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Find video protocols related to scientific articles indexed in Pubmed.
From the first drop to the first truckload: commercialization of microbial processes for renewable chemicals.
Curr. Opin. Biotechnol.
PUBLISHED: 02-27-2013
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Fermentation of carbohydrate substrates by microorganisms represents an attractive route for the manufacture of industrial chemicals from renewable resources. The technology to manipulate metabolism of bacteria and yeast, including the introduction of heterologous chemical pathways, has accelerated research in this field. However, the public literature contains very few examples of strains achieving the production metrics required for commercialization. This article presents the challenges in reaching commercial titer, yield, and productivity targets, along with other necessary strain and process characteristics. It then reviews various methods in systems biology, synthetic biology, enzyme engineering, and fermentation engineering which can be applied to strain improvement, and presents a strategy for using these tools to overcome the major hurdles on the path to commercialization.
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Metabolic engineering of Escherichia coli for direct production of 1,4-butanediol.
Nat. Chem. Biol.
PUBLISHED: 04-01-2011
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1,4-Butanediol (BDO) is an important commodity chemical used to manufacture over 2.5 million tons annually of valuable polymers, and it is currently produced exclusively through feedstocks derived from oil and natural gas. Herein we report what are to our knowledge the first direct biocatalytic routes to BDO from renewable carbohydrate feedstocks, leading to a strain of Escherichia coli capable of producing 18 g l(-1) of this highly reduced, non-natural chemical. A pathway-identification algorithm elucidated multiple pathways for the biosynthesis of BDO from common metabolic intermediates. Guided by a genome-scale metabolic model, we engineered the E. coli host to enhance anaerobic operation of the oxidative tricarboxylic acid cycle, thereby generating reducing power to drive the BDO pathway. The organism produced BDO from glucose, xylose, sucrose and biomass-derived mixed sugar streams. This work demonstrates a systems-based metabolic engineering approach to strain design and development that can enable new bioprocesses for commodity chemicals that are not naturally produced by living cells.
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Dynamic modeling of Escherichia coli metabolic and regulatory systems for amino-acid production.
J. Biotechnol.
PUBLISHED: 01-14-2010
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Our aim is to construct a practical dynamic-simulation system that can model the metabolic and regulatory processes involved in the production of primary metabolites, such as amino acids. We have simulated the production of glutamate by transient batch-cultivation using a model of Escherichia coli central metabolism. Kinetic data were used to produce both the metabolic parts of the model, including the phosphotransferase system, glycolysis, the pentose-phosphate pathway, the tricarboxylic acid cycle, the glyoxylate shunt, and the anaplerotic pathways, and the regulatory parts of the model, including regulation by transcription factors, cyclic AMP receptor protein (CRP), making large colonies protein (Mlc), catabolite repressor/activator (Cra), pyruvate dehydrogenase complex repressor (PdhR), and acetate operon repressor (IclR). RNA polymerase and ribosome concentrations were expressed as a function of the specific growth rate, mu, corresponding to the changes in the growth rate during batch cultivation. Parameter fitting was performed using both extracellular concentration measurements and in vivo enzyme activities determined by (13)C flux analysis. By manual adjustment of the parameters, we simulated the batch fermentation of glucose or fructose by a wild-type strain (MG1655) and a glutamate-producing strain (MG1655 Delta sucA). The differences caused by the carbon source, and by wild-type and glutamate-producing strains, were clearly shown by the simulation. A sensitivity analysis revealed the factors that could be altered to improve the production process. Furthermore, an in silico deletion experiments could suggested the existence of uncharacterized regulation. We concluded that our simulation model could function as a new tool for the rational improvement and design of metabolic and regulatory networks.
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Absolute metabolite concentrations and implied enzyme active site occupancy in Escherichia coli.
Nat. Chem. Biol.
PUBLISHED: 04-08-2009
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Absolute metabolite concentrations are critical to a quantitative understanding of cellular metabolism, as concentrations impact both the free energies and rates of metabolic reactions. Here we use LC-MS/MS to quantify more than 100 metabolite concentrations in aerobic, exponentially growing Escherichia coli with glucose, glycerol or acetate as the carbon source. The total observed intracellular metabolite pool was approximately 300 mM. A small number of metabolites dominate the metabolome on a molar basis, with glutamate being the most abundant. Metabolite concentration exceeds K(m) for most substrate-enzyme pairs. An exception is lower glycolysis, where concentrations of intermediates are near the K(m) of their consuming enzymes and all reactions are near equilibrium. This may facilitate efficient flux reversibility given thermodynamic and osmotic constraints. The data and analyses presented here highlight the ability to identify organizing metabolic principles from systems-level absolute metabolite concentration data.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.