Whole-cell biocatalyst for cadaverine production using stable, constitutive and high expression of lysine decarboxylase in recombinant Escherichia coli W3110

Ting, Wan-Wen; Huang, Chia-Hui; Wu, Pong-Yee; Huang, Shih-Fang; Lin, Hung‐Yi; Li, Shengfeng; Chang, Jo‐Shu; Ng, I‐Son

doi:10.1016/j.enzmictec.2021.109811

Cited by 25 publications

(6 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Finally, the cell pellet at an OD 600 value of 4 was resuspended in the reaction buffer of 146 g/L l -lysine monohydrochloride and 0.05 mM pyridoxal phosphate in a 0.9% NaCl solution at 37 °C and 200 rpm for 2 h. Lysine and diaminopentane were quantified by high-performance liquid chromatography (HPLC) analysis using the C18 column (YMC-C18 column), while mobile phase A (100% acetonitrile) and B (25 mM sodium acetate buffer at pH 4.8) at a flow rate of 1.0 mL/min were applied. Detection was carried out at 284 nm …”

Section: Materials and Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Tailoring Genetic Elements of the Plasmid-Driven T7 System for Stable and Robust One-Step Cloning and Protein Expression in Broad Escherichia coli

Tan

Hsiang

2021

ACS Synth. Biol.

Self Cite

View full text Add to dashboard Cite

The plasmid-driven T7 system (PDT7) is a flexible approach to trigger protein overexpression; however, most of the reported PDT7 rely on many auxiliary elements or inducible systems to attenuate the toxicity from the orthogonality of the T7 system, which limits its application as the one-step cloning and protein expression system. In this study, we developed a stable and robust PDT7 via tailoring the genetic elements. By error-prone mutagenesis, a mutated T7RNAP with TTTT insertion conferred a trace but enough amount of T7RNAP for stable and efficient PDT7, denoted as PDT7m. The replication origin was kept at the same level, while the ribosome binding site (RBS) of the T7 promoter was the most contributing factor, thus enhancing the protein expression twofold using PDT7m. For application as a host-independent screening platform, both constitutive and IPTG-inducible PDT7m were constructed. It was found that each strain harnessed different IPTG inducibilities for tailor-made strain selection. Constitutive PDT7m was successfully used to express the homologous protein (i.e., lysine decarboxylase) or heterologous protein (i.e., carbonic anhydrase, CA) as a one-step cloning and protein expression tool to select the best strain for cadaverine (DAP) or CA production, respectively. Additionally, PDT7m is compatible with the pET system for coproduction of DAP and CA simultaneously. Finally, PDT7m was used for in vivo high-end chemical production of aminolevulinic acid (ALA), in which addition of the T7 terminator successfully enhanced 340% ALA titer, thus paving the way to rapidly and effectively screening the superior strain as a cell factory.

show abstract

Section: Materials and Methodsmentioning

confidence: 99%

“…Detection was carried out at 284 nm. 32 Dose−Response Curve and Data Fitting. A 1% (v/v) precultured broth was inoculated in 4 mL of fresh LB medium, and the cells were induced by various concentrations of IPTG ALA Production and Quantification.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

Tailoring Genetic Elements of the Plasmid-Driven T7 System for Stable and Robust One-Step Cloning and Protein Expression in Broad Escherichia coli

Tan

Hsiang

2021

ACS Synth. Biol.

Self Cite

View full text Add to dashboard Cite

show abstract

“…1,5-diaminopentane (cadaverine), an important material monomer, could be applied into synthesis of polyamide (PA), which is a major engineering material with various excellent advantages such as high toughness, strength and resistance to abrasion [ 1 ]. PA 5.X widely applied into various industries, including plastics, fibers, rubbers, foams and other fields, is synthesized by cadaverine and aliphatic diacids, whose market was assessed up to 220 million by 2022 [ 2 ]. However, fossil fuel was the main raw material for cadaverine production in traditional method, which led to plenty of environment problem during the production process.…”

Section: Introductionmentioning

confidence: 99%

A novel co-production of cadaverine and succinic acid based on a thermal switch system in recombinant Escherichia coli

