基质金属蛋白酶抑制剂-4在大肠杆菌中的克隆与表达

基质金属蛋白酶抑制剂4(tissueinhibitorofmetalloproteinase4,TIMP4)为心脏特异性表达,在心脏代谢和肿瘤转移、侵袭过程中具有重要作用。为进一步研究其功能和作用机理,以PCR方法扩增人基质金属蛋白酶抑制剂4不含信号肽的表达序列,测序鉴定正确后,构建原核重组表达质粒pMALc2TIMP4,转化大肠杆菌,IPTG诱导阳性克隆,大量表达重组蛋白。细菌经超声破碎后,其上清中的重组蛋白经SDSPAGE初步鉴定分子量大小与理论值一致,为以后基质金属蛋白酶抑制剂4的研究创造了条件 。

Cloning and Expression of Tissue Inhibitor of Metalloproteinase-4 in E.coli

Tissue inhibitor of metalloproteinase-4 (TIMP-4) is only expressed in heart, and play important role in heart metabolism, tumor invasion and metastasis. To further study the mechanism of its structure and function, the coding sequence of TIMP-4 without signal peptide has been obtained by PCR amplifying, and the sequencing analysis demonstrates that the sequence is correct. The recombinant expression plasmid pMAL-c2/ TIMP-4 containing TIMP-4 is constructed and transformed the E.coli. The recombinant tissue inhibitor of metalloproteinase-4 is expressed in great deal after the induction by IPTG, the fused recombinant protein is released to the supernatant of the ultrasonic-fragmentated E.coli and its molecular weight is identical with natural TIMP-4 in SDS-PAGE.;