唾液富组蛋白5的原核表达、纯化与抗菌活性鉴定

设计一对3’端互补、5’端带酶切位点的引物,用PCR扩增唾液富组蛋白5(H5)基因(h5),将我体pGEX4T-2和h5双酶切、连接、转化DH5α,将筛选鉴定的重组质粒pGEX4T-2-h5转化BL21 (DE3).诱导重组菌BL21 (DE3) -pGEX4T-2-h5表达,纯化融合蛋白GST-H5,用Thrombin切去GST标签获取重组H5(rH5),测试rH5抗白色念珠菌(Candida albicans)活性.试验成功构建了重组载体pGEX4T-2-h5,在37℃重组菌BL21 (DE3)-pGEX4T-2-h5生长至OD600nm=0.6时用1 mmol/L IPTG于20℃诱导9h,可获得高水平表达的融合蛋白GST-H5；酶切后获得的rH5对白色念珠菌生长有较强的抑制作用,rH 5产率约2 mg/L.

Prokaryotic Expression, Purification and Antibacterial Activity of Histain5

The gene of histatin5(H5) was amplified by a pair of primers with complementary 3′ terminal and restriction site in 5′ terminal,and was cloned into pGEX4T-2.The recombinant plasmid was transformed into BL21(DE3);and the expression of soluble fusion protein GST-H5 was induced by 1 mmol/L IPTG at 20 ℃ for 9 h.The fusion protein was purified by Glutathione-sepharose 4B affinitychromatography and then digested by thrombin to obtain rH5.The results showed that rH5 had strong inhibition against Candida albicans.The yield of rH5 was 2 mg/L.