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通过共表达phaCAB 和 vgb 基因,并利用阿拉伯糖 P 启动子控制,在大肠杆菌中增强聚羟基丁酸酯(PHB)的生产。

Enhanced polyhydroxybutyrate (PHB) production via the coexpressed phaCAB and vgb genes controlled by arabinose P promoter in Escherichia coli.

机构信息

Department of Laboratory Medicine and Biotechnology, College of Medicine, Tzu Chi University, Hualien, Taiwan, R.O.C.

出版信息

Lett Appl Microbiol. 2010 Feb;50(2):158-67. doi: 10.1111/j.1472-765X.2009.02772.x. Epub 2009 Nov 12.

Abstract

AIM

To develop an approach to enhance polyhydroxybutyrate (PHB) production via the coexpressed phaCAB and vgb genes controlled by arabinose P(BAD) promoter in Escherichia coli.

METHOD AND RESULTS

The polyhydroxyalkanoates (PHAs) synthesis operon, (phaCAB), from Ralstonia eutropha was overexpressed under the regulation of the arabinose P(BAD) promoter in Escherichia coli, and the vgb gene encoding bacterial haemoglobin from Vitreoscilla stercoraria (VHb) was further cloned at downstream of phaCAB to form an artificial operon. The cell dry weight (CDW), PHB content and PHB concentration were enhanced around 1.23-, 1.57-, and 1.93-fold in the engineered cell harbouring phaCAB-vgb (SY-2) upon 1% arabinose induction compared with noninduction (0% arabinose). Furthermore, by using a recombinant strain harbouring P(BAD) promoter-vgb along with native promoter-phaCAB construction, the effect of vgb expression level on PHB biosynthesis was positive correlation.

CONCLUSIONS

The results exploit the possibility to improve the PHB production by fusing the genes phaCAB-vgb from different species under the arabinose regulation system in E. coli. It also demonstrates that increase in VHb level enhances the PHB production.

SIGNIFICANCE AND IMPACT OF THE STUDY

We were successful in providing a new coexpressed system for PHB synthesis in E. coli. This coexpressed system could be regulated by arabinose inducer, and is more stable and cheaper than other induced systems (e.g. IPTG). Furthermore, it could be applied in many biotechnology or fermentation processes.

摘要

目的

通过共表达phaCAB 和 vgb 基因,构建受阿拉伯糖 P(BAD)启动子调控的工程菌,提高聚羟基丁酸(PHB)产量。

方法和结果

在大肠杆菌中,受阿拉伯糖 P(BAD)启动子调控,过量表达来自恶臭假单胞菌的多聚羟基烷酸(PHA)合成操纵子(phaCAB),并进一步在其下游克隆编码细菌血绿蛋白的 vgb 基因(来自玻璃微球菌),形成人工操纵子。与非诱导(0%阿拉伯糖)相比,携带 phaCAB-vgb(SY-2)的工程菌在 1%阿拉伯糖诱导下,细胞干重(CDW)、PHB 含量和 PHB 浓度分别提高了 1.23、1.57 和 1.93 倍。此外,通过使用携带 P(BAD)启动子-vgb 以及天然启动子-phaCAB 构建体的重组菌株,vgb 表达水平对 PHB 生物合成的影响呈正相关。

结论

这些结果利用了在大肠杆菌阿拉伯糖调控系统下融合不同物种的 phaCAB-vgb 基因来提高 PHB 产量的可能性。还表明增加 VHb 水平可以提高 PHB 的产量。

研究的意义和影响

我们成功地为大肠杆菌中的 PHB 合成提供了一个新的共表达系统。该共表达系统可受阿拉伯糖诱导剂调控,比其他诱导系统(如 IPTG)更稳定、更经济。此外,它可以应用于许多生物技术或发酵过程。

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