scholarly journals High-level de novo biosynthesis of glycosylated zeaxanthin and astaxanthin in Escherichia coli

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Xixian Chen ◽  
Xiaohui Lim ◽  
Aurélie Bouin ◽  
Thomas Lautier ◽  
Congqiang Zhang
Keyword(s):  
Escherichia Coli ◽  
Water Solubility ◽  
De Novo ◽  
Biological Activities ◽  
Total Yield ◽  
Carbon Sources ◽  
Fine Tuning ◽  
Total Carotenoids ◽  
High Level ◽  
Microbial Biosynthesis

AbstractBecause of wide applications in food, feed, pharmaceutical and cosmetic industries, the carotenoid market is growing rapidly. Most carotenoids are hydrophobic, which limits their bioavailability. Glycosylation is a natural route that substantially increases the water solubility, as well as the bioavailability, photostability and biological activities of carotenoids. Here, we report metabolic engineering efforts (e.g., promoter and RBS engineering, optimization of carbon sources and supplementation of bottleneck genes) to produce glycosylated carotenoids in Escherichia coli. By fine-tuning the carotenoid-biosynthetic genes (crtX, crtZ and crtY), our strain produced up to 47.2 mg/L (~ 11,670 ppm) of zeaxanthin glucosides, ~ 78% of the total carotenoids produced. In another construct with mevalonate, astaxanthin pathway and crtX genes, the strain produced a mixture of carotenoid glucosides including astaxanthin and adonixanthin glucosides with a total yield of 8.1 mg/L (1774 ppm). Our work demonstrated a proof-of-concept study for the microbial biosynthesis of glycosylated carotenoids.

scholarly journals Metabolic engineering Escherichia coli for efficient production of icariside D2

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Xue Liu ◽  
Lingling Li ◽  
Jincong Liu ◽  
Jianjun Qiao ◽  
Guang-Rong Zhao
Keyword(s):  
Escherichia Coli ◽  
De Novo ◽  
Carbon Sources ◽  
Gene Expression Pattern ◽  
Fine Tuning ◽  
Cell Factory ◽  
Efficient Production ◽  
Uridine Diphosphate ◽  
Microbial Cell Factory ◽  
E Coli

Abstract Background Icariside D2 is a plant-derived natural glycoside with pharmacological activities of inhibiting angiotensin-converting enzyme and killing leukemia cancer cells. Production of icariside D2 by plant extraction and chemical synthesis is inefficient and environmentally unfriendly. Microbial cell factory offers an attractive route for economical production of icariside D2 from renewable and sustainable bioresources. Results We metabolically constructed the biosynthetic pathway of icariside D2 in engineered Escherichia coli. We screened the uridine diphosphate glycosyltransferases (UGTs) and obtained an active RrUGT3 that regio-specifically glycosylated tyrosol at phenolic position to exclusively synthesize icariside D2. We put heterologous genes in E. coli cell for the de novo biosynthesis of icariside D2. By fine-tuning promoter and copy number as well as balancing gene expression pattern to decrease metabolic burden, the BMD10 monoculture was constructed. Parallelly, for balancing pathway strength, we established the BMT23–BMD12 coculture by distributing the icariside D2 biosynthetic genes to two E. coli strains BMT23 and BMD12, responsible for biosynthesis of tyrosol from preferential xylose and icariside D2 from glucose, respectively. Under the optimal conditions in fed-batch shake-flask fermentation, the BMD10 monoculture produced 3.80 g/L of icariside D2 using glucose as sole carbon source, and the BMT23–BMD12 coculture produced 2.92 g/L of icariside D2 using glucose–xylose mixture. Conclusions We for the first time reported the engineered E. coli for the de novo efficient production of icariside D2 with gram titer. It would be potent and sustainable approach for microbial production of icariside D2 from renewable carbon sources. E. coli–E. coli coculture approach is not limited to glycoside production, but could also be applied to other bioproducts.

scholarly journals Genome-Wide Metabolic Reconstruction of the Synthesis of Polyhydroxyalkanoates from Sugars and Fatty Acids by Burkholderia Sensu Lato Species

2021 ◽  
Vol 9 (6) ◽  
pp. 1290
Author(s):  
Natalia Alvarez-Santullano ◽  
Pamela Villegas ◽  
Mario Sepúlveda Mardones ◽  
Roberto E. Durán ◽  
Raúl Donoso ◽  
...  
Keyword(s):  
Fatty Acids ◽  
De Novo ◽  
Reducing Power ◽  
Pentose Phosphate ◽  
Fine Tuning ◽  
Metabolic Reconstruction ◽  
Phylogenetic Groups ◽  
Outlier Group ◽  
Pha Genes ◽  
High Level

