分子伴侣蛋白Hsp60参与折叠组装新合成或胁迫变性的蛋白质。叶绿体Cpn60复合体折叠光合作用限速酶Rubisco大亚基，是Rubisco全酶生物合成的基础。Cpn60是由多个不同亚基组成的双环结构，并根据序列特异性分为α或β型。我们前期工作已经证实莱茵衣藻叶绿体内三种CPN60亚基都参与复合体的形成，且每个亚基在形成复合体中功能不同。尽管两个CPN60β亚基高度同源，但在响应ATP水解作用时表现完全不同。本项目拟利用遗传学、生物化学以及结构生物学手段，探讨CPN60复合体内三个亚基的协作与功能分化，并解析其分子机理。Chaperonin亚基含有三个结构域，体现在氨基酸序列上为五个片段。我们拟将三个CPN60亚基与GroEL亚基的五个片段重新组合，构建各种嵌合体，体内遗传互补GroEL的功能，体外生化研究其功能的分子基础。并利用结构生物学手段，解析各亚基功能分化的元件与分子机理。

Chaperones are essential for maintaining cellular protein homeostasis and exist in all kingdoms of life. Chaperonins are a subfamily of chaperones, folding and refolding of a wide variety of newly synthesised or stress-denatured polypeptides, which have diverged into two distinct groups. Group I chaperonins, found in prokaryotes or in organelles of prokaryotic origin, include GroEL/ES in E.coli, Hsp60/10 in mitochondria, and Cpn60/20 in chloroplasts.Cpn60 was discovered as Rubisco large subunit binding protein which is distinguished from other group I chaperonin by its complex composition. In contrast to homologues GroEL or Hsp60, Cpn60 consists of multiple subunits divided into two distinct α and β types. Our previous studies investigated that CPN60 complexes contain all three subunits in Chlamydomonas, and each protomer has specific function. Though CPN60β1 and CPN60β2 share very high sequence similarity (more than 90%), CPN60β oligomers were differentially affected by ATP, with ATP causing no observable effect on CPN60β1 oligomers and complete dissassembly of CPN60β2 oligomers. The project aims to investigate the cooperation and functional partition of chloroplast chaperonin, and study the mechanism of their different behaviors. The chaperonin subunit contains three structural domains: an apical domain, an intermediate domain and an equatorial domain, which were divided into five fragments in the primary sequences. We plan to swap the five fragments from three CPN60 subunits and GroEL subunit, and construct over 30 CPN60/GroEL chimeras. The structure and function of chimeras will be investigated in vivo and in vitro to determine the elements influencing the cooperation and functional partition of subunits. The underlying mechanism will be investigated by crystal structure analysis.