Mycoplasma mycoides subsp.mycoides Ben-1株一个新的膜蛋白的特性研究  

Identification of a novel membrane protein in the Mycoplasma mycoides subsp.mycoidessmall colony Ben-1 strain

作　　者：周玉梅[1] 汪洋[1] 李媛[1] 邹晓辉[1] 刘素丽[1] 白帆[2] 吴金迪[3] 辛九庆[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/国家牛传染性胸膜肺炎指定检测实验室,黑龙江哈尔滨150001 [2]吉林农业大学动物科学技术学院,吉林长春130118 [3]东北农业大学动物医学院,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2014年第5期354-358,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金(31072131)

摘　　要：为研究Mycoplasma mycoides subsp.mycoides(Mmm)Ben-1弱毒疫苗传代致弱的机制,本研究通过比较Mmm Ben-1株和其兔体内传代致弱毒株Ben-470的全基因组序列,发现在Ben-470株中缺失了一个编码165个氨基酸(分子量约19 ku)的495 bp的假定蛋白基因,命名为p588,扩增该基因,并表达重组P588(rP588)蛋白,制备兔抗血清。经细菌膜蛋白不同组份的提取及western blot鉴定,结果显示P588是一种膜蛋白,并且rP588与CBPP国际标准血清呈阳性反应。通过激光共聚焦显微镜观察到rP588对牛肺细胞(EBL)具有明显的粘附作用,并且ELISA试验也证明该蛋白的这种粘附为特异性粘附。上述结果表明P588蛋白是Mmm Ben-1的一个粘附蛋白。Mycoplasma mycoides subsp. mycoides （Mmm） is the causative agent of contagious bovine pleuropneumonia （CBPP） which had been eliminated by vaccination of attenuated vaccine of the Ben-1 strain since early 1950’s in China. To investigate the mechanism of the Mmm attenuation, we compared the entire genomes of the Ben-1 strain and the attenuated Ben-470 strain and revealed that an open reading frame of 495 bp （designed p588 gene） coding for a putative protein of 165 amino acids （about 19 ku） was absent from the Ben-470 strain. Then the p588 gene was cloned from virulent strain of Ben-1 and the recombinant P588 （rP588） was expressed in E.coli. In addition, western blot analysis demonstrated that the P588 protein was a membrane-associated protein and reacted positively with the international standard serum against CBPP. Furthermore, the rP588 was able to adhere to embryonic bovine lung cells detected by immunostaining visualised via confocal laser scanning microscopy, and this was also confirmed by a sandwich ELISA. In conclusion, the P588 protein, which exists in Ben-1 strain but deleted from the Ben-470 strain during the attenuated process, is an adhesion protein in Mmm.

关 键 词：牛传染性胸膜肺炎 基因P588 粘附 

分 类 号：S852.61[农业科学—基础兽医学]