Burkholderia sensu lato (s.l.) species have a versatile metabolism. The aims of this review are the genomic reconstruction of the metabolic pathways involved in the synthesis of polyhydroxyalkanoates (PHAs) by Burkholderia s.l. genera, and the characterization of the PHA synthases and the pha genes organization. The reports of the PHA synthesis from different substrates by Burkholderia s.l. strains were reviewed. Genome-guided metabolic reconstruction involving the conversion of sugars and fatty acids into PHAs by 37 Burkholderia s.l. species was performed. Sugars are metabolized via the Entner–Doudoroff (ED), pentose-phosphate (PP), and lower Embden–Meyerhoff–Parnas (EMP) pathways, which produce reducing power through NAD(P)H synthesis and PHA precursors. Fatty acid substrates are metabolized via β-oxidation and de novo synthesis of fatty acids into PHAs. The analysis of 194 Burkholderia s.l. genomes revealed that all strains have the phaC, phaA, and phaB genes for PHA synthesis, wherein the phaC gene is generally present in ≥2 copies. PHA synthases were classified into four phylogenetic groups belonging to class I II and III PHA synthases and one outlier group. The reconstruction of PHAs synthesis revealed a high level of gene redundancy probably reflecting complex regulatory layers that provide fine tuning according to diverse substrates and physiological conditions.

High-level De novo biosynthesis of arbutin in engineered Escherichia coli

2017 ◽  
Vol 42 ◽  
pp. 52-58 ◽  
Author(s):  
Xiaolin Shen ◽  
Jia Wang ◽  
Jian Wang ◽  
Zhenya Chen ◽  
Qipeng Yuan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
De Novo ◽  
De Novo Biosynthesis ◽  
Engineered Escherichia Coli ◽  
High Level

scholarly journals De novo Biosynthesis of Tyrosol Acetate and Hydroxytyrosol Acetate from Glucose in Engineered Escherichia Coli

2021 ◽  
Author(s):  
Daoyi Guo ◽  
Xiao Fu ◽  
Yue Sun ◽  
Xun Li ◽  
Hong Pan
Keyword(s):  
Escherichia Coli ◽  
De Novo ◽  
Virgin Olive Oil ◽  
Carbon Sources ◽  
Acetate Production ◽  
E Coli ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Alcohol Acetyltransferase ◽  
First Time

Abstract Background: Tyrosol and hydroxytyrosol derived from virgin olive oil and olives extract, have wide applications both as functional food components and as nutraceuticals. However, they have low bioavailability due to their low absorption and high metabolism in human liver and small intestine. Acetylation of tyrosol and hydroxytyrosol can effectively improve their bioavailability and thus increase their potential use in the food and cosmeceutical industries. There is no report on the bioproductin of tyrosol acetate and hydroxytyrosol acetate so far. Thus, it is of great significance to develop microbial cell factories for achieving tyrosol acetate or hydroxytyrosol acetate biosynthesis.Results: In this study, two de novo biosynthetic pathways for the production of tyrosol acetate and hydroxytyrosol acetate were constructed in Escherichia coli. First, an engineered E. coli that allows production of tyrosol from simple carbon sources was established. Four aldehyde reductases were compared, and it was found that yeaE is the best aldehyde reductase for tyrosol accumulation. Subsequently, the pathway was extended for tyrosol acetate production by further overexpression of alcohol acetyltransferase ATF1 for the conversion of tyrosol to tyrosol acetate. Finally, the pathway was further extended for hydroxytyrosol acetate production by overexpression of 4-hydroxyphenylacetate 3-hydroxylase HpaBC.Conclusion: We have successfully established the artificial biosynthetic pathway of tyrosol acetate and hydroxytyrosol acetate from fermentable sugars and demonstrated for the first time the direct fermentative production of tyrosol acetate and hydroxytyrosol acetate from glucose in engineered E. coli

IncI1 ST3 and IncI1 ST7 plasmids from CTX-M-1-producing Escherichia coli obtained from patients with bloodstream infections are closely related to plasmids from E. coli of animal origin

2019 ◽  
Vol 74 (8) ◽  
pp. 2171-2175 ◽  
Author(s):  
Adam Valcek ◽  
Louise Roer ◽  
Søren Overballe-Petersen ◽  
Frank Hansen ◽  
Valeria Bortolaia ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
De Novo ◽  
Variable Region ◽  
Bloodstream Infections ◽  
Animal Origin ◽  
Respective Group ◽  
Surveillance Programme ◽  
E Coli ◽  
High Level ◽  
Plasmid Rearrangements

