scholarly journals Optimization of Tabersonine Methoxylation to Increase Vindoline Precursor Synthesis in Yeast Cell Factories

Molecules ◽  
2021 ◽  
Vol 26 (12) ◽  
pp. 3596
Author(s):  
Pamela Lemos Cruz ◽  
Natalja Kulagina ◽  
Grégory Guirimand ◽  
Johan-Owen De Craene ◽  
Sébastien Besseau ◽  
...  
Keyword(s):  
Yeast Cell ◽  
Metabolic Flux ◽  
Madagascar Periwinkle ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Gene Copies ◽  
Precursor Synthesis ◽  
Directed Biosynthesis ◽  
High Production ◽  
Chemical Condensation

Plant specialized metabolites are widely used in the pharmaceutical industry, including the monoterpene indole alkaloids (MIAs) vinblastine and vincristine, which both display anticancer activity. Both compounds can be obtained through the chemical condensation of their precursors vindoline and catharanthine extracted from leaves of the Madagascar periwinkle. However, the extensive use of these molecules in chemotherapy increases precursor demand and results in recurrent shortages, explaining why the development of alternative production approaches, such microbial cell factories, is mandatory. In this context, the precursor-directed biosynthesis of vindoline from tabersonine in yeast-expressing heterologous biosynthetic genes is of particular interest but has not reached high production scales to date. To circumvent production bottlenecks, the metabolic flux was channeled towards the MIA of interest by modulating the copy number of the first two genes of the vindoline biosynthetic pathway, namely tabersonine 16-hydroxylase and tabersonine-16-O-methyltransferase. Increasing gene copies resulted in an optimized methoxylation of tabersonine and overcame the competition for tabersonine access with the third enzyme of the pathway, tabersonine 3-oxygenase, which exhibits a high substrate promiscuity. Through this approach, we successfully created a yeast strain that produces the fourth biosynthetic intermediate of vindoline without accumulation of other intermediates or undesired side-products. This optimization will probably pave the way towards the future development of yeast cell factories to produce vindoline at an industrial scale.

scholarly journals Synthesizing ginsenoside Rh2 in Saccharomyces cerevisiae cell factory at high-efficiency

2019 ◽  
Vol 5 (1) ◽  
Author(s):  
Pingping Wang ◽  
Wei Wei ◽  
Wei Ye ◽  
Xiaodong Li ◽  
Wenfang Zhao ◽  
...  
Keyword(s):  
Yeast Cell ◽  
Batch Fermentation ◽  
Cell Factory ◽  
Ginsenoside Rh2 ◽  
Fed Batch ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Fed Batch Fermentation ◽  
Shake Flasks ◽  
Produce Plant

AbstractSynthetic biology approach has been frequently applied to produce plant rare bioactive compounds in microbial cell factories by fermentation. However, to reach an ideal manufactural efficiency, it is necessary to optimize the microbial cell factories systemically by boosting sufficient carbon flux to the precursor synthesis and tuning the expression level and efficiency of key bioparts related to the synthetic pathway. We previously developed a yeast cell factory to produce ginsenoside Rh2 from glucose. However, the ginsenoside Rh2 yield was too low for commercialization due to the low supply of the ginsenoside aglycone protopanaxadiol (PPD) and poor performance of the key UDP-glycosyltransferase (UGT) (biopart UGTPg45) in the final step of the biosynthetic pathway. In the present study, we constructed a PPD-producing chassis via modular engineering of the mevalonic acid pathway and optimization of P450 expression levels. The new yeast chassis could produce 529.0 mg/L of PPD in shake flasks and 11.02 g/L in 10 L fed-batch fermentation. Based on this high PPD-producing chassis, we established a series of cell factories to produce ginsenoside Rh2, which we optimized by improving the C3–OH glycosylation efficiency. We increased the copy number of UGTPg45, and engineered its promoter to increase expression levels. In addition, we screened for more efficient and compatible UGT bioparts from other plant species and mutants originating from the direct evolution of UGTPg45. Combining all engineered strategies, we built a yeast cell factory with the greatest ginsenoside Rh2 production reported to date, 179.3 mg/L in shake flasks and 2.25 g/L in 10 L fed-batch fermentation. The results set up a successful example for improving yeast cell factories to produce plant rare natural products, especially the glycosylated ones.

