Sun Yan, Li Qian, Li Zhi, Zhang Yuan, Zhao Jie, Wang Lin
College of Life Sciences, Shaanxi Normal University, Xi'an, China.
Biosci Biotechnol Biochem. 2012;76(1):157-62. doi: 10.1271/bbb.110672. Epub 2012 Jan 7.
Antimicrobial peptides are effector molecules of the innate immunity of amphibians. Here, one antimicrobial peptide cDNA precursor, prepropalustrin-2CE3, from the tadpole of the Chinese brown frog Rana chensinensis was cloned. The coding sequence corresponding to the mature palustrin-2CE peptide was subcloned into pGEX-6p-1. The soluble GST-palustrin-2CE fusion protein was successfully expressed in the BL21(DE3)pLysS strain at 16 °C, and the proportion of the fusion protein reached 35%-39% of the total cellular protein. After removal of the GST-fusion tag, the purity of the palustrin-2CE obtained by Sephadex G50 chromatography was about 97%. Moreover, the purified palustrin-2CE displayed obviously inhibitory activities against the sensitive bacteria Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, and Escherichia coli, and multi-drug resistant S. aureus and E. coli. These findings suggest that the tadpole of the Chinese brown frog is a unique source of antimicrobial peptides and indicates the therapeutic potential of the palustrin-2CE peptide.
抗菌肽是两栖动物先天免疫的效应分子。在此，从中国林蛙蝌蚪中克隆出一种抗菌肽cDNA前体prepropalustrin - 2CE3。将对应成熟palustrin - 2CE肽的编码序列亚克隆到pGEX - 6p - 1中。可溶性GST - palustrin - 2CE融合蛋白在16℃下于BL21(DE3)pLysS菌株中成功表达，融合蛋白比例达到细胞总蛋白的35% - 39%。去除GST融合标签后，通过Sephadex G50层析获得的palustrin - 2CE纯度约为97%。此外，纯化后的palustrin - 2CE对敏感菌金黄色葡萄球菌、枯草芽孢杆菌、铜绿假单胞菌和大肠杆菌以及耐多药的金黄色葡萄球菌和大肠杆菌均表现出明显的抑制活性。这些发现表明中国林蛙蝌蚪是抗菌肽的独特来源，并显示出palustrin - 2CE肽的治疗潜力。
