A 21-kDa chloroplast heat shock protein assembles into high molecular weight complexes in vivo and in Organelle.
A 21-kDa chloroplast heat shock protein assembles into high molecular weight complexes in vivo and in Organelle.
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DOI：
10.1016/s0021-9258(17)36821-7
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发表时间：
1994-05
期刊：
影响因子：
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通讯作者：
Q. Chen;K. Osteryoung;E. Vierling
中科院分区：
文献类型：
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作者：
Q. Chen;K. Osteryoung;E. Vierling
小(Sm)细胞质和叶绿体热休克蛋白(HSPs)之间的羧基末端“热休克”结构域的保守性表明，这些smHSPs具有相似的功能。以往的研究已经证实，细胞质中的smHSPs存在于体内的高阶结构中(约500 kDa)。为了确定是否在类似的复合体中发现了叶绿体smHSP，我们研究了豌豆PsHSP21在非变性条件下21 kDa的叶绿体smHSP的大小。叶绿体基质提取物在蔗糖梯度上沉淀后，在10-11S对应的组分中检测到PsHSP21。在非变性凝胶电泳上，在约230 kDa和200 kDa的两个高分子量复合体中检测到PsHSP21，与蔗糖梯度数据一致。这些含有PsHSP21的颗粒在不同的盐和镁离子条件下都是稳定的，它们的完整性不受1.0%Triton X-100或10 mM ATP的影响。为了研究含有PsHSP21的高分子量复合体的组装，将体外翻译的PsHSP21导入叶绿体并测定其大小。在从热胁迫植物分离的叶绿体中导入PsHSP21后，超过50%的PsHSP21以更高分子量的形式被回收。相反，在从对照植物分离的叶绿体中输入该蛋白后，该蛋白仅以5 S(约42 kDa)的形式被回收。这些数据表明，可能需要预先存在的PsHSP21或其他热诱导因子来组装更高分子量的颗粒。我们认为10~(-11)S粒子是PsHSP21的功能形式。
The conservation of the carboxyl-terminal “heat shock” domain among small (sm) cytoplasmic and chloroplast heat shock proteins (HSPs) suggests that these smHSPs perform similar functions. Previous studies have established that cytoplasmic smHSPs are found in higher order structures in vivo (approximately 500 kDa). To determine if the chloroplast smHSP is found in similar complexes, we examined the size of the 21-kDa chloroplast smHSP from Pisum sativum, PsHSP21, under non-denaturing conditions. Following sedimentation of chloroplast stromal extracts on sucrose gradients PsHSP21 is detected in fractions corresponding to 10-11 S. Upon non-denaturing gel electrophoresis, PsHSP21 was detected in two high molecular mass complexes of approximately 230 and 200 kDa, in good agreement with the sucrose gradient data. These PsHSP21-containing particles were stable under different salt and Mg2+ conditions, and their integrity was not affected by 1.0% Triton X-100 or 10 mM ATP. To study assembly of the high molecular weight complexes containing PsHSP21, in vitro translated PsHSP21 was imported into chloroplasts and its size was examined. Following import into chloroplasts isolated from heat-stressed plants, greater than 50% of PsHSP21 was recovered in the higher molecular weight forms. In contrast, following import into chloroplasts isolated from control plants the protein was recovered exclusively in a 5 S (approximately 42-kDa) form. These data suggest that preexisting PsHSP21 or other heat-induced factors may be required for assembly of the higher molecular weight particles. We propose that the 10-11 S particles are the functional form of PsHSP21.