Metabolic engineering of Synechocystis sp. PCC 6803 for improved bisabolene production

IF 3.7 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
João S. Rodrigues, Pia Lindberg
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引用次数: 33

Abstract

Terpenoids are a wide class of organic compounds with industrial relevance. The natural ability of cyanobacteria to produce terpenoids via the methylerythritol 4-phosphate (MEP) pathway makes these organisms appealing candidates for the generation of light-driven cell factories for green chemistry. Here we address the improvement of the production of (E)-α-bisabolene, a valuable biofuel feedstock, in Synechocystis sp. PCC 6803 via sequential heterologous expression of bottleneck enzymes of the native pathway. Expression of the bisabolene synthase is sufficient to complete the biosynthetic pathway of bisabolene. Expression of a farnesyl-pyrophosphate synthase from Escherichia coli did not influence production of bisabolene, while enhancement of the MEP pathway via additional overexpression of 1-deoxy-D-xylulose-5-phosphate synthase (DXS) and IPP/DMAPP isomerase (IDI) significantly increased production per cell. However, in the absence of a carbon sink, the overexpression of DXS and IDI leads to significant growth impairment. The final engineered strain reached a volumetric titre of 9 ​mg ​L−1 culture of bisabolene after growing for 12 days. When the cultures were grown in a high cell density (HCD) system, we observed an increase in the volumetric titres by one order of magnitude for all producing-strains. The strain with improved MEP pathway presented an increase twice as much as the remaining engineered strains, yielding more than 180 ​mg ​L−1 culture after 10 days of cultivation. Furthermore, the overexpression of these two MEP enzymes prevented the previously reported decrease in the bisabolene specific titres when grown in HCD conditions, where primary metabolism is usually favoured. We conclude that fine-tuning of the cyanobacterial terpenoid pathway is crucial for the generation of microbial platforms for terpenoid production on industrial-scale.

Abstract Image

聚囊藻(Synechocystis sp. PCC 6803)代谢工程提高双abolene产量
萜类化合物是一类广泛的具有工业意义的有机化合物。蓝藻通过甲基赤藓糖醇4-磷酸(MEP)途径产生萜类物质的天然能力使这些生物成为绿色化学产生光驱动细胞工厂的有吸引力的候选者。本研究通过序列异源表达Synechocystis sp. PCC 6803的瓶颈酶,提高了生物燃料原料(E)-α-双abolene的产量。双abolene合成酶的表达足以完成双abolene的生物合成途径。来自大肠杆菌的法尼基焦磷酸合成酶的表达不影响双abolene的产量,而通过额外过表达1-脱氧-d -木醛糖-5-磷酸合成酶(DXS)和IPP/DMAPP异构酶(IDI)来增强MEP途径,显著增加了每个细胞的产量。然而,在缺乏碳汇的情况下,DXS和IDI的过表达会导致显著的生长损害。最终的工程菌株在生长12天后达到9 mg L−1双abolene培养液的体积滴度。当培养物在高细胞密度(HCD)系统中生长时,我们观察到所有生产菌株的体积滴度增加了一个数量级。改良MEP途径的菌株比其余工程菌株的产量增加了两倍,培养10天后产量超过180 mg L−1。此外,这两种MEP酶的过表达阻止了先前报道的在HCD条件下生长时双abolene特异性滴度的下降,而HCD条件通常有利于初级代谢。我们得出结论,蓝藻萜类途径的微调对于在工业规模上产生萜类生产的微生物平台至关重要。
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来源期刊
Metabolic Engineering Communications
Metabolic Engineering Communications Medicine-Endocrinology, Diabetes and Metabolism
CiteScore
13.30
自引率
1.90%
发文量
22
审稿时长
18 weeks
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.
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