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从Cupriavidus necator H16中的高度稳定的表达等离子体中基于CO2生产植物酶

Simon Arhar¹, Thomas Rauter¹, Holly Stolterfoht-Stock¹

¹Austrian Centre of Industrial Biotechnology, acib GmbH, Krenngasse 37, Graz, 8010, Austria.

Microbial cell factories

|January 3, 2024

概括

我们为Cupriavidus necator开发了新的质粒,可以提高基因表达的稳定性,并允许电穿孔. 这使得有效的植物酶生产成为可能,支持料行业的二氧化碳利用.

科学领域:

合成生物学合成生物学
微生物生物技术微生物生物技术
代谢工程是代谢工程.

背景情况:

现有的Cupriavidus necator等离子体在结合和稳定性方面存在局限性.
低等离子体稳定性和复制数阻碍了无选择的异质蛋白质的产生.
在C. necator中植物酶的产生对于酸盐在动物料中的可用性和二氧化碳的价值化是可取的.

研究的目的:

通过电穿孔开发一种新的,稳定的,高复制数表达等离子体系统,用于C. necator.
为了证明这个系统对异质植物酶生产的实用性.
为了优化植物酶表达,使用各种促进剂和种植方法.

主要方法:

设计和制造了具有分区系统的新型表达等离子体,以提高分离稳定性.
在C. necator H16 PHB-4中引入了来自大肠杆菌的AppA植物酶基因.
利用七种不同的促进剂 (构成性,酶,卡尔文-本森-巴沙姆循环) 进行植物酶表达.
进行了异构和自构培养,包括1L料批量气体发酵.

主要成果:

在C. necator H16中使用新型电穿孔等离子体成功产生活跃的AppA植物酶.
实现了植物酶活动,总蛋白质从2到50U/mg不等,取决于促进物和栽培.
在1L的料批次系统中成功升级,达到22U/ml的植物酶活性.

结论:

这种新型表达系统为C. necator.的蛋白质生产提供了一个高效的平台.
强大的促进器允许定制基因表达,包括在生长阶段过渡期间的诱导活性.
在C. necator H16中活性植物酶的生产显示出在料行业中利用二氧化碳的潜力.

关键词:

这就是Cupriavidus necator.电穿孔是一种电子穿孔.气体发酵发酵过程中的气体.植物酶是一种植物酶.活动主办人活动主办人分离的稳定性分离的稳定性

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