从Lactococcus lactis LAC460 LAC460中菌素素样细菌素LysL的异质表达和净化
Samira Mokhtari1, Per E J Saris1, Timo M Takala1
1Department of Microbiology, University of Helsinki, PO Box 56, FI-00014 Helsinki, Finland.
FEMS microbiology letters
|August 17, 2024
概括
研究人员探索了在各种宿主中产生LysL的方法,LysL是一种来自Lactococcus lactis的类似细菌素的菌素. 大肠杆菌提供了最成功的方法,为进一步的功能研究提供足够的活性lysl.
科学领域:
- 微生物学 微生物学
- 分子生物学分子生物学
- 蛋白质表达方式
背景情况:
- 乳球球菌乳糖LAC460自然产生类似细菌素的菌素,LysL和LysP.
- LysL 是异质生产和功能研究的目标.
研究的目的:
- 在各种异质宿主和体外无细胞系统中产生和分泌lysl.
- 为了优化LysL的生产,以进行后续的功能分析.
主要方法:
- 将lysL基因克隆成乳球菌cremoris NZ9000和L. lactis N8具有或没有usp45信号序列.
- 研究LysL表达和活性在复合乳球菌株.
- 使用一个体外无细胞表达系统.
- 在大肠杆菌BL21(DE3) 中生产和净化用Ni-NTA亲和力染色法标记的His-标记的LysL.
主要成果:
- LysL 主要是在 L. lactis N8 中细胞内产生,分泌量有限.
- 再组合的L. cremoris NZ9000在诱导时被溶解,证实了lysL的表达.
- 与USP45信号的融合降低了LysL的活性.
- 从乳球菌宿主和体外表达物中获得了低产量的活性lysl.
- 成功地从大肠杆菌中生产和净化了4.34毫克的被标记为His的活性lysl.
结论:
- 在乳球菌株中LysL的异质表达带来了分泌和活性方面的挑战.
- 大肠杆菌是生产大量活性lysL的合适宿主.
- 纯化的His标记的LysL可用于未来的功能研究.
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