牛乳铁蛋白肽衍生肽结构设计及其在毕赤酵母中表达后的活性分析

Structural Design of Bovine Lactoferricin-derived Peptide and Its Activity Assay after Expressed in Pichia pastoris

本研究通过抗菌肽数据库和生物信息学等工具对牛乳铁蛋白肽衍生肽(LfcinBD)的结构进行优化设计,将衍生肽LfcinB-W4的基因片段在毕赤酵母GS115表达。发酵实验结果表明当甲醇诱导浓度为2.5%,发酵时间为96h时,发酵上清液对金黄色葡萄球菌和沙门氏菌具有很强的抑制作用,其效果与50mg/mL的氨苄青霉素抑菌效果相当,高于LFcinB。发酵上清液冷冻干燥浓缩后,10倍的发酵上清浓缩液其抑菌效果为同体积50mg/mL的氨苄青霉素的2.5倍。用Tricine-SDS-PAGE检测到相对分子质量约为3.2kDa的衍生肽LFcinB-W4。实验通过生物信息学工具设计得到抑菌活性高于LFcinB的衍生肽,为下一步分析牛乳铁蛋白肽功能与结构关系奠定了基础。

In this study,the structure of bovine lactoferricin peptide-derived( LfcinBD) was optimized by antimicrobial peptide database and bioinformatics tools,and the gene fragment of the LfcinB-W4 was expressed in Pichia pastoris GS115.The results of fermentation experiments showed that when the methanol induction concentration was 2.5% and the fermentation time was 96 h,the fermentation supernatant had a strong inhibitory effect on Staphylococcus aureus and Salmonella,and its effect was compared with the antibacterial effect of 50 mg /mL ampicillin.After the fermentation supernatant was freeze - dried and concentrated,the bacteriostatic effect of the 10 - fold concentrate was 2.5 times that of the same volume of 50 mg /mL ampicillin.LFcinB - W4 with a relative molecular mass of about 3.2 kDa was detected by Tricine - SDS - PAGE.In this experiment,bio-informatics tools were designed to obtain derived peptides with higher antibacterial activity than LFcinB,which laid a foundation for further analysis of the relationship between bovine lactoferrin peptide function and structure.