Isobutene production in Synechocystis sp. PCC 6803 by introducing α-ketoisocaproate dioxygenase from Rattus norvegicus

IF 3.7 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Metabolic Engineering Communications Pub Date : 2021-06-01 DOI:10.1016/j.mec.2021.e00163
Henna Mustila , Amit Kugler, Karin Stensjö
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引用次数: 11

Abstract

Cyanobacteria can be utilized as a platform for direct phototrophic conversion of CO2 to produce several types of carbon-neutral biofuels. One promising compound to be produced photobiologically in cyanobacteria is isobutene. As a volatile compound, isobutene will quickly escape the cells without building up to toxic levels in growth medium or get caught in the membranes. Unlike liquid biofuels, gaseous isobutene may be collected from the headspace and thus avoid the costly extraction of a chemical from culture medium or from cells. Here we investigate a putative synthetic pathway for isobutene production suitable for a photoautotrophic host. First, we expressed α-ketoisocaproate dioxygenase from Rattus norvegicus (RnKICD) in Escherichia coli. We discovered isobutene formation with the purified RnKICD with the rate of 104.6 ​± ​9 ​ng (mg protein)-1 min-1 using α-ketoisocaproate as a substrate. We further demonstrate isobutene production in the cyanobacterium Synechocystis sp. PCC 6803 by introducing the RnKICD enzyme. Synechocystis strain heterologously expressing the RnKICD produced 91 ​ng ​l−1 OD750−1 ​h−1. Thus, we demonstrate a novel sustainable platform for cyanobacterial production of an important building block chemical, isobutene. These results indicate that RnKICD can be used to further optimize the synthetic isobutene pathway by protein and metabolic engineering efforts.

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引入褐家鼠α-酮异己酸双加氧酶对褐家鼠PCC 6803产异丁烯的影响
蓝藻可以作为直接光养转换二氧化碳的平台,以生产几种碳中性生物燃料。异丁烯是一种在蓝藻中产生光生物学的有前途的化合物。作为一种挥发性化合物,异丁烯会迅速逃离细胞,而不会在生长培养基中形成毒性水平,也不会被细胞膜捕获。与液体生物燃料不同,气态异丁烯可以从顶空收集,从而避免了从培养基或细胞中提取化学物质的昂贵费用。在这里,我们研究了一种适用于光自养寄主的异丁烯合成途径。首先,我们在大肠杆菌中表达褐家鼠α-酮异己酸双加氧酶(RnKICD)。我们发现纯化的RnKICD以α-酮异己酸酯为底物形成异丁烯的速率为104.6±9 ng (mg蛋白)-1 min-1。通过引入RnKICD酶,我们进一步证明了蓝细菌synnechocystis sp. PCC 6803中异丁烯的产生。异源表达RnKICD的胞囊菌产生91 ng l−1 OD750−1 h−1。因此,我们展示了一个新的可持续平台的蓝藻生产的一个重要的构建块化学品,异丁烯。这些结果表明,RnKICD可以通过蛋白质和代谢工程的努力进一步优化合成异丁烯途径。
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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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