scholarly journals Enhancement of β-Alanine Biosynthesis in Escherichia coli Based on Multivariate Modular Metabolic Engineering

Biology ◽  
2021 ◽  
Vol 10 (10) ◽  
pp. 1017
Author(s):  
Jian Xu ◽  
Li Zhou ◽  
Zhemin Zhou
Keyword(s):  
Escherichia Coli ◽  
Metabolic Engineering ◽  
Industrial Production ◽  
Metabolic Flux ◽  
Coli Strain ◽  
Biosynthesis Pathway ◽  
Fed Batch ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Fed Batch Fermentation

β-alanine is widely used as an intermediate in industrial production. However, the low production of microbial cell factories limits its further application. Here, to improve the biosynthesis production of β-alanine in Escherichia coli, multivariate modular metabolic engineering was recruited to manipulate the β-alanine biosynthesis pathway through keeping the balance of metabolic flux among the whole metabolic network. The β-alanine biosynthesis pathway was separated into three modules: the β-alanine biosynthesis module, TCA module, and glycolysis module. Global regulation was performed throughout the entire β-alanine biosynthesis pathway rationally and systematically by optimizing metabolic flux, overcoming metabolic bottlenecks and weakening branch pathways. As a result, metabolic flux was channeled in the direction of β-alanine biosynthesis without huge metabolic burden, and 37.9 g/L β-alanine was generated by engineered Escherichia coli strain B0016-07 in fed-batch fermentation. This study was meaningful to the synthetic biology of β-alanine industrial production.

scholarly journals Enzymatic Hydrolysate of Cinnamon Waste Material as Feedstock for the Microbial Production of Carotenoids

2021 ◽  
Vol 18 (3) ◽  
pp. 1146
Author(s):  
Stefano Bertacchi ◽  
Stefania Pagliari ◽  
Chiara Cantù ◽  
Ilaria Bruni ◽  
Massimo Labra ◽  
...  
Keyword(s):  
Industrial Sector ◽  
Waste Material ◽  
Fungal Cell ◽  
Enzymatic Hydrolysate ◽  
Carbon And Nitrogen ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Separate Hydrolysis And Fermentation ◽  
Additional Processing ◽  
Microbial Cultivation

In the context of the global need to move towards circular economies, microbial cell factories can be employed thanks to their ability to use side-stream biomasses from the agro-industrial sector to obtain additional products. The valorization of residues allows for better and complete use of natural resources and, at the same time, for the avoidance of waste management to address our needs. In this work, we focused our attention on the microbial valorization of cinnamon waste material after polyphenol extraction (C-PEW) (Cinnamomum verum J.Presl), generally discarded without any additional processing. The sugars embedded in C-PEW were released by enzymatic hydrolysis, more compatible than acid hydrolysis with the subsequent microbial cultivation. We demonstrated that the yeast Rhodosporidium toruloides was able to grow and produce up to 2.00 (±0.23) mg/L of carotenoids in the resulting hydrolysate as a sole carbon and nitrogen source despite the presence of antimicrobial compounds typical of cinnamon. To further extend the potential of our finding, we tested other fungal cell factories for growth on the same media. Overall, these results are opening the possibility to develop separate hydrolysis and fermentation (SHF) bioprocesses based on C-PEW and microbial biotransformation to obtain high-value molecules.

scholarly journals Protein-Engineered Polymers Functionalized with Antimicrobial Peptides for the Development of Active Surfaces

2021 ◽  
Vol 11 (12) ◽  
pp. 5352
Author(s):  
Ana Margarida Pereira ◽  
Diana Gomes ◽  
André da Costa ◽  
Simoni Campos Dias ◽  
Margarida Casal ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Antimicrobial Peptides ◽  
Recombinant Dna ◽  
Biological Properties ◽  
Resistant Bacteria ◽  
Recombinant Dna Technology ◽  
Free Standing ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Antimicrobial Materials

