Zymomonas mobilis as an emerging biotechnological chassis for the production of industrially relevant compounds

AbstractZymomonas mobilis is a well-recognized ethanologenic bacterium with outstanding characteristics which make it a promising platform for the biotechnological production of relevant building blocks and fine chemicals compounds. In the last years, research has been focused on the physiological, genetic, and metabolic engineering strategies aiming at expanding Z. mobilis ability to metabolize lignocellulosic substrates toward biofuel production. With the expansion of the Z. mobilis molecular and computational modeling toolbox, the potential of this bacterium as a cell factory has been thoroughly explored. The number of genomic, transcriptomic, proteomic, and fluxomic data that is becoming available for this bacterium has increased. For this reason, in the forthcoming years, systems biology is expected to continue driving the improvement of Z. mobilis for current and emergent biotechnological applications. While the existing molecular toolbox allowed the creation of stable Z. mobilis strains with improved traits for pinpointed biotechnological applications, the development of new and more flexible tools is crucial to boost the engineering capabilities of this bacterium. Novel genetic toolkits based on the CRISPR-Cas9 system and recombineering have been recently used for the metabolic engineering of Z. mobilis. However, they are mostly at the proof-of-concept stage and need to be further improved. Graphical Abstract

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Reassessing Escherichia coli as a cell factory for biofuel production

Current Opinion in Biotechnology ◽

10.1016/j.copbio.2017.02.010 ◽

2017 ◽

Vol 45 ◽

pp. 92-103 ◽

Cited By ~ 28

Author(s):

Chonglong Wang ◽

Brian F Pfleger ◽

Seon-Won Kim

Keyword(s):

Escherichia Coli ◽

Biofuel Production ◽

Cell Factory ◽

A Cell

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Lactic acid bacteria as a cell factory: rerouting of carbon metabolism in Lactococcus lactis by metabolic engineering

Enzyme and Microbial Technology ◽

10.1016/s0141-0229(00)00180-0 ◽

2000 ◽

Vol 26 (9-10) ◽

pp. 840-848 ◽

Cited By ~ 61

Author(s):

Michiel Kleerebezemab ◽

Pascal Hols ◽

Jeroen Hugenholtz

Keyword(s):

Lactic Acid ◽

Metabolic Engineering ◽

Lactic Acid Bacteria ◽

Lactococcus Lactis ◽

Carbon Metabolism ◽

Cell Factory ◽

A Cell

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A metabolic reconstruction of Lactobacillus reuteri JCM 1112 and analysis of its potential as a cell factory

Microbial Cell Factories ◽

10.1186/s12934-019-1229-3 ◽

2019 ◽

Vol 18 (1) ◽

Cited By ~ 2

Author(s):

Thordis Kristjansdottir ◽

Elleke F. Bosma ◽

Filipe Branco dos Santos ◽

Emre Özdemir ◽

Markus J. Herrgård ◽

...

Keyword(s):

Experimental Data ◽

Metabolic Engineering ◽

Growth Rates ◽

Lactobacillus Reuteri ◽

Metabolic Model ◽

Metabolic Reconstruction ◽

Cell Factory ◽

A Genome ◽

A Cell ◽

Genome Scale

Abstract Background Lactobacillus reuteri is a heterofermentative Lactic Acid Bacterium (LAB) that is commonly used for food fermentations and probiotic purposes. Due to its robust properties, it is also increasingly considered for use as a cell factory. It produces several industrially important compounds such as 1,3-propanediol and reuterin natively, but for cell factory purposes, developing improved strategies for engineering and fermentation optimization is crucial. Genome-scale metabolic models can be highly beneficial in guiding rational metabolic engineering. Reconstructing a reliable and a quantitatively accurate metabolic model requires extensive manual curation and incorporation of experimental data. Results A genome-scale metabolic model of L. reuteri JCM 1112T was reconstructed and the resulting model, Lreuteri_530, was validated and tested with experimental data. Several knowledge gaps in the metabolism were identified and resolved during this process, including presence/absence of glycolytic genes. Flux distribution between the two glycolytic pathways, the phosphoketolase and Embden–Meyerhof–Parnas pathways, varies considerably between LAB species and strains. As these pathways result in different energy yields, it is important to include strain-specific utilization of these pathways in the model. We determined experimentally that the Embden–Meyerhof–Parnas pathway carried at most 7% of the total glycolytic flux. Predicted growth rates from Lreuteri_530 were in good agreement with experimentally determined values. To further validate the prediction accuracy of Lreuteri_530, the predicted effects of glycerol addition and adhE gene knock-out, which results in impaired ethanol production, were compared to in vivo data. Examination of both growth rates and uptake- and secretion rates of the main metabolites in central metabolism demonstrated that the model was able to accurately predict the experimentally observed effects. Lastly, the potential of L. reuteri as a cell factory was investigated, resulting in a number of general metabolic engineering strategies. Conclusion We have constructed a manually curated genome-scale metabolic model of L. reuteri JCM 1112T that has been experimentally parameterized and validated and can accurately predict metabolic behavior of this important platform cell factory.

