Photoaffinity analogues of farnesyl pyrophosphate transferable by protein farnesyl transferase
Photoaffinity analogues of farnesyl pyrophosphate transferable by protein farnesyl transferase
复制标题
DOI：
10.1021/ja0124717
复制
发表时间：
2002-07-17
影响因子：
15
通讯作者：
Spielmann, HP
中科院分区：
文献类型：
--
作者：
Chehade, KAH;Kiegiel, K;Spielmann, HP
法尼化是一种翻译后的脂质修饰，在蛋白质法尼基转移酶(FTase)的催化下，15碳的法尼基异戊二烯通过硫醚键连接到蛋白质的特定半胱氨酸残基上。我们合成了法尼基焦磷酸(FPP)的类似物(3-6)，以探索FPP骨架可能的修饰范围，从而允许FTase有效地转移。用全氟苯叠氮官能团取代FPP的欧米茄末端异戊二烯，制备了FPP(5，6)的光亲和类似物。取代的苯胺取代了类似物3和4中的欧米伽末端的异戊二烯。化合物3-5是通过适当的苯胺与8-氧锗乙酸酯还原胺化，然后经酯水解、氯化和焦磷酸化得到的。再用三个亚甲基取代FPP的β-异戊二烯，得到9个步骤的光电探针6。类似物的制备需要在NaBH(OAc)(3)还原之前由TiCl4介导亚胺的形成。叠氮部分不受Ph3PCl2将烯丙醇13-16转化为相应的氯化物17-20的影响。类似物3-6通过哺乳动物FTase有效地转移到靶N-Dansyl-GCVLS多肽底物上。与FPP的类似物结构和转移动力学的比较表明，环氟化和对位取代基对类似物焦磷酸对FT酶的亲和力和转移效率几乎没有影响。FTase活性部位类似物结合模式的模型也支持这些结果。可转移叠氮光探针5可使FTase失活。可转移的类似物5和6允许形成适当的翻译后修饰的异戊二烯功能的光反应多肽探针。
Farnesylation is a posttranslational lipid modification in which a 15-carbon farnesyl isoprenoid is linked via a thioether bond to specific cysteine residues of proteins in a reaction catalyzed by protein farnesyltransferase (FTase). We synthesized analogues (3-6) of farnesyl pyrophosphate (FPP) to probe the range of modifications possible to the FPP skeleton which allow for efficient transfer by FTase. Photoaffinity analogues of FPP (5, 6) were prepared by substituting perfluorophenyl azide functional groups for the omega-terminal isoprene of FPP. Substituted anilines replace the omega-terminal isoprene in analogues 3 and 4. Compounds 3-5 were prepared by reductive amination of the appropriate anilines with 8-oxogeranyl acetate, followed by ester hydrolysis, chlorination, and pyrophosphorylation. Additional substitution of three methylenes for the beta-isoprene of FPP gave photoprobe 6 in nine steps. Preparation of the analogues required TiCl4-mediated imine formation prior to NaBH(OAc)(3) reduction for anilines with a pK(a) < 1. The azide moiety was not affected by Ph3PCl2 conversion of allylic alcohols 13-16 into corresponding chlorides 17-20. Analogues 3-6 are efficiently transferred to target N-dansyl-GCVLS peptide substrate by mammalian FTase. Comparison of analogue structures and kinetics of transfer to those of FPP reveals that ring fluorination and para substituents have little effect on the affinity of the analogue pyrophosphate for FTase and its transfer efficiency. These results are also supported with models of the analogue binding modes in the active site of FTase. The transferable azide photoprobe 5 photoinactivates FTase. Transferable analogues 5 and 6 allow the formation of appropriately posttranslationally modified photoreactive peptide probes of isoprene function.