Morra Giulia, Neves Marco A C, Plescia Christopher J, Tsustsumi Shinji, Neckers Len, Verkhivker Gennady, Altieri Dario C, Colombo Giorgio
Istituto di Chimica del Riconoscimento Molecolare, CNR, Via Mario Bianco 9, 20131 Milano, Italy, Department of Cancer Biology, University of Massachusetts Medical School, Worcester, Massachusetts 01605, Urologic Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland 20892, and Department of Pharmaceutical Chemistry, School of Pharmacy and Center for Bioinformatics, University of Kansas, Lawrence, Kansas 66047.
J Chem Theory Comput. 2010 Sep 14;6(9):2978-89. doi: 10.1021/ct100334n. Epub 2010 Aug 30.
The study of allosteric functional modulation in dynamic proteins is attracting increasing attention. In particular, the discovery of new allosteric sites may generate novel opportunities and strategies for drug development, overcoming the limits of classical active-site oriented drug design. In this paper, we report on the results of a novel, ab initio, fully computational approach for the discovery of allosteric inhibitors based on the physical characterization of signal propagation mechanisms in proteins and apply it to the important molecular chaperone Hsp90. We first characterize the allosteric "hot spots" involved in interdomain communication pathways from the nucleotide-binding site in the N-domain to the distal C-domain. On this basis, we develop dynamic pharmacophore models to screen drug libraries in the search for small molecules with the functional and conformational properties necessary to bind these "hot spot" allosteric sites. Experimental tests show that the selected moelcules bind the Hsp90 C-domain, exhibit antiproliferative activity in different tumor cell lines, while not affecting proliferation of normal human cells, destabilize Hsp90 client proteins, and disrupt association with several cochaperones known to bind the N- and M-domains of Hsp90. These results prove that the hits alter Hsp90 function by affecting its conformational dynamics and recognition properties through an allosteric mechanism. These findings provide us with new insights on the discovery and development of new allosteric inhibitors that are active on important cellular pathways through computational biology. Though based on the specific case of Hsp90, our approach is general and can readily be extended to other target proteins and pathways.
对动态蛋白质中变构功能调节的研究正吸引着越来越多的关注。特别是，新变构位点的发现可能为药物开发带来新的机遇和策略，克服传统活性位点导向药物设计的局限性。在本文中，我们报告了一种基于蛋白质中信号传播机制的物理特征来发现变构抑制剂的全新的、从头开始的完全计算方法的结果，并将其应用于重要的分子伴侣Hsp90。我们首先表征了从N结构域中的核苷酸结合位点到远端C结构域的结构域间通讯途径中涉及的变构“热点”。在此基础上，我们开发了动态药效团模型来筛选药物库，以寻找具有结合这些“热点”变构位点所需功能和构象特性的小分子。实验测试表明，所选分子与Hsp90 C结构域结合，在不同肿瘤细胞系中表现出抗增殖活性，同时不影响正常人细胞的增殖，使Hsp90客户蛋白不稳定，并破坏与已知结合Hsp90 N和M结构域的几种共伴侣的结合。这些结果证明，命中的分子通过变构机制影响Hsp90的构象动力学和识别特性，从而改变Hsp90的功能。这些发现为通过计算生物学发现和开发对重要细胞途径有活性的新型变构抑制剂提供了新的见解。尽管基于Hsp90的具体案例，但我们的方法具有通用性，可轻松扩展到其他靶蛋白和途径。