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A metabolic reconstruction of Lactobacillus reuteri JCM 1112 and analysis of its potential as a cell factory

10.1101/708875 ◽

2019 ◽

Author(s):

Thordis Kristjansdottir ◽

Elleke F. Bosma ◽

Filipe Branco dos Santos ◽

Emre Özdemir ◽

Markus J. Herrgård ◽

...

Keyword(s):

Experimental Data ◽

Metabolic Engineering ◽

Growth Rates ◽

Lactobacillus Reuteri ◽

Metabolic Model ◽

Metabolic Reconstruction ◽

Cell Factory ◽

A Genome ◽

A Cell ◽

Genome Scale

AbstractBackgroundLactobacillus reuteri is a heterofermentative Lactic Acid Bacterium (LAB) that is commonly used for food fermentations and probiotic purposes. Due to its robust properties, it is also increasingly considered for use as a cell factory. It produces several industrially important compounds such as 1,3-propanediol and reuterin natively, but for cell factory purposes, developing improved strategies for engineering and fermentation optimization is crucial. Genome-scale metabolic models can be highly beneficial in guiding rational metabolic engineering. Reconstructing a reliable and a quantitatively accurate metabolic model requires extensive manual curation and incorporation of experimental data.ResultsA genome-scale metabolic model of L. reuteri JCM 1112T was reconstructed and the resulting model, Lreuteri_530, was validated and tested with experimental data. Several knowledge gaps in the metabolism were identified and resolved during this process, including presence/absence of glycolytic genes. Flux distribution between the two glycolytic pathways, the phosphoketolase and Embden-Meyerhof-Parnas pathways, varies considerably between LAB species and strains. As these pathways result in different energy yields, it is important to include strain-specific utilization of these pathways in the model. We determined experimentally that the Embden-Meyerhof-Parnas pathway carried at most 7% of the total glycolytic flux. Predicted growth rates from Lreuteri_530 were in good agreement with experimentally determined values. To further validate the prediction accuracy of Lreuteri_530, the predicted effects of glycerol addition and adhE gene knock-out, which results in impaired ethanol production, were compared to in vivo data. Examination of both growth rates and uptake- and secretion rates of the main metabolites in central metabolism demonstrated that the model was able to accurately predict the experimentally observed effects. Lastly, the potential of L. reuteri as a cell factory was investigated, resulting in a number of general metabolic engineering strategies.ConclusionWe have constructed a manually curated genome-scale metabolic model of L. reuteri JCM 1112T that has been experimentally parameterized and validated and can accurately predict metabolic behavior of this important platform cell factory.

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Metabolic engineering of Rhodococcus ruber Chol-4: A cell factory for testosterone production

PLoS ONE ◽

10.1371/journal.pone.0220492 ◽

2019 ◽

Vol 14 (7) ◽

pp. e0220492 ◽

Cited By ~ 1

Author(s):

Govinda Guevara ◽

Yamileth Olortegui Flores ◽

Laura Fernández de las Heras ◽

Julián Perera ◽

Juana María Navarro Llorens

Keyword(s):

Metabolic Engineering ◽

Cell Factory ◽

Rhodococcus Ruber ◽

Testosterone Production ◽

A Cell

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Sporulation in solventogenic and acetogenic clostridia

Applied Microbiology and Biotechnology ◽

10.1007/s00253-021-11289-9 ◽

2021 ◽

Author(s):

Mamou Diallo ◽

Servé W. M. Kengen ◽

Ana M. López-Contreras

Keyword(s):

Current Knowledge ◽

Carbon Sources ◽

Cost Effective ◽

Biotechnological Production ◽

Key Points ◽

Wide Range ◽

Potential Applications ◽

A Cell ◽

Sporulation Initiation ◽

Solventogenic Clostridia

AbstractThe Clostridium genus harbors compelling organisms for biotechnological production processes; while acetogenic clostridia can fix C1-compounds to produce acetate and ethanol, solventogenic clostridia can utilize a wide range of carbon sources to produce commercially valuable carboxylic acids, alcohols, and ketones by fermentation. Despite their potential, the conversion by these bacteria of carbohydrates or C1 compounds to alcohols is not cost-effective enough to result in economically viable processes. Engineering solventogenic clostridia by impairing sporulation is one of the investigated approaches to improve solvent productivity. Sporulation is a cell differentiation process triggered in bacteria in response to exposure to environmental stressors. The generated spores are metabolically inactive but resistant to harsh conditions (UV, chemicals, heat, oxygen). In Firmicutes, sporulation has been mainly studied in bacilli and pathogenic clostridia, and our knowledge of sporulation in solvent-producing or acetogenic clostridia is limited. Still, sporulation is an integral part of the cellular physiology of clostridia; thus, understanding the regulation of sporulation and its connection to solvent production may give clues to improve the performance of solventogenic clostridia. This review aims to provide an overview of the triggers, characteristics, and regulatory mechanism of sporulation in solventogenic clostridia. Those are further compared to the current knowledge on sporulation in the industrially relevant acetogenic clostridia. Finally, the potential applications of spores for process improvement are discussed.Key Points• The regulatory network governing sporulation initiation varies in solventogenic clostridia.• Media composition and cell density are the main triggers of sporulation.• Spores can be used to improve the fermentation process.

