【作者】 钟雅；

【导师】 陈正望；

【作者基本信息】 华中科技大学 ， 生物化学与分子生物学， 2007， 硕士

【摘要】 抗菌肽是生物体内产生的一种具有抗菌功能的小分子多肽,是生物先天非特异性免疫的重要组成部分。这类活性多肽具有分子量小、热稳定性强、水溶性好、抗菌谱广、杀菌活力高的特点,能杀灭对传统抗生素产生抗性的菌株,其杀菌机制使病原菌不易产生耐药性突变,使其可能成为抗生素的理想替代物。此外,抗菌肽对一些病毒和寄生虫也有抑杀作用;还能特异性地抑制某些肿瘤细胞的生长,且对人体正常细胞无害,这给抗癌药物的开发带来了新的希望。抗菌肽作为一大类新型抗菌机理的天然多肽抗生素和抗癌药物,具有广泛的应用前景。寻找新的抗菌肽已成为生命科学领域中的热门研究课题之一。本论文从野生家蝇中分离出一个新抗菌肽,并对其部分性质进行了研究。家蝇(Musca domestica)属昆虫纲,双翅目,环裂亚目、蝇科,是我国大部分地区最常见、数量最多的一种蝇类。家蝇中存在大量抗菌活性物质,对抗菌药物的开发具有重要的意义。但家蝇中抗菌肽的种类较多,成分复杂,很难分离纯化出一个单一抗菌肽,目前国内外对其研究较少。本研究利用短时沸水处理、稀醋酸低温浸提、海藻酸吸附、稀盐酸低温洗脱、盐析、Sephadex G25凝胶过滤、CMC23弱阳离子交换柱层析、反相高效液相色谱层析、电洗脱回收等方法,结合高灵敏度的杀菌活性检测手段,从家蝇蛆中分离纯化出一个20 kD的新抗菌肽,对革兰氏阳性菌如苏云金芽孢杆菌( Bacillus thuringiensis)、枯草芽胞杆菌( Bacillus subtilis )、金黄色葡萄球菌( Staphylococcus aureus )均有杀灭作用。采用胞内荧光测钙技术,发现mM级浓度的此抗菌肽能引起大鼠胰腺β-细胞胞内钙浓度升高,且这种效应具有持续性。更多还原

【Abstract】 Antibacterial peptides are small Mr peptides with antibacterial function produced by organisms, which are important components of innate immunity system in organisms. These peptides have characters of highly thermal stability, well water solubility, broad antibacterial spectrum and strong bactericidal activity. Some of them can kill bacteria which have resistance against traditional antibiotics, and the bactericidal mechanism impossibly makes the pathogenic bacteria occur resistance mutation. These advantages could make antibacterial peptides become perfect substitutes of antibiotics. In addition, some antibacterial peptides can inhibit or kill some virus and parasite. Furthermore, some of them can inhibit some tumour cell growing, and are harmless to human normal cell. It brings new hope for the development of anticancer drugs. Antibacterial peptide has extensive applied perspective as a natural peptide antibiotic with new bactericidal mechanism and a new anticancer drug. It has become a popular research subject in life science field to find new antibacterial peptides.Musca domestica, which belongs to insecta, diptera, cyclorrhapha, muscidae, is the most common muscae and the richest resource in our country. It is very significant and valuable to separating antibacterial peptides from Musca domestica and to develop these peptides into antibacterial medicine. Due to purify a pure peptide from natural material is very difficult, few research is in progress. Based on a highly sensitive bactericidal detection method, using dilute acetic acid extraction, alginic acid absorption, NaCl salting-out, Sephadex G25 gel filtration, CMC23 ion-exchange chromatography, RP-HPLC and electro-elution, a novel 20 kD antibacterial peptide had been purified from Musca domestica dry larvae. The peptide has antibacterial activity against Gram-positive bacteria such as Bacillus thuringiensis, Bacillus subtilis and Staphylococcus aureus. Using intracellular Ca2+ fluorimetry technique, it has been detected that the antibacterial peptide in mM make the intracellular Ca2+ concentration of pancreasβ-cell increase. And the effect is durative and not related to the dose of the antibacterial peptide.更多还原