Mammalian protein RAP46: an interaction partner and modulator of 70 kDa heat shock proteins
Mammalian protein RAP46: an interaction partner and modulator of 70 kDa heat shock proteins
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DOI：
10.1093/emboj/16.18.5483
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发表时间：
1997-09
期刊：
影响因子：
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通讯作者：
M. Zeiner;Mathias Gebauer;U. Gehring
中科院分区：
文献类型：
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作者：
M. Zeiner;Mathias Gebauer;U. Gehring
最近发现了一种广泛表达的核受体相关蛋白（RAP 46）。与体外翻译的蛋白质和细胞提取物中含有的蛋白质的相互作用实验表明，多种细胞调节剂与RAP 46相关。然而，在通过远Western技术的直接相互作用测试中，仅70 kDa蛋白质出现并被鉴定为70 kDa热休克蛋白（hsp 70）家族的成员。相互作用是特异性的，因为不是所有的hsp 70家族成员都与RAP 46结合;相互作用通过它们的ATP结合结构域发生。RAP 46在哺乳动物细胞中与hsp 70形成复合物，并在酵母双杂交系统中与hsp 70相互作用。与hsp 70可以结合多种蛋白质的事实一致，我们鉴定了RAP 46-hsp 70与一些选定蛋白质的异聚复合物，最值得注意的是c-Jun。通过用碱性磷酸酶预处理，复合物形成显著增加，从而表明通过蛋白磷酸化调节相互作用。我们观察到RAP 46干扰热变性荧光素酶的有效重折叠。此外，错误折叠蛋白与hsp 70的ATP依赖性结合被RAP 46极大地抑制。这些数据表明RAP 46在高等真核生物中作为hsp 70的调节剂起作用。
A ubiquitously expressed nuclear receptor‐associating protein of ∼46 kDa (RAP46) was identified recently. Interaction experiments with in vitro‐translated proteins and proteins contained in cell extracts revealed that a great variety of cellular regulators associate with RAP46. However, in direct interaction tests by the far‐Western technique, only 70 kDa proteins showed up and were identified as members of the 70 kDa heat shock protein (hsp70) family. Interaction is specific since not all members of the hsp70 family bind to RAP46; interaction occurs through their ATP‐binding domain. RAP46 forms complexes with hsp70 in mammalian cells and interacts with hsp70 in the yeast two‐hybrid system. Consistent with the fact that hsp70 can bind a multitude of proteins, we identified heteromeric complexes of RAP46–hsp70 with some selected proteins, most notably c‐Jun. Complex formation is increased significantly by pre‐treatment with alkaline phosphatase, thus suggesting modulation of interactions by protein phosphorylation. We observed that RAP46 interferes with efficient refolding of thermally denatured luciferase. Moreover, ATP‐dependent binding of misfolded proteins to hsp70 was greatly inhibited by RAP46. These data suggest that RAP46 functions as a regulator of hsp70 in higher eukaryotes.