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TEMPO-oxidized cellulose poly-ionic drawn fiber, a cell support system proof of concept

Journal of Materials Science ◽

10.1007/s10853-021-06373-4 ◽

2021 ◽

Author(s):

Mariana Alves Rios ◽

Paula Aboud Barbugli ◽

Mônica Rosas Costa Iemma ◽

Rafael Grande ◽

Antônio José Felix Carvalho ◽

...

Keyword(s):

Support System ◽

Oxidized Cellulose ◽

Proof Of Concept ◽

A Cell ◽

Cell Support

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Future trends in diagnosis using laboratory-on-a-chip technologies

Parasitology ◽

10.1017/s0031182099004126 ◽

1999 ◽

Vol 117 (7) ◽

pp. 191-203 ◽

Cited By ~ 16

Author(s):

M. S. TALARY ◽

J. P. H. BURT ◽

R. PETHIG

Keyword(s):

Gene Expression ◽

Lower Cost ◽

Building Blocks ◽

Cancer Diagnostics ◽

Future Trends ◽

Biotechnological Applications ◽

Environmental Testing ◽

Current State ◽

Microelectronic Fabrication ◽

Wide Range

There has been an enormous growth in the development of biotechnological applications, where advances in the techniques of microelectronic fabrication and the technologies of miniaturization and integration in semiconductor industries are being applied to the production of Laboratory-on-a-Chip devices. The aim of this development is to create devices that will perform the same processes that are currently carried out in the laboratory in reduced timescales, at a lower cost, requiring less reagents, and with a greater resolution of detection and specificity. The expectations of this Laboratory-on-a-Chip revolution is that this technology will facilitate rapid advances in gene discovery, genetic mapping and gene expression with broader applications ranging from infectious diseases and cancer diagnostics to food quality and environmental testing. A review of the current state of development in this field reveals the scale of the ongoing revolution and serves to highlight the advances that can be perceived in the development of Laboratory-on-a-Chip technologies. Since miniaturization can be applied to such a wide range of laboratory processes, some of the sub-units that can be used as building blocks in these devices are described, with a brief description of some of the fabrication processes that can be used to create them.

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Metabolic engineering of Escherichia coli for de novo production of 3-phenylpropanol via retrobiosynthesis approach

Microbial Cell Factories ◽

10.1186/s12934-021-01615-1 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Zhenning Liu ◽

Xue Zhang ◽

Dengwei Lei ◽

Bin Qiao ◽

Guang-Rong Zhao

Keyword(s):

Escherichia Coli ◽

Metabolic Engineering ◽

De Novo ◽

Microbial Cell ◽

Cell Factory ◽

Phosphopantetheinyl Transferase ◽

Chromosome Engineering ◽

Microbial Cell Factory ◽

E Coli ◽

Artificial Pathway

Abstract Background 3-Phenylpropanol with a pleasant odor is widely used in foods, beverages and cosmetics as a fragrance ingredient. It also acts as the precursor and reactant in pharmaceutical and chemical industries. Currently, petroleum-based manufacturing processes of 3-phenypropanol is environmentally unfriendly and unsustainable. In this study, we aim to engineer Escherichia coli as microbial cell factory for de novo production of 3-phenypropanol via retrobiosynthesis approach. Results Aided by in silico retrobiosynthesis analysis, we designed a novel 3-phenylpropanol biosynthetic pathway extending from l-phenylalanine and comprising the phenylalanine ammonia lyase (PAL), enoate reductase (ER), aryl carboxylic acid reductase (CAR) and phosphopantetheinyl transferase (PPTase). We screened the enzymes from plants and microorganisms and reconstructed the artificial pathway for conversion of 3-phenylpropanol from l-phenylalanine. Then we conducted chromosome engineering to increase the supply of precursor l-phenylalanine and combined the upstream l-phenylalanine pathway and downstream 3-phenylpropanol pathway. Finally, we regulated the metabolic pathway strength and optimized fermentation conditions. As a consequence, metabolically engineered E. coli strain produced 847.97 mg/L of 3-phenypropanol at 24 h using glucose-glycerol mixture as co-carbon source. Conclusions We successfully developed an artificial 3-phenylpropanol pathway based on retrobiosynthesis approach, and highest titer of 3-phenylpropanol was achieved in E. coli via systems metabolic engineering strategies including enzyme sources variety, chromosome engineering, metabolic strength balancing and fermentation optimization. This work provides an engineered strain with industrial potential for production of 3-phenylpropanol, and the strategies applied here could be practical for bioengineers to design and reconstruct the microbial cell factory for high valuable chemicals.

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