Protomer Roles in Chloroplast Chaperonin Assembly and Function  

作　　者：Cuicui Bai Peng Guo Qian Zhao Zongyang Lv Shijia Zhang Fei Gao Liyan Gao Yingchun Wang Zhixi Tian Jifeng Wang Fuquan Yang Cuimin Liu 

机构地区：[1]State Key Laboratory of Plant Cell and Chromosome Engineering, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences,Beijing 100101, China [2]Key Laboratory of Molecular Development Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China [3]University of Chinese Academy of Sciences, Beijing 100101, China [4]Laboratory of Protein and Peptide Pharmaceuticals and Laboratory of Proteomics, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China [5]These authors contributed equally to this article.

出　　处：《Molecular Plant》2015年第10期1478-1492,共15页分子植物（英文版）

摘　　要：The individual roles of three chloroplast CPN60 protomers （CPN60α, CPN60β1, and CPN60β2） and whether and how they are assembled into functional chaperonin complexes are investigated in Chlamydomonas reinhardtii. Protein complexes containing all three potential subunits were identified in Chlamydomonas, and their co-expression in Escherichia coil yielded a homogeneous population of oligomers containing all three subunits （CPN60α1β11β2）, with a molecular weight consistent with a tetradecameric structure. While homo-oligomers of CPN60β could form, they were dramatically reduced when CPN60α was present and homo-oligomers of CPN60β2 were readily changed into hetero-oligomers in the presence of ATP and other protomers. ATP hydrolysis caused CPN60 oligomers to disassemble and drove the purified protomers to reconstitute oligomers in vitro, suggesting that the dynamic nature of CPN60 oligomers is dependent on ATP. Only hetero-oligomeric CPN60α1β1β2, containing CPN60α, CPN60β1, and CPN60β2 subunits in a 5：6：3 ratio, cooperated functionally with GroES. The combination of CPN60α and CPN60β subunits, but not the individual subunits alone, complemented GroEL function in E. coil with subunit recognition specificity. Down-regulation of the CPN60α subunit in Chlamydomonas resulted in a slow growth defect and an inability to grow autotrophically, indicating the essential role of CPN60α in vivo.The individual roles of three chloroplast CPN60 protomers （CPN60α, CPN60β1, and CPN60β2） and whether and how they are assembled into functional chaperonin complexes are investigated in Chlamydomonas reinhardtii. Protein complexes containing all three potential subunits were identified in Chlamydomonas, and their co-expression in Escherichia coil yielded a homogeneous population of oligomers containing all three subunits （CPN60α1β11β2）, with a molecular weight consistent with a tetradecameric structure. While homo-oligomers of CPN60β could form, they were dramatically reduced when CPN60α was present and homo-oligomers of CPN60β2 were readily changed into hetero-oligomers in the presence of ATP and other protomers. ATP hydrolysis caused CPN60 oligomers to disassemble and drove the purified protomers to reconstitute oligomers in vitro, suggesting that the dynamic nature of CPN60 oligomers is dependent on ATP. Only hetero-oligomeric CPN60α1β1β2, containing CPN60α, CPN60β1, and CPN60β2 subunits in a 5：6：3 ratio, cooperated functionally with GroES. The combination of CPN60α and CPN60β subunits, but not the individual subunits alone, complemented GroEL function in E. coil with subunit recognition specificity. Down-regulation of the CPN60α subunit in Chlamydomonas resulted in a slow growth defect and an inability to grow autotrophically, indicating the essential role of CPN60α in vivo.

关 键 词：CHAPERONIN Cpn60 PHOTOSYNTHESIS protomer ASSEMBLY 

分 类 号：Q51[生物学—生物化学] S511.01[农业科学—作物学]