利用ELP自断裂标签在大肠杆菌中生产抗菌肽Oxysterlin 1

Production of antimicrobial peptide (Oxysterlin 1) in Escherichia coli with ELP self-cleavage tag

摘要:

抗菌肽是目前最有希望的抗生素替代品，但是使用重组技术生产抗菌肽的策略大多步骤烦琐且价格昂贵，不利于抗菌肽的规模化生产。Oxysterlin 1是一种新型的天蚕素抗菌肽，主要对革兰氏阴性菌有抗菌活性，具有较低的细胞毒性。文中利用一种简单经济的方法在大肠杆菌中实现Oxysterlin 1的表达和纯化。将Oxysterlin 1基因克隆到含有弹性蛋白样多肽Elastin-like polypeptide (ELP) 和蛋白质内含肽 (Intein) 的载体中，构建重组表达质粒pET-ELP-I-Oxysterlin 1。重组蛋白在大肠杆菌中主要以可溶性形式表达，进而通过简单的盐析和pH改变便可对目标小肽进行纯化。最终得到的Oxysterlin 1的产量约为1.2 mg/L，抑菌试验显示出预期活性，为抗菌肽的规模化生产及深入研究其抑菌机理奠定基础。

Abstract:

Antimicrobial peptides are the most promising alternatives to antibiotics. However, the strategy of producing antimicrobial peptides by recombinant technology is complicated and expensive, which is not conducive to the large-scale production. Oxysterlin 1 is a novel type of cecropin antimicrobial peptide mainly targeting on Gram-negative bacteria and is of low cytotoxicity. In this study, a simple and cost-effective method was developed to produce Oxysterlin 1 in Escherichia coli. The Oxysterlin 1 gene was cloned into a plasmid containing elastin-like polypeptide (ELP) and protein splicing elements (intein) to construct the recombinant expression plasmid (pET-ELP-I-Oxysterlin 1). The recombinant protein was mainly expressed in soluble form in E. coli, and then the target peptide can be purified with a simple salting out method followed by pH changing. The final yield of Oxysterlin 1 was about 1.2 mg/L, and the subsequent antimicrobial experiment showed the expected antimicrobial activity. This study holds promise for large-scale production of antimicrobial peptides and the in-depth study of its antimicrobial mechanism.