Module assembly for protein-surface recognition: Geranylgeranyltransferase I bivalent inhibitors for simultaneous targeting of interior and exterior protein surfaces
Module assembly for protein-surface recognition: Geranylgeranyltransferase I bivalent inhibitors for simultaneous targeting of interior and exterior protein surfaces
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DOI：
10.1002/chem.200701634
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发表时间：
2008-01-01
影响因子：
4.3
通讯作者：
Ohkanda, Junko
中科院分区：
文献类型：
--
作者：
Machida, Shinnosuke;Usuba, Kakeru;Ohkanda, Junko
设计用于同时靶向蛋白质表面多个位点的合成化学探针因其作为蛋白质-蛋白质相互作用的位点特异性调节剂的潜在应用而受到关注。报道了一种基于模块组装的哺乳动物 I 型香叶基香叶基转移酶 (GGTase 1) 二价抑制剂的新方法，用于同时识别内部和外部蛋白质表面。本研究中合成的抑制剂由两个通过烷基间隔基连接的模块组成；一个是四肽 CVIL 模块，用于与内部蛋白质表面（活性口袋）结合，另一个是 3,4,5-烷氧基取代的苯甲酰基基序，其中包含三个氨基烷基，设计用于与活性位点附近带负电荷的蛋白质外表面结合。通过基于荧光光谱和 [H-3] 标记的异戊二烯基团在蛋白质底物上的掺入的两种不同的酶抑制测定来筛选化合物。该二价抑制剂以亚微摩尔范围内的 K-i 值阻断 GGTase I 酶活性，其效果分别比四肽 CVIL 和苯甲酸甲酯衍生物高约一个数量级和 150 倍以上。二价化合物 6 和 8 被证明是竞争性抑制剂，表明 CVIL 模块将整个分子锚定到 GGTase I 活性位点，并将另一个模块递送到靶向蛋白质表面。因此，我们的模块组装方法导致同时多位点识别，从而协同抑制 GGTase I 活性，从而为设计用于靶向蛋白质-蛋白质相互作用的蛋白质表面定向抑制剂提供了一种新方法。
Synthetic chemical probes designed to simultaneously targeting multiple sites of protein surfaces are of interest owing to their potential application as site specific modulators of protein-protein interactions. A new approach toward bivalent inhibitors of mammalian type I geranylgeranyltransferase (GGTase 1) based on module assembly for simultaneous recognition of both interior and exterior protein surfaces is reported. The inhibitors synthesized in this study consist of two modules linked by an alkyl spacer; one is the tetrapeptide CVIL module for binding to the interior protein surface (active pocket) and the other is a 3,4,5-alkoxy substituted benzoyl motif that contains three aminoalkyl groups designed to bind to the negatively charged protein exterior surface near the active site. The compounds were screened by two distinct enzyme inhibition assays based on fluorescence spectroscopy and incorporation of a [H-3]-labeled prenyl group onto a protein substrate. The bivalent inhibitors block GGTase I enzymatic activity with K-i values in the submicromolar range and are approximately one order of magnitude and more than 150 times more effective than the tetrapeptide CVIL and the methyl benzoate derivatives, respectively. The bivalent compounds 6 and 8 were shown to be competitive inhibitors, suggesting that the CVIL module anchors the whole molecule to the GGTase I active site and delivers the other module to the targeting protein surface. Thus, our module-assembly approach resulted in simultaneous multiple-site recognition, and as a consequence, synergetic inhibition of GGTase I activity, thereby providing a new approach in designing protein-surface-directed inhibitors for targeting protein-protein interactions.