Interaction of a small heat shock protein of the fission yeast, Schizosaccharomyces pombe, with a denatured protein at elevated temperature
Interaction of a small heat shock protein of the fission yeast, Schizosaccharomyces pombe, with a denatured protein at elevated temperature
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DOI：
10.1074/jbc.m504121200
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发表时间：
2005-09-23
影响因子：
4.8
通讯作者：
Yohda, M
中科院分区：
文献类型：
--
作者：
Hirose, M;Tohda, H;Yohda, M
我们表达，纯化，并表征了一个小的热休克蛋白裂殖酵母裂殖酵母，SpHSP 16.0。SpHsp 16.0能够高效地保护柠檬酸合酶免受45 ℃下的热聚集。在生理生长温度附近，它以十六聚体球状寡聚体的形式存在。在升高的温度下，低聚物解离成小的物质，可能是二聚体。解离是完全可逆的，并且在温度下降后立即重新形成原始低聚物。在45摄氏度下孵育然后冷却后，通过尺寸排阻层析和电子显微镜观察到SpHsp16.0和变性柠檬酸合酶的大复合物。然而，这样的大复合物不能从在45 ℃下孵育的尺寸排阻柱中分离出来。保护免于聚集的变性柠檬酸合酶在45 ℃下被GroEL陷阱突变体捕获。这些结果表明，复合物的SpHsp16.0和变性柠檬酸合酶在高温下是在瞬态，并具有疏水性。通过荧光互相关光谱法对SpHsp 16.0和变性柠檬酸合酶之间的相互作用的分析也表明，SpHsp 16.0-变性柠檬酸合酶复合物在升高的温度下的特征不同于在转移到降低的温度后获得的大复合物的特征。
We have expressed, purified, and characterized one small heat shock protein of the fission yeast Schizosaccharomyces pombe, SpHsp16.0. SpHsp16.0 was able to protect citrate synthase from thermal aggregation at 45 degrees C with high efficiency. It existed as a hexadecameric globular oligomer near the physiological growth temperature. At elevated temperatures, the oligomer dissociated into small species, probably dimers. The dissociation was completely reversible, and the original oligomer reformed immediately after the temperature dropped. Large complexes of SpHsp16.0 and denatured citrate synthase were observed by size exclusion chromatography and electron microscopy following incubation at 45 degrees C and then cooling. However, such large complexes did not elute from the size exclusion column incubated at 45 degrees C. The denatured citrate synthase protected from aggregation was trapped by a GroEL trap mutant at 45 degrees C. These results suggest that the complex of SpHsp16.0 and denatured citrate synthase at elevated temperatures is in the transient state and has a hydrophobic nature. Analyses of the interaction between SpHsp16.0 and denatured citrate synthase by fluorescence cross- correlation spectrometry have also shown that the characteristics of SpHsp16.0- denatured citrate synthase complex at the elevated temperature are different from those of the large complex obtained after the shift to lowered temperatures.