Ken W. Lu, Chris T. Wang, Hengray Chang, Ryan S. Wang, Claire R. Shen
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引用次数: 4
Abstract
Biosynthesis of itaconic acid occurs through decarboxylation of the TCA cycle intermediate cis-aconitate. Engineering of efficient itaconate producers often requires elimination of the highly active isocitrate dehydrogenase to conserve cis-aconitate, leading to 2-ketoglutarate auxotrophy in the producing strains. Supplementation of glutamate or complex protein hydrolysate then becomes necessary, often in large quantities, to support the high cell density desired during itaconate fermentation and adds to the production cost. Here, we present an alternative approach to overcome the glutamate auxotrophy in itaconate producers by synthetically introducing the Weimberg pathway from Burkholderia xenovorans for 2-ketoglutarate biosynthesis. Because of its independence from natural carbohydrate assimilation pathways in Escherichia coli, the Weimberg pathway is able to provide 2-ketoglutarate using xylose without compromising the carbon flux toward itaconate. With xylose concentration carefully tuned to minimize excess 2-ketoglutarate flux in the stationary phase, the final strain accumulated 20 g/L of itaconate in minimal medium from 18 g/L of xylose and 45 g/L of glycerol. Necessity of the recombinant Weimberg pathway for growth also allowed us to maintain multi-copy plasmids carrying in operon the itaconate-producing genes without addition of antibiotics. Use of the Weimberg pathway for growth restoration is applicable to other production systems with disrupted 2-ketoglutarate synthesis.
期刊介绍:
Metabolic Engineering Communications, a companion title to Metabolic Engineering (MBE), is devoted to publishing original research in the areas of metabolic engineering, synthetic biology, computational biology and systems biology for problems related to metabolism and the engineering of metabolism for the production of fuels, chemicals, and pharmaceuticals. The journal will carry articles on the design, construction, and analysis of biological systems ranging from pathway components to biological complexes and genomes (including genomic, analytical and bioinformatics methods) in suitable host cells to allow them to produce novel compounds of industrial and medical interest. Demonstrations of regulatory designs and synthetic circuits that alter the performance of biochemical pathways and cellular processes will also be presented. Metabolic Engineering Communications complements MBE by publishing articles that are either shorter than those published in the full journal, or which describe key elements of larger metabolic engineering efforts.