Novel signal peptides improve the secretion of recombinant Staphylococcus aureus Alpha toxin(H35L) in Escherichia coli.
Novel signal peptides improve the secretion of recombinant Staphylococcus aureus Alpha toxin(H35L) in Escherichia coli.
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DOI：
10.1186/s13568-017-0394-1
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发表时间：
2017-12
期刊：
影响因子：
3.7
通讯作者：
Schoner R
中科院分区：
文献类型：
--
作者：
Han S;Machhi S;Berge M;Xi G;Linke T;Schoner R
将异源蛋白质分泌到大肠杆菌细胞培养基中，为下游加工提供了优于包涵体生产的显着优势;包括节省成本和时间，并减少内毒素。信号肽在靶向蛋白跨细胞质膜易位至周质空间并在分泌过程中释放至培养基中中起重要作用。选择α毒素H35 L（ATH 35 L）作为针对金黄色葡萄球菌感染的疫苗开发的抗原。成功地将其分泌到E.大肠杆菌通过使用连接到蛋白质的N-末端的细菌信号肽。为了提高分泌型ATH 35 L的水平，我们通过交换修饰dsbA和pelB信号肽的各个结构域来设计一系列新的信号肽，并在补料分批发酵过程中测试它们。结果表明，部分信号肽的修饰提高了ATH 35 L的分泌效率。来自dsbA、pelB和phoA蛋白的大肠杆菌信号肽。事实上，一种新的信号肽在补料分批发酵过程中将分泌的ATH 35 L的产率提高了3.5倍，同时在信号肽切割的预期位点保持加工。这些新的信号肽可用于提高其他异源蛋白在大肠杆菌中的分泌效率。杆菌此外，对合成信号肽氨基酸序列的分析提供了对影响分泌效率的信号肽内的序列特征的一些了解。本文的在线版本（doi：10.1186/s13568-017-0394-1）包含补充材料，可供授权用户使用。
Secretion of heterologous proteins into Escherichia coli cell culture medium offers significant advantages for downstream processing over production as inclusion bodies; including cost and time savings, and reduction of endotoxin. Signal peptides play an important role in targeting proteins for translocation across the cytoplasmic membrane to the periplasmic space and release into culture medium during the secretion process. Alpha toxinH35L (ATH35L) was selected as an antigen for vaccine development against Staphylococcus aureus infections. It was successfully secreted into culture medium of E. coli by using bacterial signal peptides linked to the N-terminus of the protein. In order to improve the level of secreted ATH35L, we designed a series of novel signal peptides by swapping individual domains of modifying dsbA and pelB signal peptides and tested them in a fed-batch fermentation process. The data showed that some of the modified signal peptides improved the secretion efficiency of ATH35L compared with E. coli signal peptides from dsbA, pelB and phoA proteins. Indeed, one of the novel signal peptides improved the yield of secreted ATH35L by 3.5-fold in a fed-batch fermentation process and at the same time maintained processing at the expected site for signal peptide cleavage. Potentially, these new novel signal peptides can be used to improve the secretion efficiency of other heterologous proteins in E. coli. Furthermore, analysis of the synthetic signal peptide amino acid sequences provides some insight into the sequence features within the signal peptide that influence secretion efficiency. The online version of this article (doi:10.1186/s13568-017-0394-1) contains supplementary material, which is available to authorized users.