家蝇Cecropin A分子体外表达及抗菌活性鉴定

Expression and identification of cecropin A from Musca domestica

目的体外重组表达家蝇天蚕素多肽分子,并对重组表达产物活性进行初步鉴定。方法克隆家蝇天蚕素成熟肽(Mature Cecropin,MC)基因,构建pET32a-MC重组质粒。转化大肠杆菌(E.coli)BL21(DE3)中。1mmol/L终浓度的IPTG诱导重组蛋白Thioredoxin(硫氧还蛋白,Trx)-MC表达。重组蛋白质纯化回收后经胃蛋白酶水解获得天蚕素多肽分子。用重组蛋白和天蚕素多肽分子进行抑菌试验鉴定其生物活性。结果 MC多肽分子体外具有较强的抑菌活性,且抑菌活性持续时间长。结论硫氧还蛋白在重组蛋白质中作为分子伴侣改变了天蚕素多肽分子生物活性,使其对宿主菌低毒性。MC多肽分子的体外长效抑菌能力使其具有良好的应用前景。

We conducted expression of cecropin A from the Musca domesticaand identified its activity in vitro.Housefly cecropin mature peptide gene was cloned,and pET32a-mc plasmid was constructed.The vector pET32a-mc was transformed into BL21(DE3).After induction with IPTG,the Trx&MC fusion protein was purified and digested with pepsin in buffer.The hydrolyzate was analyzed with tricine-SDS-PAGE.The antibacterial activity of MC was detected by liquid growth inhibition.Results indicated that MCinvitrohas strong and long duration antibacterial activity.Thioredoxin,as a molecular partner,could affect MC biological activity and reduce toxicity to bacteria,suggesting that MC molecules will have a good prospect of application.