Light-dependent fragmentation of the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase in chloroplasts isolated from wheat leaves
Light-dependent fragmentation of the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase in chloroplasts isolated from wheat leaves
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DOI：
10.1007/s004250050260
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发表时间：
1998-03-01
期刊：
影响因子：
4.3
通讯作者：
Mae, T
中科院分区：
文献类型：
--
作者：
Ishida, H;Shimizu, S;Mae, T
在光照下，核酮糖-1，5-二磷酸羧化酶/加氧酶(Rubisco；EC 4.1.1.39)的大亚基(LSU)被叶绿体裂解产物中的羟基自由基降解为37 kDa的N端侧片段和16 kDa的C端侧片段(H.Ishida等)。1997，植物细胞生理学38：471-479)。在本研究中，我们证明了LSU的这种碎裂也以同样的方式发生在完整的叶绿体中，并讨论了这种碎裂的机制。将完整的小麦叶片叶绿体在KCN或NaN_3的光照下孵育，观察到LSU的断裂。在叶绿体中发现的片段的性质，如分子质量和与特定部位的抗LSU抗体的交叉反应，与裂解物中的相同。这些结果表明，与裂解产物一样，完整叶绿体中LSU的断裂也是由光产生的羟基自由基引起的。3-(3‘，4’-二氯苯基)-1，1-二甲基脲(DCMU)能完全抑制LSU的裂解，而甲基紫精只能部分抑制裂解产物中LSU的裂解。在裂解物中加入过氧化氢可以刺激LSU在光下的裂解，但在黑暗中不会引起任何裂解。因此，我们得出结论，在光下产生过氧化氢和在类囊体膜上产生还原力都是LSU碎裂的基本要求。
The large subunit (LSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco; EC 4.1.1.39) is degraded into an N-terminal side fragment of 37 kDa and a C-terminal side fragment of 16 kDa by the hydroxyl radical in the lysates of chloroplasts in light (H. Ishida et al. 1997, Plant Cell Physiol 38: 471-479). In the present study, we demonstrate that this fragmentation of the LSU also occurs in the same manner in intact chloroplasts, and discuss the mechanisms of the fragmentation. The fragmentation of the LSU was observed when intact chloroplasts from wheat leaves were incubated under illumination in the presence of KCN or NaN3, which is a potent inhibitor of active oxygen-scavenging enzyme(s). The properties, such as molecular masses and cross-reactivities against the site-specific anti-LSU antibodies, of the fragments found in the chloroplasts were the same as those found in the lysates. These results indicate that, as in the lysates, the fragmentation of the LSU in the intact chloroplasts was also caused by the hydroxyl radical generated in light. The fragmentation of the LSU was completely inhibited by 3-(3',4'-dichlorophenyl)-1,1-dimethylurea (DCMU), and only partially inhibited by methyl viologen in the lysates. The addition of hydrogen peroxide to the lysates stimulated LSU fragmentation in light, but did not induce any fragmentation in darkness. Thus, we conclude that both production of hydrogen peroxide and generation of the reducing power at thylakoid membranes in light are essential requirements for fragmentation of the LSU.