Gao

Wang

et al. 2022

Microb Cell Fact

View full text Add to dashboard Cite

Background Polyamide (nylon) is an important material, which has aroused plenty of attention from all aspects. PA 5.4 is one kind of nylon with excellent property, which consists of cadaverine and succinic acid. Due to the environmental pollution, bio-production of cadaverine and succinic acid has been more attractive due to the less pollution and environmental friendliness. Microbes, like Escherichia coli, has been employed as cell factory to produce cadaverine and succinic acid. However, the accumulation of cadaverine will cause severe damage on cells resulting in inhibition on cell growth and cadaverine production. Herein, a novel two stage co-production of succinic acid and cadaverine was designed based on an efficient thermos-regulated switch to avoid the inhibitory brought by cadaverine. Results The fermentation process was divided into two phase, one for cell growth and lysine production and the other for cadaverine and succinic acid synthesis. The genes of ldhA and ackA were deleted to construct succinic acid pathway in cadaverine producer strain. Then, a thermal switch system based on pR/pL promoter and CI857 was established and optimized. The fermentation conditions were investigated that the optimal temperature for the first stage was determined as 33 ℃ and the optimal temperature for the second stage was 39 ℃. Additionally, the time to shifting temperature was identified as the fermentation anaphase. For further enhance cadaverine and succinic acid production, a scale-up fermentation in 5 L bioreactor was operated. As a result, the titer, yield and productivity of cadaverine was 55.58 g/L, 0.38 g/g glucose and 1.74 g/(L·h), respectively. 28.39 g/L of succinic acid was also obtained with yield of 0.19 g/g glucose. Conclusion The succinic acid metabolic pathway was constructed into cadaverine producer strain to realize the co-production of succinic acid and cadaverine. This study provided a novel craft for industrial co-production of cadaverine and succinic acid.

show abstract

“…Among all E. coli strains, W3110 is the K‐12 derivative that performs as robust and highly resistant to toxic condition; thus it has been frequently exploited for versatile chemicals production including cadaverine (Qian et al, 2011; Ting et al, 2021), l ‐malate (Dong et al, 2017), l ‐Dopa (Das et al, 2018) and other compounds (Koma et al, 2020; Xue et al, 2021b). Therefore, W3110 was chosen as the 5‐ALA producing host with T7RNAP integration at different chromosomal loci (Ting et al, 2022), and investigated the efficiency of T7RNAP for RcALAS expression.…”

Section: Introductionmentioning

confidence: 99%

Effective 5‐aminolevulinic acid production via T7 RNA polymerase and RuBisCO equipped Escherichia coli W3110

Ting

2022

Biotech & Bioengineering

Self Cite

View full text Add to dashboard Cite

Chromosome-based engineering is a superior approach for gene integration generating a stable and robust chassis. Therefore, an effective amplifier, T7 RNA polymerase (T7RNAP) from bacteriophage, has been incorporated into Escherichia coli W3110 by site-specific integration. Herein, we performed the 5-aminolevulinic acid (5-ALA) production in four T7RNAP-equipped W3110 strains using recombinant 5-aminolevulinic synthase and further explored the metabolic difference in best strain. The fastest glucose consumption resulted in the highest biomass and the 5-ALA production reached to 5.5 g/L; thus, the least by-product of acetate was shown in RH strain in which T7RNAP was inserted at HK022 phage attack site.Overexpression of phosphoenolpyruvate (PEP) carboxylase would pull PEP to oxaloacetic acid in tricarboxylic acid cycle, leading to energy conservation and even no acetate production, thus, 6.53 g/L of 5-ALA was achieved. Amino acid utilization in RH deciphered the major metabolic flux in α-ketoglutaric acid dominating 5-ALA production. Finally, the ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and phosphoribulokinase were expressed for carbon dioxide recycling; a robust and efficient chassis toward low-carbon assimilation and high-level of 5-ALA production up to 11.2 g/L in fed-batch fermentation was established.

show abstract

Whole-cell biocatalyst for cadaverine production using stable, constitutive and high expression of lysine decarboxylase in recombinant Escherichia coli W3110

Cited by 25 publications

References 33 publications

Tailoring Genetic Elements of the Plasmid-Driven T7 System for Stable and Robust One-Step Cloning and Protein Expression in Broad Escherichia coli

Tailoring Genetic Elements of the Plasmid-Driven T7 System for Stable and Robust One-Step Cloning and Protein Expression in Broad Escherichia coli

A novel co-production of cadaverine and succinic acid based on a thermal switch system in recombinant Escherichia coli

Effective 5‐aminolevulinic acid production via T7 RNA polymerase and RuBisCO equipped Escherichia coli W3110

Contact Info

Product

Resources

About