摘要
目的将儿茶酚胺氧位甲基转移酶基因(COMT)克隆到原核表达载体进行可溶性表达,制备纯化COMT蛋白,为深入研究COMT的功能提供材料。方法运用分子生物学方法构建融合表达质粒Pet22b+-COMT。将Pet22b+-COMT转入E.coli BL21(DE3)表达菌株,利用IPTG诱导表达,并用亲和色谱纯化COMT。结果经测序分析成功构建了融合表达质粒Pet22b+-COMT。COMT基因在E.coli BL21(DE3)中表达相对分子质量约为28000的蛋白质,纯化后,COMT蛋白的纯度大于90%。结论COMT基因在原核中得到良好的表达,得到了纯度较高的蛋白质,为酶学活性研究奠定了基础。
Purpose To clon the human soluble catachol-O-Methyltransferase（COMT） into the Pet22b^＋ vector, expression and purification the COMT protein for researching the function of COMT. Methods After constructing Pet22b^＋ -COMT plasmid, transferred it into E. coli BL21（DE3）, then induced by IPTG,and pudried by affinity chromatography. Results It is success to build a fusion expresion vector Pet22b^＋ -COMT analyzed by nucleotide sequencing. COMT gene express 28 000 of protein in E. coli BL21 （DE3）, through puring, the purity of COMT protein can be more than 90%. Conclusion COMT gene was expressed well in BL21 （DE3） E. coli, and highly purified COMT protein are obtained.
出处
《中国生化药物杂志》
CAS
CSCD
2007年第1期17-19,共3页
Chinese Journal of Biochemical Pharmaceutics
