猪CD58分子基因克隆、表达及其结构功能预测

Porcine CD58: cDNA Cloning,Expression and Prediction of the Structure and Function

CD58在机体免疫系统中具有重要作用,本研究通过对人、绵羊CD58 mRNA序列比对,设计兼并引物,应用反转录PCR技术克隆猪CD58基因,并在大肠杆菌中进行原核表达,同时运用生物信息学方法对其核苷酸序列、编码的氨基酸序列以及蛋白结构进行预测。结果表明:克隆的猪CD58 cDNA全长800 bp,ORF为735 bp;将其在大肠杆菌中进行表达,产物可被CD58抗血清识别;序列比对结果显示猪、羊和人的CD58核苷酸序列及氨基酸序列同源性并不高,但蛋白结构预测表明三者蛋白结构非常相似,尤其是V区三维结构,这是异种动物淋巴细胞和红细胞发生黏附的分子基础。该研究为CD58作为疫苗佐剂或免疫调节药物在临床中的应用、进一步研究CD58分子结构及CD2-CD58复合物激活免疫系统的机理奠定了基础。

CD58 plays an important role in mammal immune system. In this study, a pair of primers was designed according to human and sheep CD58 sequence and used to clone porcine CD58 gene by RT-PCR method. In addition, the protein structure of porcine CD58 were predicted through bioinformatics methods and compared with human and sheep CD58. The results showed that an 800 bp fragment of porcine CD58 cDNA with an ORF of 735 bp was obtained. Then, this cDNA was ligated into pGEX-4T-1 and transformed to E. coli BL21 cells. The recombinant transformant was induced with IPTG and the expressed product could be recognized by anti-CD58 serum. The prediction of the protein structure indicated that the porcine, sheep and human CD58 have high similarity, especially in V region which was the molecular foundation of xeno-adhesion between lymphocyte and RBC. This research will help us to develop new adjuvant and/or immune-regulated drugs, and laid a solid foundation for further study of structure of CD58 molecule and the mechanism of CD2-CD58 complex triggering immune system.