JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2022
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IF 5.895
Ya’nan Shi,Rongshan Li,Jie Zheng,Yubin Xue,Yong Tao,Bo Yu
Bio-based propionate attracts increasing attention owing to its green nature and specific food additive market. To date, the time-consuming and costly fermentation process by strict anaerobes makes propionate production not ideal. In this study, we designed a new route for propionate production, in which 1,2-propanediol was first dehydrated to propionaldehyde and then to propionate by taking advantage of the robust oxidization capacity of the Pseudomonas putida KT2440 strain. The high atom economy (0.97 g/g) in this proposed pathway is more advantageous than the previous l-threonine-derived route (0.62 g/g). The molecular mechanism of the extraordinary oxidation capacity of P. putida KT2440 was first deciphered. The propionate production was realized in P. putida KT2440 by screening suitable glycerol dehydratases and optimizing the expression to eliminate the formation of 1-propanol and the accumulation of the intermediate propionaldehyde. The engineered strain produced propionate with a molar conversion rate of >99% from 1,2-propanediol. A high titer of 46.5 g/L pure propionic acid with a productivity of 1.55 g/L/h and a mass yield of 0.96 g/g was achieved in fed-batch biotransformation. Thus, this study provides another idea for the production of high-purity bio-based propionate from renewable materials with high atom economy.
研究领域:
生物技术
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微生物学
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化学工程
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代谢工程
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DOI
PubMed
高影响因子
Frontiers in Bioengineering and Biotechnology
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2020
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IF 3.644
Sun Jun,Lu Li-Bing,Liang Tian-Xin,Yang Li-Rong,Wu Jian-Ping
Pseudomonas putida (P. putida) KT2440 is a paradigmatic environmental-bacterium that possesses significant potential in synthetic biology, metabolic engineering and biodegradation applications. However, most genome editing methods of P. putida KT2440 depend on heterologous repair proteins and the provision of donor DNA templates, which is laborious and inefficient. In this report, an efficient cytosine base editing system was established by using cytidine deaminase (APOBEC1), enhanced specificity Cas9 nickase (eSpCas9ppD10A) and the uracil DNA glycosylase inhibitor (UGI). This constructed base editor converts C-G into T-A in the absence of DNA strands breaks and donor DNA templates. By introducing a premature stop codon in target spacers, we successfully applied this system for gene inactivation with an efficiency of 25–100% in various Pseudomonas species, including P. putida KT2440, P. aeruginosa PAO1, P. fluorescens Pf-5 and P. entomophila L48. We engineered an eSpCas9ppD10A-NG variant with a NG protospacer adjacent motif to expand base editing candidate sites. By modifying the APOBEC1 domain, we successfully narrowed the editable window to increase gene inactivation efficiency in cytidine-rich spacers. Additionally, multiplex base editing in double and triple loci was achieved with mutation efficiencies of 90–100% and 25–35%, respectively. Taken together, the establishment of a fast, convenient and universal base editing system will accelerate the pace of future research undertaken with P. putida KT2440 and other Pseudomonas species.
研究领域:
合成生物学
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代谢工程
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微生物遗传学
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生物技术
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产品关联
DOI
PubMed
高影响因子
SCIENCE OF THE TOTAL ENVIRONMENT
·
2023
·
IF 9.8
Kaiyue Huo,Siqi Wang,Wanwan Zhao,Hongfu Guo,Weini Xiong,Ruihua Liu,Chao Yang
Currently, 1,2-dichloroethane (DCA) is frequently detected in groundwater and has been listed as a potential human carcinogen by the U.S. EPA. Owing to its toxicity and recalcitrant nature, inefficient DCA mineralization has become a bottleneck of DCA bioremediation. In this study, the first engineered DCA-mineralizing strain KTU-P8DCA was constructed by functional assembly of DCA degradation pathway and enhancing pathway expression with a strong promoter P8 in the biosafety strain Pseudomonas putida KT2440. Strain KTU-P8DCA can metabolize DCA to produce CO 2 and utilize DCA as the sole carbon source for cell growth by quantifying 13 C stable isotope ratios in collected CO 2 and in lyophilized cells. Strain KTU-P8DCA exhibited superior tolerance to high concentrations of DCA. Excellent genetic stability was also observed in continuous passage culture. Therefore, strain KTU-P8DCA has enormous potential for use in bioremediation of sites heavily contaminated with DCA. In the future, our strategy for pathway construction and optimization is expected to be developed as a standard pipeline for creating a wide variety of new contaminants-mineralizing microorganisms. The present study also highlights the power of synthetic biology in creating novel degraders for environmental remediation.
