绵羊生殖道抗菌肽

摘要

The female sheep reproductive tracts were freshly collected from a local meat processing plant and used as experimental materials. Two antibacterial peptides were isolated and characterized from female sheep reproductive tracts by two consecutive chromatographic steps. The peptide isolation procedures included acetic acid extraction, dialyzed, gel filtration chromatography on Sephadex G-50, and reverse phase high-performance liquid chromatography (RP-HPLC). Their molecular mass were 4 820.47 u and 4 012.5 u, respectively, analyzed by MALDI-TOF-MS. The partial N-terminal amino acid sequences of two peptides were determined as AYVLDEPKP and YDSGA, respectively, by Edman degradation. The antimicrobial activity was tested during each purification step by the radial diffusion plate assay and broth microdilution method. These two peptides showed good antimicrobial activities against reference strains of G~+(S. Aureus ATCC2592 and Streptococcu ATCC55121), G~-(E. Coli ATCC25922) and fungi(C. Albicans ATCC2002). The peptides did not show active hemolytic activity against rabbit blood red cells and had no significant effects on human blood coagulation system. The discovery of antibacterial peptides in sheep reproductive system reveals that antibacterial peptides may play a role in innate immunity against microorganisms in a wide range of animal species.%以屠宰场收集的新鲜、健康、雌性绵羊牛殖器官为原材料.采用乙酸浸提、透析、Sephadex G-50凝胶过滤层析和反相高效液相色谱(RP-HPLC)等方法分离纯化绵羊生殖道抗菌肽.以G+、G-和真菌为抗菌活性检测指示菌株,利用薄层琼脂糖孔穴扩散法、微量肉汤稀释法进行抗菌活性检测.对分离纯化所得纯品进行分子质量质谱测定、纯度鉴定、N端测序,并对其性质进行研究.结果表明:分离纯化所得两个绵羊生殖道抗菌肽分子质量分别为4820.47 u和4012.5 u,N端部分氨基酸序列分别为AYVLDEPKP和YDSGA.对G+细菌(S.aureus ATCC2592、Streptococcu ATCC55121)、G-细菌(E.coli ATCC25922)、真菌(C.albicans ATCC2002)均具有良好的抑菌活性.对家兔红细胞无溶血活性,对人血液凝固无影响.目前未见有从绵羊生殖道分离纯化得到抗菌肽的报道,并且这一研究结果进一步证实抗菌肽在多种动物生殖道天然免疫防御方面起着重要作用.