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通过转录组分析,探索和设计DB104中Bacillus subtilisDB104中高水平蛋白质表达的新型强促剂.
Ji-Su Jun1, Hyang-Eun Jeong1, Kwang-Won Hong1
1Department of Food Science and Biotechnology, College of Life Science and Biotechnology, Dongguk University, Goyang-si 10326, Republic of Korea.
Microorganisms
|December 23, 2023
概括
这项研究设计了一个强大的Bacillus subtilis表达系统,用于快速,无诱导剂的重组蛋白质生产. 像PSDP-4这样的优化促进剂在24小时内实现了生物活性人体表皮生长因子 (hEGF) 的高产量.
科学领域:
- 微生物生物技术和合成生物学.
- 再组合蛋白的表达和优化.
背景情况:
- 细菌是重组蛋白质生产的关键宿主.
- 缺少蛋白酶的衍生品DB104,为细胞外蛋白质表达提供了优势.
- 转录组数据为促进体发现提供了对高度表达的基因的洞察.
研究的目的:
- 为了识别和表征来自Bacillus subtilis DB104转录组数据的强促进体.
- 为改善重组蛋白质表达而设计增强型促进体.
- 为工业应用开发一种高效,无诱导剂的表达系统.
主要方法:
- 对 Bacillus subtilis DB104.4 的时间过程转录组分析.
- 使用eGFP报告员识别高度表达的基因并评估它们的促进体强度.
- 通过对监管元素和序列元素的优化进行推动者工程.
- 构建和评估人类表皮生长因子 (hEGF) 的表达盒.
主要成果:
- 与对照P43促进剂相比,PSDP和PskfA促进剂的强度显著更高.
- 改造后的PSDP-4促进剂的强度比原始PSDP增加了3.84倍.
- Psdp-4磁带在24小时内实现了His标记的hEGF的103.9微克/毫升的高产量.
- 表达的hEGF蛋白被净化,并通过细胞增殖试验证实具有生物活性.
结论:
- 一个高效的,转录基因驱动的表达系统 Bacillus subtilis DB104 已成功构建.
- 设计的PSDP-4促进体使得蛋白质快速表达,没有诱导剂.
- 该系统是工业规模重组蛋白质生产的宝贵工具.
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