Enlist the applications of metabolic engineering.
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Cyanobacteria are promising microorganisms for sustainable biotechnologies, yet unlocking their potential requires radical re-engineering and application of cutting-edge synthetic biology techniques. In recent years, the available devices and strategies for modifying cyanobacteria have been increasing, including advances in the design of genetic promoters, ribosome binding sites, riboswitches, reporter proteins, modular vector systems, and markerless selection systems. Because of these new toolkits, cyanobacteria have been successfully engineered to express heterologous pathways for the production of a wide variety of valuable compounds. Cyanobacterial strains with the potential to be used in real-world applications will require the refinement of genetic circuits used to express the heterologous pathways and development of accurate models that predict how these pathways can be best integrated into the larger cellular metabolic network.
In 1990s, a new technique called metabolic engineering emerged. This technique analyzes the metabolic pathway of a microorganism, and determines the constraints and their effects on the production of desired compounds. It then uses genetic engineering to relieve these constraints. Some examples of successful metabolic engineering are the following: (i) Identification of constraints to lysine production in Corynebacterium glutamicum and insertion of new genes to relieve these constraints to improve production[5] (ii) Engineering of a new fatty acid biosynthesis pathway, called reversed beta oxidation pathway, that is more efficient than the native pathway in producing fatty acids and alcohols which can potentially be catalytically converted to chemicals and fuels[6] (iii) Improved production of DAHP an aromatic metabolite produced by E. coli that is an intermediate in the production of aromatic amino acids.[7] It was determined through metabolic flux analysis that the theoretical maximal yield of DAHP per glucose molecule utilized, was 3/7. This is because some of the carbon from glucose is lost as carbon dioxide, instead of being utilized to produce DAHP. Also, one of the metabolites (PEP, or phosphoenolpyruvate) that are used to produce DAHP, was being converted to pyruvate (PYR) to transport glucose into the cell, and therefore, was no longer available to produce DAHP. In order to relieve the shortage of PEP and increase yield, Patnaik et al. used genetic engineering on E. coli to introduce a reaction that converts PYR back to PEP. Thus, the PEP used to transport glucose into the cell is regenerated, and can be used to make DAHP. This resulted in a new theoretical maximal yield of 6/7 – double that of the native E. coli system.
At the industrial scale, metabolic engineering is becoming more convenient and cost effective. According to the Biotechnology Industry Organization, "more than 50 biorefinery facilities are being built across North America to apply metabolic engineering to produce biofuels and chemicals from renewable biomass which can help reduce greenhouse gas emissions". Potential biofuels include short-chain alcohols and alkanes (to replace gasoline), fatty acid methyl esters and fatty alcohols (to replace diesel), and fatty acid-and isoprenoid-based biofuels (to replace diesel).[8]
Metabolic engineering continues to evolve in efficiency and processes aided by breakthroughs in the field of synthetic biology and progress in understanding metabolite damage and its repair or preemption. Early metabolic engineering experiments showed that accumulation of reactive intermediates can limit flux in engineered pathways and be deleterious to host cells if matching damage control systems are missing or inadequate.[9][10] Researchers in synthetic biology optimize genetic pathways, which in turn influence cellular metabolic outputs. Recent decreases in cost of synthesized DNA and developments in genetic circuits help to influence the ability of metabolic engineering to produce desired outputs.[11]