论文部分内容阅读
目的在毕赤酵母中表达抗菌肽Catesbeiania-1,并检测其体外生物活性。方法将从牛蛙皮肤上新分离的一条抗菌肽基因Catesbeiania-1克隆至真核表达载体pPICZaA,构建重组真核表达质粒pPICZaA-Catesbeiania-1。在毕赤酵母GS115中表达重组Catesbeiania-1蛋白,并进行体外生物活性测定。结果所构建的重组表达质粒pPICZaA-Catesbeiania-1序列完整,经Tricine-SDS-PAGE分析,表达的重组Catesbeiania-1蛋白相对分子质量约为5800,分泌性蛋白的表达量为36.3mg/ml。重组蛋白对几种常见的革兰阳性菌和阴性菌均具有较好的抑菌活性,无胰蛋白酶水解活性和抑制剂活性,具有很强的抗氧化能力。结论已成功表达了抗菌肽Catesbeiania-1,表达的重组蛋白具有一定的抑菌活性。
Objective To express the antibacterial peptide Catesbeiania-1 in Pichia pastoris and test its in vitro biological activity. Methods An antimicrobial peptide gene Catesbeiania-1 newly isolated from the skin of bullfrog was cloned into the eukaryotic expression vector pPICZaA to construct the recombinant eukaryotic expression plasmid pPICZaA-Catesbeiania-1. Recombinant Catesbeiania-1 protein was expressed in Pichia pastoris GS115 and in vitro bioassay was performed. Results The constructed recombinant plasmid pPICZaA-Catesbeiania-1 had a complete sequence. The relative molecular mass of the recombinant Catesbeiania-1 protein was about 5800 and the secreted protein was 36.3 mg / ml after Tricine-SDS-PAGE analysis. Recombinant proteins have good antibacterial activity against several common Gram-positive and -negative bacteria, without trypsin hydrolysis activity and inhibitor activity, and have strong antioxidant capacity. Conclusion The antimicrobial peptide Catesbeiania-1 has been successfully expressed and the expressed recombinant protein has certain antibacterial activity.