抗菌肽Pexiganan和IB-367基因在大肠杆菌中的克隆与表达
发布时间：2019-04-25 14:05
【摘要】： 抗菌肽是生物体产生的一种具有强抗菌作用的阳离子多肽,是生物先天免疫的重要组成部分,是机体理想的第一道防线。抗菌肽具有分子量小、合成速度快、扩散迅速、强碱性、热稳定性高、水溶性好、抗菌机理独特等特性。抗菌肽具有广谱抗菌能力的同时,甚至对病毒及肿瘤细胞也能发挥杀灭作用,更重要的是抗菌肽对高等动物正常细胞无明显的毒害作用。 传统抗生素耐药菌株种类不断增多,全球抗生素的滥用加速了这一过程,抗菌肽的开发将为解决这一棘手难题提供更多的途径。抗菌肽所具备的药用价值使其被开发成为新一类抗感染药物成为可能。 本课题通过人工合成两种抗菌肽Pexiganan和IB-367的基因,构建了表达载体并筛选了两株表达抗菌肽的工程菌株,在大肠杆菌中分别表达了Pexiganan和IB-367与GST的融合蛋白,摸索出了融合蛋白以可溶形式大量存在于上清中的最佳诱导条件,并通过亲和层析对表达载上清中的融合蛋白进行纯化,纯化后的产物经过Factor Xa切割得到两种目的小肽——Pexiganan和IB-367。抑菌活性检测证明Pexiganan和IB-367对金黄色葡萄球菌及铜绿假单胞菌均有抑制作用。
[Abstract]:Antibacterial peptide is a kind of cationic polypeptide with strong antibacterial effect produced by organisms. It is an important part of biological innate immunity and an ideal first line of defense. Antibacterial peptides are characterized by low molecular weight, rapid synthesis, rapid diffusion, strong alkalinity, high thermal stability, good water solubility and unique antibacterial mechanism. Antibacterial peptides not only have broad spectrum antibacterial ability, but also kill virus and tumor cells. More importantly, antibacterial peptides have no obvious toxic effect on normal cells of higher animals. The abuse of antibiotics in the world has accelerated the development of antibiotic-resistant strains, and the development of antimicrobial peptides will provide more ways to solve this difficult problem. The medicinal value of antibacterial peptides makes it possible to develop a new class of anti-infective drugs. In this paper, two kinds of antibacterial peptides Pexiganan and IB-367 genes were synthesized, the expression vectors were constructed and two engineering strains expressing antibacterial peptides were screened. The fusion proteins of Pexiganan, IB-367 and GST were expressed in E. coli, respectively, and the fusion proteins of Pexiganan, IB-367 and GST were expressed in E. coli. The optimal induction conditions for fusion protein to exist in the supernatant in a large amount in soluble form were found, and the fusion protein expressed in the supernatant was purified by affinity chromatography. The purified products were cleaved by Factor Xa to obtain two small peptides-Pexiganan and IB-367.. The antibacterial activity test showed that both Pexiganan and IB-367 could inhibit Staphylococcus aureus and Pseudomonas aeruginosa.
【学位授予单位】：天津大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R346
本文编号：2465182
[Abstract]:Antibacterial peptide is a kind of cationic polypeptide with strong antibacterial effect produced by organisms. It is an important part of biological innate immunity and an ideal first line of defense. Antibacterial peptides are characterized by low molecular weight, rapid synthesis, rapid diffusion, strong alkalinity, high thermal stability, good water solubility and unique antibacterial mechanism. Antibacterial peptides not only have broad spectrum antibacterial ability, but also kill virus and tumor cells. More importantly, antibacterial peptides have no obvious toxic effect on normal cells of higher animals. The abuse of antibiotics in the world has accelerated the development of antibiotic-resistant strains, and the development of antimicrobial peptides will provide more ways to solve this difficult problem. The medicinal value of antibacterial peptides makes it possible to develop a new class of anti-infective drugs. In this paper, two kinds of antibacterial peptides Pexiganan and IB-367 genes were synthesized, the expression vectors were constructed and two engineering strains expressing antibacterial peptides were screened. The fusion proteins of Pexiganan, IB-367 and GST were expressed in E. coli, respectively, and the fusion proteins of Pexiganan, IB-367 and GST were expressed in E. coli. The optimal induction conditions for fusion protein to exist in the supernatant in a large amount in soluble form were found, and the fusion protein expressed in the supernatant was purified by affinity chromatography. The purified products were cleaved by Factor Xa to obtain two small peptides-Pexiganan and IB-367.. The antibacterial activity test showed that both Pexiganan and IB-367 could inhibit Staphylococcus aureus and Pseudomonas aeruginosa.
【学位授予单位】：天津大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R346
【引证文献】
相关博士学位论文 前1条
1 唐勇;黄鳝肠道cDNA文库构建、EST分析及抗菌肽基因的原核表达[D];四川农业大学;2010年
，本文编号：2465182
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