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通过缺失cyAbrB2转录调节因子,增强了集胞藻PCC 6803中游离脂肪酸的产生。

Free fatty acid production in the cyanobacterium Synechocystis sp. PCC 6803 is enhanced by deletion of the cyAbrB2 transcriptional regulator.

作者信息

Kawahara Akihito, Sato Yusuke, Saito Yujiro, Kaneko Yasuko, Takimura Yasushi, Hagihara Hiroshi, Hihara Yukako

机构信息

Biological Science Laboratories, KAO Corporation, Wakayama 640-8580, Japan.

Graduate School of Science and Engineering, Saitama University, Saitama 338-8570, Japan.

出版信息

J Biotechnol. 2016 Feb 20;220:1-11. doi: 10.1016/j.jbiotec.2015.12.035. Epub 2015 Dec 29.

Abstract

The cyAbrB2 (Sll0822) transcriptional regulator in Synechocystis sp. PCC 6803 is involved in coordination of carbon and nitrogen metabolism and its deletion causes distinct phenotypes such as decreased expression levels of nitrogen-regulated genes and high accumulation of glycogen granules. From the viewpoint of metabolic engineering, the highly accumulated glycogen granules in the ΔcyabrB2 mutant could be a valuable source for the production of biofuels. Here, by disruption of the aas gene (slr1609) encoding acyl-acyl carrier protein synthetase and introduction of a gene encoding thioesterase from Umbellularia californica (UcTE), we conferred the ability of production and secretion of free fatty acids on the ΔcyabrB2 mutant. Notable features of the resulting ΔcyabrB2Δaas::UcTE strain compared with ΔcyabrB2 by RNA-seq analysis were decrease in expression levels of genes related to uptake and subsequent metabolism of nitrogen and carbon and increase in the expression level of sigE encoding a group 2 sigma factor. These changes in gene expression profile were not observed when the same genetic modification was introduced in the wild-type background. The ΔcyabrB2Δaas::UcTE strain showed two-folds higher free fatty acid productivity on a per OD730 basis compared with the Δaas::UcTE strain, without expense of the accumulated glycogen granules. This shows the potential of the ΔcyabrB2 mutant as the platform of biofuel production. The effective utilization of the accumulated glycogen must be the next task to be pursued.

摘要

集胞藻6803中的cyAbrB2(Sll0822)转录调节因子参与碳氮代谢的协调,其缺失会导致明显的表型,如氮调节基因表达水平降低和糖原颗粒高度积累。从代谢工程的角度来看,ΔcyabrB2突变体中高度积累的糖原颗粒可能是生产生物燃料的宝贵来源。在此,通过破坏编码酰基 - 酰基载体蛋白合成酶的aas基因(slr1609)并引入来自加州月桂树(Umbellularia californica)的硫酯酶基因(UcTE),我们赋予了ΔcyabrB2突变体生产和分泌游离脂肪酸的能力。通过RNA测序分析,与ΔcyabrB2相比,所得的ΔcyabrB2Δaas::UcTE菌株的显著特征是与氮和碳的摄取及后续代谢相关的基因表达水平降低,以及编码2类sigma因子的sigE表达水平增加。当在野生型背景中引入相同的基因修饰时,未观察到基因表达谱的这些变化。与Δaas::UcTE菌株相比,ΔcyabrB2Δaas::UcTE菌株在每OD730基础上的游离脂肪酸生产率高出两倍,且不消耗积累的糖原颗粒。这表明ΔcyabrB2突变体作为生物燃料生产平台的潜力。有效利用积累的糖原必定是接下来要探索的任务。

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