Acid via Integrative Strain Engineering and RuBisCo-Equipped Escherichia coli

Xue

2021

ACS Sustainable Chem. Eng.

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“…To obtain industrially applied strains, multicopy chromosomal integration of target pathway genes may be a solution. The CRISPR-Cas system allows the knock-in of heterologous genes, integration of large synthetic pathways as well as the combinatorial and multiplex modifications in chromosomes [85][86][87][88][89]. Using CRISPR-associated transposases (MUCICAT) and targeting the crRNA to multicopy loci of the E. coli genome, up to 10 copies of integration were achieved [90].…”

Section: Increasing Production Strain Stabilitymentioning

confidence: 99%

Toward bioproduction of oxo chemicals from C1 feedstocks using isobutyraldehyde as an example

Guo

Sun

Huo

2022

Biotechnol Biofuels

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Oxo chemicals are valuable chemicals for synthesizing a wide array of industrial and consumer products. However, producing of oxo chemicals is predominately through the chemical process called hydroformylation, which requires petroleum-sourced materials and generates abundant greenhouse gas. Current concerns on global climate change have renewed the interest in reducing greenhouse gas emissions and recycling the plentiful greenhouse gas. A carbon–neutral manner in this regard is producing oxo chemicals biotechnologically using greenhouse gas as C1 feedstocks. Exemplifying isobutyraldehyde, this review demonstrates the significance of using greenhouse gas for oxo chemicals production. We highlight the current state and the potential of isobutyraldehyde synthesis with a special focus on the in vivo and in vitro scheme of C1-based biomanufacturing. Specifically, perspectives and scenarios toward carbon– and nitrogen–neutral isobutyraldehyde production are proposed. In addition, key challenges and promising approaches for enhancing isobutyraldehyde bioproduction are thoroughly discussed. This study will serve as a reference case in exploring the biotechnological potential and advancing oxo chemicals production derived from C1 feedstocks.

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“…glmS encoding glucosamine-6-phosphate synthase, glmM encoding phosphoglucosamine mutase, glmU encod-ing N-acetylglucosamine-1-phosphate uridyltransferase/glucosamine-1-phosphate acetyltransferase, pgi encoding glucose-6-phosphate isomerase, pgm encoding phosphoglucomutase, galE encoding UDPglucose 4-epimerase, lacZ encoding β-D-galactosidase, ugd encoding UDP-glucose 6-dehydrogenase, nagB encoding glucosamine-6-phosphate deaminase, manA encoding mannose-6-phosphate isomerase, murA encoding UDP-N-acetylglucosamine 1-carboxyvinyltransferase, wecB encoding UDP-N-acetyl glucosamine-2-epimerase, lgtA β-1,3-N-acetylglucosaminyltransferase, lgtB β-1,4-galactosyltransferase and the primers used were listed in Supplementary Table S3. The 16S ribosomal RNA was chosen as the reference gene for relative quantification [10,34].…”

Section: Real-time Quantitative Pcr (Rt-qpcr)mentioning

confidence: 99%

Metabolic engineering of Escherichia coli for the production of Lacto-N-neotetraose (LNnT)

Zhang

Liu²,

Gong

et al. 2021

Syst Microbiol and Biomanuf

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Lacto-N-neotetraose (LNnT), one of the most important human milk oligosaccharides, can be used as infants' food additives. Nowadays, extraction, chemical and biological synthesis were utilized to obtain LNnT, while these methods still face some problems such as low yield and high cost. The aim of current work is to construct a de novo biosynthesis pathway of LNnT in E. coli K12 MG1655. The lgtA and lgtB were first expressed by a plasmid, resulting in a LNnT titer of 0.04 g/L. To improve the yield of LNnT on substrate lactose, lacZ and lacI were knocked out, and lacY was over-expressed. As a result, the yield of LNnT on lactose increased from 0.01 to 0.09 mol/mol, and the titer of LNnT elevated to 0.41 g/L. In addition, the pathway was regulated using the titer of Lacto-N-triose II (LNTII) as a measure, and obtained a high titer strain of LNnT for 1.04 g/L. Finally, the gene expressions were fine-tuned, the titer of LNnT reached 1.2 g/L, which was 93% higher than the control strain, and the yield on lactose reached 0.28 mol/mol. The engineering strategy of pathway construction and modulation used in this study is applicable to facilitate the microbial production of other metabolites in E. coli.

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High-yield and plasmid-free biocatalytic production of 5-methylpyrazine-2-carboxylic acid by combinatorial genetic elements engineering and genome engineering of Escherichia coli

Cited by 18 publications

References 32 publications

Low-Carbon-Footprint Production of High-End 5-Aminolevulinic Acid via Integrative Strain Engineering and RuBisCo-Equipped Escherichia coli

Low-Carbon-Footprint Production of High-End 5-Aminolevulinic Acid via Integrative Strain Engineering and RuBisCo-Equipped Escherichia coli

Toward bioproduction of oxo chemicals from C1 feedstocks using isobutyraldehyde as an example

Metabolic engineering of Escherichia coli for the production of Lacto-N-neotetraose (LNnT)

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