人肽基脯氨酰顺反异构酶基因的克隆、表达、纯化及热变性交联  

Cloning, expression, purification and protein cross-linking during thermal unfolding of human peptidylprolyl-cis- trans-isomerase cyclophilin

作　　者：周霞[1] 刘凤华[1] 席慧[1] 万平[1] 何宏伟[1] 高音[1] 

机构地区：[1]首都师范大学生物系,北京100037

出　　处：《首都师范大学学报（自然科学版）》2004年第2期52-56,65,共6页Journal of Capital Normal University：Natural Science Edition

基　　金：国家自然科学基金项目 (No .3 980 0 0 2 8) ;北京市科技新星计划项目 (No .9612 8)

摘　　要：蛋白质分子间交联是普遍存在的现象 .然而 ,蛋白质交联的分子机理还不太清楚 .为了进一步探测蛋白质交联的分子机理 ,以及交联能否在异源肽链间发生 ,本实验室克隆了人肽基脯氨酰顺反异构酶 (humanPeptidylproly cis trans isomerase ,hPPI)cyclophilincDNA基因 ,并纯化出了PPI蛋白 .最后 ,将PPI和lysozyme蛋白进行热变性交联实验 ,结果显示在同源和异源肽链间都有二聚体和多聚体形成 .并证实蛋白质交联可经三步完成 :1 )蛋白质构象包括二级结构改变 ;2 )形成分子间二硫键 ;3)Protein interchain cross\|linking is a common phenomenon.However the molecular mechanisms for protein cross\|linking are not well understood.For further exploring the molecular mechanisms for protein cross\|linking, and detect if cross\|linking can happen in heterogeneous peptides, the investigation was extended with human peptidylprolyl\|cis\|trans\|isomerase cyclophilin (hPPI cyclophilin). hPPI cyclophilin cDNA was cloned and PPI protein was priufied.At last,hPPI cyclophilin and lysozyme were mixed during thermal unfolding.The observation revealed that heterodimer/oligomer formation was as common as homodimer/oligomer formation.It also proved protein cross\|linking can be accomplished in three concerted steps:(1) a change in protein conformation;(2) formation of interchain disulfide bonds; and (3) formation of interchain isopeptide cross\|links.

关 键 词：肽基脯氨酰顺反异构酶 克隆 基因表达 纯化 CYCLOPHILIN 固定化金属亲和层析 构象病 热变性交联 

分 类 号：Q558.2[生物学—生物化学] Q78