Rapid Optimization of Mcl-1 Inhibitors using Stapled Peptide Libraries Including Non-Natural Side Chains.
Rapid Optimization of Mcl-1 Inhibitors using Stapled Peptide Libraries Including Non-Natural Side Chains.
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DOI：
10.1021/acschembio.5b01002
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发表时间：
2016-05-20
影响因子：
4
通讯作者：
Keating AE
中科院分区：
文献类型：
--
作者：
Rezaei Araghi R;Ryan JA;Letai A;Keating AE
-螺旋是许多蛋白-蛋白相互作用结合界面的关键部分，化学稳定的合成螺旋肽可以有效抑制这种螺旋介导的复合物。特别是，烃类钉接肽以产生受限的螺旋可以改善肽的结合亲和力和其他特性，但确定最佳的钉接肽变体往往需要艰苦的试验和错误。在这里，我们描述了一种与Mcl-1结合的钉状螺旋肽的快速发现和优化，Mcl-1是一种在许多化疗耐药癌症中过表达的抗凋亡蛋白。为了加速发现，我们开发了一种肽库合成和筛选方案，能够识别mcl -1结合的钉接肽之间的细微亲和力差异。我们使用我们的方法对Mcl-1抑制剂中引入的非天然氨基酸取代的组合进行了采样，这些氨基酸取代是在固定的螺旋稳定碳氢化合物主要成分的背景下增加肽螺旋含量并减少蛋白质水解的。在我们的筛选中发现的肽在天然结合伙伴中保守的位点上含有令人惊讶的取代。文库鉴定的肽M3d是迄今为止在Mcl-1依赖性细胞系中选择性触发线粒体通透性的最有效分子。我们优化螺旋肽抑制剂的文库方法可以很容易地应用于其他生物医学重要靶点的研究。
Alpha helices form a critical part of the binding interface for many protein-protein interactions, and chemically stabilized synthetic helical peptides can be effective inhibitors of such helix-mediated complexes. In particular, hydrocarbon stapling of peptides to generate constrained helices can improve binding affinity and other peptide properties, but determining the best stapled peptide variant often requires laborious trial and error. Here we describe the rapid discovery and optimization of a stapled-helix peptide that binds to Mcl-1, an anti-apoptotic protein that is overexpressed in many chemoresistant cancers. To accelerate discovery, we developed a peptide library synthesis and screening scheme capable of identifying subtle affinity differences among Mcl-1-binding stapled peptides. We used our method to sample combinations of non-natural amino-acid substitutions that we introduced into Mcl-1 inhibitors in the context of a fixed helix-stabilizing hydrocarbon staple that increased peptide helical content and reduced proteolysis. Peptides discovered in our screen contained surprising substitutions at sites that are conserved in natural binding partners. Library-identified peptide M3d is the most potent molecule yet tested for selectively triggering mitochondrial permeabilization in Mcl-1 dependent cell lines. Our library approach for optimizing helical peptide inhibitors can be readily applied to the study of other biomedically important targets.