Synthesis and characterization of potent bivalent amyloidosis inhibitors that bind prior to transthyretin tetramerization
Synthesis and characterization of potent bivalent amyloidosis inhibitors that bind prior to transthyretin tetramerization
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DOI：
10.1021/ja030294z
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发表时间：
2003-11-05
影响因子：
15
通讯作者：
Kelly, JW
中科院分区：
文献类型：
--
作者：
Green, NS;Palaninathan, SK;Kelly, JW
甲状腺素运载蛋白（TTR）的错误折叠，包括限速性四聚体解离和部分单体变性，足以使TTR错误组装成淀粉样蛋白和其他与老年系统性淀粉样变性、家族性淀粉样多发性神经病和家族性淀粉样心肌病相关的异常四级结构。结合到TTR四级结构界面处的一个或两个未占据的甲状腺激素结合位点的单价小分子稳定天然状态，充分提高四聚体解离的动力学屏障，使得解离速率以及因此淀粉样变性变慢。本文报道了同时与两个结合位点结合的二价淀粉样蛋白抑制剂。候选二价抑制剂通常不能结合天然TTR四聚体，并且由于强烈的抑制剂取向偏好而通常不参与单价结合。然而，如果抑制剂在组装发生之前拦截蛋白质，则TTR四级结构可以围绕几个二价抑制剂组装。以这种方式制备的一些野生型TTR-二价抑制剂复合物在经受实质性变性应激（8 M尿素，120 h）时保留四聚体结构。最好的二价抑制剂将酸介导的TTR（3.6 μ M）淀粉样蛋白原纤维形成减少到不存在抑制剂时TTR所表现的6%，与最好的单价抑制剂（3.6 μ M，72小时）所观察到的类似30%相比有显著改善。最好的二价抑制剂的表观解离速率实际上为零，这与TTR四聚体解离和抑制剂解离是连接的这一想法一致--这是作为模板的四聚体组装的结果。两个复合物的X-射线共晶结构表明，二价抑制剂同时占据TTR中的两个位点，与来自HPLC分析的1：1结合化学计量一致。本研究的目的是证明二价抑制剂可能是有用的;结果是迄今为止生产的最好的抑制剂。在这种情况下，能够在内质网腔内折叠和组装期间拦截TTR的分子将是明显的兴趣。
The misfolding of transthyretin (TTR), including rate-limiting tetramer dissociation and partial monomer denaturation, is sufficient for TTR misassembly into amyloid and other abnormal quaternary structures associated with senile systemic amyloidosis, familial amyloid polyneuropathy, and familial amyloid cardiomyopathy. Monovalent small molecules that bind to one or both of the unoccupied thyroid hormone binding sites at the TTR quaternary structure interface stabilize the native state, raising the kinetic barrier for tetramer dissociation sufficiently that the rate of dissociation, and therefore amyloidosis, becomes slow. Bivalent amyloid inhibitors that bind to both binding sites simultaneously are reported herein. The candidate bivalent inhibitors are generally unable to bind to the native TTR tetramer and typically do not engage in monovalent binding owing to a strong inhibitor orientation preference. However, the TTR quaternary structure can assemble around several of the bivalent inhibitors if the inhibitor intercepts the protein before assembly occurs. Some of the wild-type TTR-bivalent inhibitor complexes prepared in this fashion retain a tetrameric structure when subjected to substantial denaturation stresses (8 M urea, 120 h). The best bivalent inhibitor reduced acid-mediated TTR (3.6 muM) amyloid fibril formation to 6% of that exhibited by TTR in the absence of inhibitor, a significant improvement over the similar to30% observed for the best monovalent inhibitors (3.6 muM, 72 h). The apparent dissociation rate of the best bivalent inhibitor is effectively zero, consistent with the idea that TTR tetramer dissociation and inhibitor dissociation are linked-as a result of the inhibitor-templating tetramer assembly. X-ray cocrystal structures of two of the complexes demonstrate that the bivalent inhibitors simultaneously occupy both sites in TTR, consistent with the 1:1 binding stoichiometry derived from HPLC analysis. The purpose of this study was to demonstrate that bivalent inhibitors could be useful; what resulted are the best inhibitors produced to date. In this context, molecules capable of intercepting TTR during folding and assembly in the lumen of the endoplasmic reticulum would be of obvious interest.