猪肺炎支原体P159蛋白的截短表达及黏附活性研究

Truncated expression of P159 of Mycoplasma hyopneumoniae and identification of its adhesion activity

为了解猪肺炎支原体(Mycoplasma hyopneumoniae,Mhp)P159蛋白的黏附活性,本实验利用PCR从Mhp NJ株扩增P159基因片段(3'端),克隆于pET-28a(+)进行表达,并用表达的截短蛋白进行黏附试验。结果表明,PCR扩增的目的基因片段约为450 bp;SDS-PAGE检测重组蛋白分子量为46 ku;western blot检测表明该重组蛋白能够与Mhp阳性血清发生特异性反应,表明该蛋白具有良好的免疫反应原性;表达的截短蛋白与Mhp的黏附试验结果表明,该重组蛋白可以粘附于猪肺上皮细胞(SJPL)表面,并部分抑制Mhp对SJPL的粘附作用。本研究结果为Mhp致病机制的进一步研究奠定了基础。

To investigate the Mycoplasma hyopneumoniae （Mhp） P159 adhesion activity, the 3＇ end of P159 gene fragment was amplified by PCR from Mhp NJ strain, and inserted into expression vector pET-28a（＋） for expression in E.coli. The results showed that the target gene was 450 bp. Molecular weight of the recombinant protein was 46 ku by SDS-PAGE detection and westem blot analysis indicated that the recombinant protein had specific reaction with pig anti Mhp serum. Furthermore, the recombinant protein was able to adhere to porcine lung epithelial cells and partially blocked the Mhp to adhere the cell which was specially inhibited by anti Mhp serum. The results provide a basis for the figther study on the pathogenesis of Mhp.