STRUCTURE-ACTIVITY STUDIES OF RAPAMYCIN ANALOGS - EVIDENCE THAT THE C-7 METHOXY GROUP IS PART OF THE EFFECTOR DOMAIN AND POSITIONED AT THE FKBP12-FRAP INTERFACE
STRUCTURE-ACTIVITY STUDIES OF RAPAMYCIN ANALOGS - EVIDENCE THAT THE C-7 METHOXY GROUP IS PART OF THE EFFECTOR DOMAIN AND POSITIONED AT THE FKBP12-FRAP INTERFACE
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DOI：
10.1016/1074-5521(95)90264-3
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发表时间：
1995-07-01
影响因子：
--
通讯作者：
HOLT, DA
中科院分区：
文献类型：
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作者：
LUENGO, JI;YAMASHITA, DS;HOLT, DA
背景：雷帕霉素是一种免疫抑制性天然产物，可阻断T细胞有丝分裂和酵母增殖。在细胞质中，雷帕霉素与亲免素FKBP 12结合，这两种分子的复合物与最近发现的蛋白质FRAP结合。雷帕霉素分子具有两个功能结构域，通过它们与FKBP 12（结合结构域）或与FRAP（效应结构域）的相互作用来定义。我们以前表明，烯丙基甲氧基在C-7的雷帕霉素可以被各种不同的取代基取代。我们着手研究这种取代对FKBP 12结合和生物活性的影响。结果：雷帕霉素C-7-修饰的R和S构型的类似物被证明对FKBP 12具有高亲和力，但这些同源物在脾细胞和酵母增殖试验中显示出广泛的效力。测定了与FKBP 12复合的四种雷帕霉素类似物的X射线晶体结构，并揭示了蛋白质和配体骨架构象与母体雷帕霉素-FKBP 12复合物中观察到的构象基本相同，并且C-7基团仍暴露于溶剂。然后，我们制备了具有光反应性官能团作为C-7取代基的一部分的雷帕霉素类似物。该化合物特异性标记，在FKBP 12依赖的方式，一个类似于250 kDa的蛋白质，这comigrates与重组FRAP.Conclusions：我们得出结论，C-7甲氧基雷帕霉素的效应域的一部分。在三元复合物中，该组位于FRAP和FKBP 12之间的界面处，紧邻FRAP。
Background: Rapamycin is an immunosuppressant natural product, which blocks T-cell mitogenesis and yeast proliferation. In the cytoplasm, rapamycin binds to the immunophilin FKBP12 and the complex of these two molecules binds to a recently discovered protein, FRAP. The rapamycin molecule has two functional domains, defined by their interaction with FKBP12 (binding domain) or with FRAP (effector domain). We previously showed that the allylic methoxy group at C-7 of rapamycin could be replaced by a variety of different substituents. We set out to examine the effects of such substitutions on FKBP12 binding and on biological activity.Results: Rapamycin C-7-modified analogs of both R and S configurations were shown to have high affinities for FKBP12, yet these congeners displayed a wide range of potencies in splenocyte and yeast proliferation assays. The X-ray crystal structures of four rapamycin analogs in complexes with FKBP12 were determined and revealed that protein and ligand backbone conformations were essentially the same as those observed for the parent rapamycin-FKBP12 complex and that the C-7 group remained exposed to solvent. We then prepared a rapamycin analog with a photoreactive functionality as part of the C-7 substituent. This compound specifically labeled, in an FKBP12-dependent manner, a protein of similar to 250 kDa, which comigrates with recombinant FRAP.Conclusions: We conclude that the C-7 methoxy group of rapamycin is part of the effector domain. In the ternary complex, this group is situated in close proximity to FRAP, at the interface between FRAP and FKBP12.