衣藻叶绿体分裂相关基因CrFtsZ3的克隆及其原核表达  

Cloning and Expression in E. coli of Chlamydomonas reinhardtii CrFtsZ3 Gene

作　　者：胡勇[1] 孔冬冬[2] 王东[3] 何奕昆[1] 

机构地区：[1]首都师范大学生物系,北京100037 [2]中国科学院植物研究所,北京100093 [3]北京师范大学生命科学学院,北京100875

出　　处：《生物化学与生物物理学报》2003年第11期998-1004,共7页

基　　金：国家自然科学基金 (No .GN3 9970 3 5 6);北京市自然科学基金(No .GN5 992 0 0 3 )资助项目~~

摘　　要：FtsZ蛋白在原核细胞以及植物细胞叶绿体的分裂过程中发挥着重要作用。为了研究叶绿体分裂装置的进化 ,运用RT PCR方法从莱茵衣藻中克隆了叶绿体分裂相关基因CrFtsZ3。由于已经从衣藻细胞中克隆了一个ftsZ基因 ,所以CrFtsZ3的克隆表明衣藻中已经存在两类不同的 ftsZ基因 ,这说明 ftsZ基因的复制与分歧发生于绿藻的分化之前。序列分析结果显示 ,CrFtsZ3所编码的蛋白质具有FtsZ蛋白的典型模体。进一步的原核表达与定位分析表明CrFtsZ3 GFP融合蛋白沿着宿主菌体的纵轴方向有规律地聚集成荧光点或荧光带 ,并且CrFtsZ3蛋白过量表达明显干挠了宿主菌正常的细胞分裂过程 ,说明衣藻CrFtsZ3蛋白能够识别宿主细胞内的分裂位点并影响细胞分裂过程 。FtsZ protein plays a key role in the division of bacteria and chloroplast. To investigate the evolution of the chloroplast dividing apparatus, cloning and molecular characterization of a second chloroplast division gene, CrFtsZ3, from Chlamydomonas reinhardtii is performed. As there are two ftsZ genes in Chlamydomonas reinhardtii, duplication and divergence of the ftsZ genes might occur in an early stage before the emergence of green algae during the course of plant evolution. Sequence analysis showed that CrFtsZ3 gene had significant sequence homology with other ftsZs. It encoded a precursor of 479 amino acid residues with a putative transit peptide in its N terminal . To study the function of CrFtsZ3, a recombinant plasmid expressing the full length CrFtsZ3/EGFP fusion protein was constructed. By using IPTG inducing, overexpression of CrFtsZ3/EGFP in E. coli was achieved, and this overexpression blocked cell division and resulted in filament formation. GFP derived fluorescence showed regularly spaced dots along the bacterial filaments. This suggests that CrFtsZ3 could still recognize the signals of cell division site in E. coli and could take part in the process of bacterial division.

关 键 词：莱茵衣藻 叶绿体 分裂 基因 CrFtsZ3 克隆 RT-PCR 融合表达 系统发生树 

分 类 号：Q943.2[生物学—植物学]