高等植物Rubisco的组装及其中间产物的鉴定  

Rubisco Assembly in Higher Plant and Characterization of Its Intermediates

作　　者：陈根云[1] 肖元珍[1] 李立人[1] 

机构地区：[1]中国科学院上海植物生理研究所,上海200032

出　　处：《植物生理学报（0257-4829)》2000年第1期16-22,共7页Acta Phytophysiologica Sinica

基　　金：国家重点基础研究发展规划及国家自然科学基金

摘　　要：将新鲜制备的不含Ｒｕｂｉｓｃｏ的水稻叶片低分子量蛋白组分在离体条件下于室温保温４８ｈ，ＮＤＰＡＧＥ分析发现在ＡＴＰ５ｍｍｏｌ／Ｌ和Ｍｇ２＋５ｍｍｏｌ／Ｌ的作用条件下，在分子量为５６０ｋＤ位置上有一蛋白带生成。高浓度的Ｋ＋在一定程度上抑制它的形成，在保温介质中没有ＡＴＰ存在时，不能产生５６０ｋＤ分子量的条带，但有一更高分子量（约６００ｋＤ）蛋白条带的产生，这一条带能在ＡＴＰ和Ｍｇ２＋作用下发生解离。Ｗｅｓｔｅｒｎｂｌｏｔ和ＳＤＳＰＡＧＥ分析证实在ＡＴＰ和Ｍｇ２＋存在的情况下，组装成的蛋白确实是Ｒｕｂｉｓｃｏ全酶；而６００ｋＤ的蛋白是由Ｒｕｂｉｓｃｏ大小亚基和ｃｐｎ６０的α、β亚基组成的一个复合物，用３５ＳＭｅｔ同位素标记的水稻Ｒｕｂｉｓｃｏ小亚基前体通过跨膜运输与豌豆叶绿体中的大亚基进行异源重组后，发现除了组装成的异源Ｒｕｂｉｓｃｏ杂合酶外，还发现存在有多种组装中间体。根据这些中间体分子量的测定，推测了高等植物Ｒｕｂｉｓｃｏ在ｃｐｎ６０和ｃｐｎ１０作用下完成组装的可能途径。The Rubisco deleted low molecular weight (LMW) protein components was prepared by Superose 6 column with FPLC from the crude extracts of rice leaves, then the LMW protein components were incubated with ATP, Mg 2+ or K + in a different combination at room temperature for 48 h. The results of ND PAGE analysis indicated that ATP and Mg 2+ were necessary for Rubisco assembly, and K + inhibited the assembling process to some extent. When no ATP was added to the incubation medium, a high molecular weight (HMW) protein complex (～600 kD) was observed (Fig.2). The results of Western blot and SDS PAGE indicated that the HMW protein complex consisted of two kinds of ch cpn60 (α&β) subunits,the large subunit(LS) and small subunit (SS) of Rubisco, and some unknown proteins subunits(Fig.4). When the rice prSS labeled with 35 S Met was incubated under light with intact chloroplasts from pea, and the supernatant of broken chloroplasts was analyzed by ND PAGE and autoradiography, several radioactive protein bands appeared in the photo besides the assembled hybrid Rubisco. Their molecular weights varied from 70 to 600 kD (Fig.5). Some of their intermediates have been confirmed to contain LS and SS by Western blot analysis against anti rice LS serum. According to the molecular weights of the intermediates found, we proposed a pathway of Rubisco assembly in higher plant (Fig.6).

关 键 词：RUBISCO 组装 中间体 cpn60 cpn10 光合作用 

分 类 号：Q946[生物学—植物学]