Abstract Objectives Fully sequenced IncI1 plasmids obtained from CTX-M-1-producing Escherichia coli of human and animal origin were compared. Methods Twelve E. coli isolates sharing identical ESBL genes and plasmid multilocus STs sequenced on Illumina and MinION platforms were obtained from the Danish antimicrobial resistance surveillance programme, DANMAP. After de novo assembly, the sequences of plasmids harbouring blaCTX-M-1 were manually curated and ORFs annotated. Within-group comparisons were performed separately for the IncI1 ST3 plasmid type and the IncI1 ST7 plasmid type. The IncI1 ST3 plasmid group was obtained from 10 E. coli isolates (2 from patients with bloodstream infections, 6 from food and 2 from animals). The IncI1 ST7 plasmids originated from E. coli isolates obtained from a patient with bloodstream infection and from a pig. Sequences of IncI1 ST3 and IncI1 ST7 plasmids harbouring blaCTX-M-1 with determined origin were retrieved from GenBank and used for comparison within the respective group. Results The 10 IncI1 ST3 blaCTX-M-1 plasmids were highly similar in structure and organization with only minor plasmid rearrangements and differences in the variable region. The IncI1 ST7 blaCTX-M-1 plasmids also showed high similarity in structure and organization. The high level of similarity was also observed when including plasmids from E. coli of animal origin from Australia, Switzerland, the Netherlands and France. Conclusions This study shows broad spread of a very successful CTX-M-1-producing IncI1 type plasmid among E. coli of both human and animal origin.

scholarly journals Metabolic engineering of Saccharomyces cerevisiae for enhanced production of caffeic acid

2020 ◽  
Author(s):  
Pingping Zhou ◽  
Chunlei Yue ◽  
Bin Shen ◽  
Yi Du ◽  
Nannan Xu ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Metabolic Engineering ◽  
Caffeic Acid ◽  
Acid Production ◽  
De Novo ◽  
Feedback Inhibition ◽  
Biological Activities ◽  
Acid Product ◽  
Enhanced Production ◽  
Microbial Biosynthesis

Abstract Background As a natural phenolic acid product of plant source, caffeic acid displays diverse biological activities and acts as an important precursor for the synthesis of other valuable compounds. Limitations in chemical synthesis or plant extraction of caffeic acid trigger interest in its microbial biosynthesis. Recently, Saccharomyces cerevisiae has been reported sporadically for biosynthesis of caffeic acid via free plasmid‑mediated pathway assembly. However, the production was far from satisfactory and even relied on the addition of precursor. Results In this study, we first established a controllable caffeic acid pathway by employing a modified GAL regulatory system in S. cerevisiae and realized de novo biosynthesis of 313.8 mg/L caffeic acid from glucose. Combinatorial engineering strategies including eliminating the tyrosine-induced feedback inhibition, deleting genes involved in competing pathways and overexpressing rate-limiting enzymes led to about 2.5-fold improvement in the caffeic acid production, reaching up to 769.3 mg/L in shake-flask cultures. To our knowledge, this is the highest ever reported titer of caffeic acid de novo synthesized by engineered yeast. Conclusions Caffeic acid production in S. cerevisiae strain was successfully improved by adopting a glucose-regulated GAL system and comprehensive metabolic engineering strategies. This work showed the prospect for microbial biosynthesis of caffeic acid and laid the foundation for constructing biosynthetic pathways of its derived metabolites.

scholarly journals Cyt1Ca from Bacillus thuringiensis subsp. israelensis: production in Escherichia coli and comparison of its biological activities with those of other Cyt-like proteins

Microbiology ◽  
2006 ◽  
Vol 152 (9) ◽  
pp. 2651-2659 ◽  
Author(s):  
Robert Manasherob ◽  
Mark Itsko ◽  
Nadine Sela-Baranes ◽  
Eitan Ben-Dov ◽  
Colin Berry ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Bacillus Thuringiensis ◽  
Biological Activities ◽  
Nitrilotriacetic Acid ◽  
Binding Motif ◽  
Carbohydrate Binding ◽  
High Level Expression ◽  
E Coli ◽  
C Terminus ◽  
High Level

The larvicidal activity of Bacillus thuringiensis subsp. israelensis against dipteran larvae is determined by four major polypeptides of the parasporal crystalline body produced during sporulation. Cyt1Aa shows the lowest toxicity when used alone but is the most synergistic with any of the other proteins. The sequence of the plasmid pBtoxis, which contains all the toxin genes in this subspecies, revealed a new cyt-like coding sequence named cyt1Ca. In addition to the Cyt-like region, the predicted Cyt1Ca contained an extra domain at the C terminus, which appeared to be a β-trefoil carbohydrate-binding motif, as found in several ricin-like toxins. The gene was PCR-amplified from pBtoxis and cloned in several vectors, allowing high-level expression in Escherichia coli. Cyt1Ca was purified by nickel-nitrilotriacetic acid affinity chromatography, characterized, and its biological activity was determined. Toxicity against larvae of Aedes aegypti of Cyt1Ca in recombinant E. coli cells was compared with that of Cyt1Aa and Cyt2Ba, and the ability of these proteins to enhance the activity of Cry4Aa was assessed. Although Cyt2Ba appeared able to interact with Cry4Aa, no activity for Cyt1Ca was observed, even when produced in truncated form. Furthermore, in contrast to Cyt1Aa, Cyt1Ca did not lyse sheep erythrocytes, and it was not bactericidal to the host cell.

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