研究领域:
环境科学
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合成生物学
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微生物学
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2024
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IF 5.7
Yihan Wang,Jie Zheng,Yubin Xue,Bo Yu
Dipicolinic acid (DPA), a cyclic diacid, has garnered significant interest due to its potential applications in antimicrobial agents, antioxidants, chelating reagents, and polymer precursors. However, its natural bioproduction is limited since DPA is only accumulated in Bacillus and Clostridium species during sporulation. Thus, heterologous production by engineered strains is of paramount importance for developing a sustainable biological route for DPA production. Pseudomonas putida KT2440 has emerged as a promising host for the production of various chemicals thanks to its robustness, metabolic versatility, and genetic tractability. The dominant Entner–Doudoroff (ED) pathway for glucose metabolism in this strain offers an ideal route for DPA production due to the advantage of NADPH generation and the naturally balanced flux between glyceraldehyde-3-phosphate and pyruvate, which are both precursors for DPA synthesis. In this study, DPA production via the ED pathway was in silico designed in P. putida KT2440. The systematically engineered strain produced dipicolinate with a titer of 11.72 g/L from glucose in a 5 L fermentor. This approach not only provides a sustainable green route for DPA production but also expands our understanding of the metabolic potential of the ED pathway in P. putida KT2440.
研究领域:
微生物学
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生物化学
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代谢工程
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DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2024
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IF 5.7
Huoyong Jiang,Xuexiu Yu,Jingjing Guo,Guangdong Shang,Yijun Dai
The amides 4-trifluoromethylnicotinamide, acrylamide, and benzamide are widely used in agriculture and industry, posing hazards to the environment and animals. Immobilized bacteria are preferred in wastewater treatment, but degradation of these amides by immobilized engineered bacteria has not been explored. Here, engineered Pseudomonas putida KT2440 pLSJ15-amiA was constructed by introducing a new amidase gene expression vector into environmentally safe P. putida KT2440. P. putida KT2440 pLSJ15-amiA had high amidase activity, even at 80 °C. P. putida KT2440 pLSJ15-amiA immobilized with calcium alginate exhibited a greater environmental tolerance than free cells. The amides were rapidly degraded by the immobilized cells, but the activity was inhibited by high concentrations of substrates. The substrate inhibition model revealed that the optimum initial concentrations of 4-trifluoromethylnicotinamide, acrylamide, and benzamide for degradation by immobilized cells were 197.65, 350.76, and 249.40 μmol/L, respectively. This study develops a novel and excellent immobilized biocatalyst for remediation of wastewater containing hazardous amides.
研究领域:
微生物学
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环境科学
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基因工程
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PubMed
高影响因子
BIOPROCESS AND BIOSYSTEMS ENGINEERING
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2019
·Vol. 43
·期 1
·67-73
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IF 2.371
Xu Qianqian,Zheng Zhaojuan,Zou Lihua,Zhang Chen,Yang Fen,Zhou Kaijie,Ouyang Jia
Currently, biotransformation of 5-hydroxymethylfurfural (HMF) into a series of high-value bio-based platform chemicals is massively studied. In this study, selective biooxidation of HMF to 5-hydroxymethyl-2-furancarboxylic acid (HMFCA) by Pseudomonas putida KT2440 with superior titer, yield, and productivity was reported. The biocatalytic performances of P. putida KT2440 were optimized separately. Under optimal conditions, 100% yield of HMFCA was obtained when HMF concentration was less than 150 mM, while the maximum concentration of 155 mM was achieved from 160 mM HMF in 12 h. P. putida KT2440 was highly tolerate to HMF, up to 190 mM. Besides, it was capable of selective oxidation of other furan aldehydes to the corresponding carboxylic acids with good yield of 100%. This study further demonstrates the potential of P. putida KT2440 as a biocatalyst for biomass conversion, as this strain has been proved the capacity to convert and utilize many kinds of biomass-derived sugars and ligin-derived aromatic compounds.
研究领域:
生物技术
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生物化学
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化学工程
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）