Design of Superior Cell Factories for a Sustainable Biorefinery By Synthetic Bioengineering

ABSTRACT In eukaryotic cells, membrane-surrounded organelles are orchestrally organized spatiotemporally under environmental situations. Among such organelles, vesicular transports and membrane contacts occur to communicate each other, so-called membrane traffic. Filamentous fungal cells are highly polarized and thus membrane traffic is developed to have versatile functions. Early endosome (EE) is an endocytic organelle that dynamically exhibits constant long-range motility through the hyphal cell, which is proven to have physiological roles, such as other organelle distribution and signal transduction. Since filamentous fungal cells are also considered as cell factories, to produce valuable proteins extracellularly, molecular mechanisms of secretory pathway including protein glycosylation have been well investigated. In this review, molecular and physiological aspects of membrane traffic especially related to EE dynamics and protein secretion in filamentous fungi are summarized, and perspectives for application are also described.

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Molecular characterization of HEK293 cells as emerging versatile cell factories

Current Opinion in Biotechnology ◽

10.1016/j.copbio.2021.05.001 ◽

2021 ◽

Vol 71 ◽

pp. 18-24

Author(s):

Michela Pulix ◽

Vera Lukashchuk ◽

Daniel C Smith ◽

Alan J Dickson

Keyword(s):

Molecular Characterization ◽

Hek293 Cells ◽

Cell Factories

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

International Journal of Environmental Research and Public Health ◽

10.3390/ijerph18031146 ◽

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.

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

Applied Sciences ◽

10.3390/app11125352 ◽

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.

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

Fuel ◽

10.1016/j.fuel.2021.120858 ◽

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

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Production of small ruminant morbillivirus, rift valley fever virus and lumpy skin disease virus in CelCradle™ -500A bioreactors

BMC Veterinary Research ◽

10.1186/s12917-021-02801-4 ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Halima Rhazi ◽

Najete Safini ◽

Karima Mikou ◽

Meryeme Alhyane ◽

Khalid Omari Tadlaoui ◽

...

Keyword(s):

Disease Virus ◽

Skin Disease ◽

Rift Valley ◽

Rift Valley Fever ◽

Rift Valley Fever Virus ◽

Fever Virus ◽

Valley Fever ◽

Lumpy Skin Disease ◽

Lumpy Skin Disease Virus ◽

Cell Factories

Abstract Background Animal vaccination is an important way to stop the spread of diseases causing immense damage to livestock and economic losses and the potential transmission to humans. Therefore effective method for vaccine production using simple and inexpensive bioprocessing solutions is very essential. Conventional culture systems currently in use, tend to be uneconomic in terms of labor and time involved. Besides, they offer a limited surface area for growth of cells. In this study, the CelCradle™-500A was evaluated as an alternative to replace conventional culture systems in use such as Cell factories for the production of viral vaccines against small ruminant morbillivirus (PPR), rift valley fever virus (RVF) and lumpy skin disease virus (LSD). Results Two types of cells Vero and primary Lamb Testis cells were used to produce these viruses. The study was done in 2 phases as a) optimization of cell growth and b) virus cultivation. Vero cells could be grown to significantly higher cell densities of 3.04 × 109 using the CelCradle™-500A with a shorter doubling time as compared to 9.45 × 108 cells in Cell factories. This represents a 19 fold increase in cell numbers as compared to seeding vs only 3.7 fold in Cell factories. LT cells achieved modestly higher cell densities of 6.7 × 108 as compared to 6.3 × 108 in Cell factories. The fold change in densities for these cells was 3 fold in the CelCradle™-500A vs 2.5 fold in Cell factories. The titers in the conventional system and the bioreactor were not significantly different. However, the Cell-specific virus yield for rift valley fever virus and lumpy skin disease virus are higher (25 virions/cell for rift valley fever virus, and 21.9 virions/cell for lumpy skin disease virus versus 19.9 virions/cell for rift valley fever virus and 10 virions/cell for lumpy skin disease virus). Conclusions This work represents a novel study for primary lamb testis cell culture in CellCradle™-500A bioreactors. In addition, on account of the high cell densities obtained and the linear scalability the titers could be further optimized using other culture process such us perfusion.

