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Recombinant expression of the precursor of rat lung surfactant protein B in Escherichia coli and its antibacterial mechanism
Protein Expression and Purification ( IF 1.4 ) Pub Date : 2020-11-25 , DOI: 10.1016/j.pep.2020.105801
Wulong Cao 1 , Qin Liu 1 , Ting Wang 1 , Qiuhan Zhang 1 , Fu Cheng 1 , Yishan Tang 1 , Chenchen Mei 1 , Fang Wen 1 , Wanneng Wang 1
Affiliation  

While the discovery of antibiotics has made a huge contribution to medicine, bacteria that are resistant to many antibiotics pose new challenges to medicine. Antimicrobial peptides (AMPs), a new kind of antibiotics, have attracted people's attention because they are not prone to drug resistance. In this study, glutathione transferase (GST) was used as a fusion partner to recombinantly expressed rat lung surfactant protein B precursor (proSP-B) in E. coli pLySs. Cck-8 evaluated the cytotoxicity of the fusion protein and calculated its 50% inhibitory concentration (IC50). The purified peptides showed broad-spectrum antibacterial activity using filter paper method and MIC, and propidium iodide (PI) was used to explore the antibacterial mechanism against Staphylococcus aureus. In addition, the pEGFP-N2-proSP-B vector was constructed to explore the localization of proSP-B in CCL-149 cells. We found that proSP-B has obvious antibacterial activity against Gram-positive bacteria, Gram-negative bacteria and fungi, and has broad-spectrum antibacterial activity. Besides, proSP-B fusion protein has low toxicity and can change the permeability of Staphylococcus aureus cell membrane to realize its antibacterial. For these reasons, proSP-B can be used as a potential natural antibacterial drug.



中文翻译:

大鼠肺表面活性蛋白B前体在大肠杆菌中的重组表达及其抗菌机制

虽然抗生素的发现对医学做出了巨大贡献,但对许多抗生素具有抗药性的细菌对医学提出了新的挑战。抗菌肽(AMPs)作为一种新型抗生素,由于不易产生耐药性而受到人们的关注。在这项研究中,谷胱甘肽转移酶 (GST) 被用作融合伙伴,在大肠杆菌pLySs 中重组表达大鼠肺表面活性蛋白 B 前体 (proSP-B) 。Cck-8 评估融合蛋白的细胞毒性并计算其 50% 抑制浓度 (IC50)。纯化的肽段采用滤纸法和 MIC 显示出广谱抗菌活性,碘化丙啶 (PI) 用于探索对金黄色葡萄球菌的抗菌机制. 此外,构建了 pEGFP-N2-proSP-B 载体以探索 proSP-B 在 CCL-149 细胞中的定位。我们发现proSP-B对革兰氏阳性菌、革兰氏阴性菌和真菌具有明显的抗菌活性,具有广谱抗菌活性。此外,proSP-B融合蛋白毒性低,可以改变金黄色葡萄球菌细胞膜的通透性,实现其抗菌作用。由于这些原因,proSP-B 可以用作潜在的天然抗菌药物。

更新日期:2020-12-11
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