乳酸菌细菌素plantaricin JK的异源表达、分离纯化及抗菌活性

Heterologous expression and purification of plantaricin JK and analysis of its antibacterial activity

为获得高产双肽细菌素plantaricin JK,通过PCR扩增获得pln J和pln K基因,构建表达载体p ET32a(+)-pln J-BL21(DE3)和p ET32a(+)-pln K-BL21(DE3),经IPTG诱导表达后,利用镍亲和层析柱进行分离纯化,重组蛋白经肠激酶酶切纯化后,细菌素pln J和pln K的产量分别为2~3、5~8 mg·L-1。利用牛津杯平板抑菌法研究抗菌活性,结果显示细菌素pln J和pln K对柠檬色葡萄球菌(Staphylococcus citreus)、大肠埃希菌(Escherichia coli)、藤黄微球菌(Micrococcus luteus)和单核细胞增生李斯特菌(Listeria monocytogenes)都有明显抑制作用,且双肽细菌素plantaricin JK的抑菌圈直径大于单独的细菌素pln J或pln K。

In order to obtain the high-yield two-peptide bacteriocin plantaricin JK,pln J and pln K were successfully heterologously expressed in Escherichia coli BL21（ DE3）,and were induced by IPTG. Then the two peptides were expressed as His6-tag fusion proteins and were separated by Ni2 ＋chelating affinity chromatography. The fusion proteins were cleaved by enterokinase and further purified. The yields of pln J and pln K were around 2-3 and 5-8 mg·L-1. The antibacterial activity of the peptides against 4 indicator strains,i. e. Staphylococcus citreus,Escherichia coli,Micrococcus luteus,Listeria monocytogenes,was analyzed by agar diffusion method. It was shown that pln J and pln K both could inhibit the growth of the 4 indicator strains,but plantaricin JK had larger inhibition zone than pln J or pln K alone.