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Multi-omics analysis of the bioactive constituents biosynthesis of glandular trichome in Perilla frutescens

BMC Plant Biology ◽

10.1186/s12870-021-03069-4 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Peina Zhou ◽

Mengjiao Yin ◽

Shilin Dai ◽

Ke Bao ◽

Chenglin Song ◽

...

Keyword(s):

Glandular Trichomes ◽

Glandular Trichome ◽

Perilla Frutescens ◽

Morphological Features ◽

Limited Information ◽

Bioactive Constituents ◽

Edible Plant ◽

Cell Factories ◽

Custom Made ◽

Trichome Types

Abstract Background Perilla frutescens (L.) Britt is a medicinal and edible plant widely cultivated in Asia. Terpenoids, flavonoids and phenolic acids are the primary source of medicinal ingredients. Glandular trichomes with multicellular structures are known as biochemical cell factories which synthesized specialized metabolites. However, there is currently limited information regarding the site and mechanism of biosynthesis of these constituents in P. frutescens. Herein, we studied morphological features of glandular trichomes, metabolic profiling and transcriptomes through different tissues. Results Observation of light microscopy and scanning electron microscopy indicated the presence of three distinct glandular trichome types based on their morphological features: peltate, capitate, and digitiform glandular trichomes. The oil of peltate glandular trichomes, collected by custom-made micropipettes and analyzed by LC–MS and GC–MS, contained perillaketone, isoegomaketone, and egomaketone as the major constituents which are consistent with the components of leaves. Metabolomics and transcriptomics were applied to explore the bioactive constituent biosynthesis in the leaves, stem, and root of P. frutescens. Transcriptome sequencing profiles revealed differential regulation of genes related to terpenoids, flavonoids, and phenylpropanoid biosynthesis, respectively with most genes expressed highly in leaves. The genes affecting the development of trichomes were preliminarily predicted and discussed. Conclusions The current study established the morphological and chemical characteristics of glandular trichome types of P. frutescens implying the bioactive constituents were mainly synthesized in peltate glandular trichomes. The genes related to bioactive constituents biosynthesis were explored via transcriptomes, which provided the basis for unraveling the biosynthesis of bioactive constituents in this popular medicinal plant.

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Complete biosynthesis of a sulfated chondroitin in Escherichia coli

Nature Communications ◽

10.1038/s41467-021-21692-5 ◽

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.

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Oleaginous Yeasts as Cell Factories for the Sustainable Production of Microbial Lipids by the Valorization of Agri-Food Wastes

Fermentation ◽

10.3390/fermentation7020050 ◽

2021 ◽

Vol 7 (2) ◽

pp. 50

Author(s):

Antonio Caporusso ◽

Angela Capece ◽

Isabella De De Bari

Keyword(s):

Value Chain ◽

Carbon Sources ◽

Research Progress ◽

Food Wastes ◽

Microbial Lipids ◽

Oleaginous Yeasts ◽

Cell Factories ◽

Side Stream ◽

Versatile Tool ◽

Single Cell Oils

The agri-food industry annually produces huge amounts of crops residues and wastes, the suitable management of these products is important to increase the sustainability of agro-industrial production by optimizing the entire value chain. This is also in line with the driving principles of the circular economy, according to which residues can become feedstocks for novel processes. Oleaginous yeasts represent a versatile tool to produce biobased chemicals and intermediates. They are flexible microbial factories able to grow on different side-stream carbon sources such as those deriving from agri-food wastes, and this characteristic makes them excellent candidates for integrated biorefinery processes through the production of microbial lipids, known as single cell oils (SCOs), for different applications. This review aims to present an extensive overview of research progress on the production and use of oleaginous yeasts and present discussions on the current bottlenecks and perspectives of their exploitation in different sectors, such as foods, biofuels and fine chemicals.

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