可溶性尿激酶受体的表达、纯化和结晶  

The Expression, Purification and Crystallization of Monomeric Soluble Human Urokinase Receptor

作　　者：袁彩[1,3] 淮清 卞传兵[1,3] 黄明东[1] 

机构地区：[1]中国科学院福建物质结构研究所,结构化学国家重点实验室 [2]Center for Haemostasis and Thrombosis Research,Beth Israel Deaconess Medical Center,Harvard Medical School,Boston,MA 02215,USA [3]中国科学院研究生院,北京100049

出　　处：《生物化学与生物物理进展》2006年第3期277-281,共5页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金重点项目(30430190)~~

摘　　要：克隆尿激酶受体可溶区域(suPAR)到果蝇胚胎细胞分泌表达载体pMT/Bip/v5-his,重组质粒与pCoHygro共转染果蝇S2细胞,筛选多拷贝稳定表达细胞系suPARS2.suPAR表达蛋白经尿激酶N端片段亲和柱、ResourceQ阴离子交换柱两步纯化后,得到高纯度的、稳定的suPAR单体.纯化后的蛋白质,与其抗体ATN615及尿激酶N端片段结构域等摩尔混合,浓缩至10g/L,以透析法进行该三元复合物晶体生长,获得衍射分辨率为1.9#的蛋白质晶体.Soluble form of human urokinase receptor was cloned into Drosophila Schneider 2 secretion expression vector pMT/Bip/vS-his, and co-transfected with pCoHygro into S2 cells, to establish stably S2 cells line. The expressed suPAR from such procedure tends to form oligomer and is unstable, presenting difficulties for its structural studies. Here a purification procedure that yields monomeric suPAR was reported. SuPAR obtained through such procedure is monomeric and quite stable. SuPAR complex with amino terminal fragment （ATF） of uPA and an anti-suPAR antibody （ATN615） was crystallized by dialysis method into diffracting crystals （1.9A）.

关 键 词：可溶性尿激酶受体 纯化 复合物结晶 

分 类 号：Q71[生物学—分子生物学]