Antibacterial resistance is a major worldwide threat due to the increasing number of infections caused by antibiotic-resistant bacteria with medical devices being a major source of these infections. This suggests the need for new antimicrobial biomaterial designs able to withstand the increasing pressure of antimicrobial resistance. Recombinant protein polymers (rPPs) are an emerging class of nature-inspired biopolymers with unique chemical, physical and biological properties. These polymers can be functionalized with antimicrobial molecules utilizing recombinant DNA technology and then produced in microbial cell factories. In this work, we report the functionalization of rPBPs based on elastin and silk-elastin with different antimicrobial peptides (AMPs). These polymers were produced in Escherichia coli, successfully purified by employing non-chromatographic processes, and used for the production of free-standing films. The antimicrobial activity of the materials was evaluated against Gram-positive and Gram-negative bacteria, and results showed that the polymers demonstrated antimicrobial activity, pointing out the potential of these biopolymers for the development of new advanced antimicrobial materials.

Developing fourth-generation biofuels secreting microbial cell factories for enhanced productivity and efficient product recovery; a review

Fuel ◽  
2021 ◽  
Vol 298 ◽  
pp. 120858
Author(s):  
Sana Malik ◽  
Ayesha Shahid ◽  
Chen-Guang Liu ◽  
Aqib Zafar Khan ◽  
Muhammad Zohaib Nawaz ◽  
...  
Keyword(s):  
Product Recovery ◽  
Microbial Cell ◽  
Fourth Generation ◽  
Microbial Cell Factories ◽  
Cell Factories

scholarly journals Complete biosynthesis of a sulfated chondroitin in Escherichia coli

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Abinaya Badri ◽  
Asher Williams ◽  
Adeola Awofiranye ◽  
Payel Datta ◽  
Ke Xia ◽  
...  
Keyword(s):  
Chondroitin Sulfate ◽  
Scale Up ◽  
Production Methods ◽  
E Coli ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Microbial Product ◽  
Dry Cell Weight ◽  
One Step ◽  
Dry Cell

AbstractSulfated glycosaminoglycans (GAGs) are a class of important biologics that are currently manufactured by extraction from animal tissues. Although such methods are unsustainable and prone to contamination, animal-free production methods have not emerged as competitive alternatives due to complexities in scale-up, requirement for multiple stages and cost of co-factors and purification. Here, we demonstrate the development of single microbial cell factories capable of complete, one-step biosynthesis of chondroitin sulfate (CS), a type of GAG. We engineer E. coli to produce all three required components for CS production–chondroitin, sulfate donor and sulfotransferase. In this way, we achieve intracellular CS production of ~27 μg/g dry-cell-weight with about 96% of the disaccharides sulfated. We further explore four different factors that can affect the sulfation levels of this microbial product. Overall, this is a demonstration of simple, one-step microbial production of a sulfated GAG and marks an important step in the animal-free production of these molecules.

scholarly journals Metabolic engineering strategy for synthetizing trans-4-hydroxy-l-proline in microorganisms

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Zhenyu Zhang ◽  
Pengfu Liu ◽  
Weike Su ◽  
Huawei Zhang ◽  
Wenqian Xu ◽  
...  
Keyword(s):  
Metabolic Engineering ◽  
Biosynthetic Pathway ◽  
Industrial Applications ◽  
Microbial Cell Factories ◽  
Important Amino Acid ◽  
Cell Factories ◽  
Complex Process ◽  
Engineering Strategy ◽  
Hydrolysis Of ◽  
New Strategies

AbstractTrans-4-hydroxy-l-proline is an important amino acid that is widely used in medicinal and industrial applications, particularly as a valuable chiral building block for the organic synthesis of pharmaceuticals. Traditionally, trans-4-hydroxy-l-proline is produced by the acidic hydrolysis of collagen, but this process has serious drawbacks, such as low productivity, a complex process and heavy environmental pollution. Presently, trans-4-hydroxy-l-proline is mainly produced via fermentative production by microorganisms. Some recently published advances in metabolic engineering have been used to effectively construct microbial cell factories that have improved the trans-4-hydroxy-l-proline biosynthetic pathway. To probe the potential of microorganisms for trans-4-hydroxy-l-proline production, new strategies and tools must be proposed. In this review, we provide a comprehensive understanding of trans-4-hydroxy-l-proline, including its biosynthetic pathway, proline hydroxylases and production by metabolic engineering, with a focus on improving its production.

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