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在大肠杆菌和其应用中,细胞质二烯酸酶的高效分泌表达

Lan Qin¹, Youhao Tian², Shuaishuai Zhao²

¹State Key Laboratory of Biobased Material and Green Papermaking, Qilu University of Technology, Shandong Academy of Science, Jinan 250353, PR China; School of Bioengineering, Qilu University of Technology, Shandong Academy of Science, Jinan 250353, PR China.

International journal of biological macromolecules

|March 19, 2025

概括

这项研究通过使用信号,在大肠杆菌中实现了细胞质2-酶 (CE) 的直接细胞外分泌. 优化的发酵条件产生了高酶产量,在特定反应参数下实现了56%的乳糖转化率.

科学领域:

生物技术是生物技术.
酶工程是什么? 酶工程是什么?
微生物发酵微生物发酵

背景情况:

纤维素2-表皮酶 (CE) 对于乳糖转化至关重要.
重组蛋白表达通常需要有效的分泌策略.
直接的细胞外分泌简化了下游处理,降低了成本.

研究的目的:

通过使用信号来实现细胞质2-酶 (CE) 的直接细胞外分泌.
为了优化发酵条件,提高复合CE生产.
为了确定最大限度的乳糖转化最佳反应条件.

主要方法:

一个信号 (N20) 与 Caldicellulosiruptor bescii. 的 CE 基因的融合.
在使用pET-22b(+) 载体的Escherichia coli BL21 (DE3) 中进行重组表达.
优化发酵,通过硫酸沉净化酶,以及活性测定.
高性能液体色谱 (HPLC) 用于分析乳糖转化率.

主要成果:

通过SDS-PAGE证实了CE的成功细胞外分泌.
确定了最佳发酵条件:2xYT介质,0.8毫米IPTG,0.1毫米Fe2+,25毫米甘氨酸,37°C,每分钟200转,36小时化后-2.5小时诱导.
纯化CE产量为1529.61毫克/升,特异活性为19.25单位/毫克.
在24小时内以1.53g/L的酶度达到56%的最大乳糖转化率.

结论:

在大肠杆菌中成功实现了CE的直接细胞外分泌.
优化的发酵和反应条件使得生产效率高,转化率高.
这项研究为工业生产和CE的应用提供了基础.

关键词:

蜂素2-聚酶酶是什么?埃舍里希亚大肠杆菌 (Escherichia coli) 是一个大肠杆菌.细胞外分泌的细胞外分泌物信号的信号.

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