标题: | 区域-区域交互作用地图探索蛋白质交互作用介面 Site-site interaction maps for exploring protein-protein interfaces |
作者: | 杨方琪 杨进木 Yang, Fang-Chi Yang, Jinn-Moon 生物资讯及系统生物研究所 |
关键字: | 区域-区域交互作用地图;蛋白质交互作用;核心锚点;专一性锚点;site-site interaction maps;protein-protein interaction;PPI family;core anchor;specific anchor |
公开日期: | 2016 |
摘要: | 蛋白质交互作用(PPIs)的机制对许多生物途径以及设计药物上都扮演着重要的角色。目前在PPIs的分析上已经有研究利用单一残基或一对交互作用残基的保守性来推论蛋白质交互作用介面上的热点(hotspots)。然而这些表面包含着许多交互作用的残基,因此在解释交互作用介面上的功能性以及在结构上扮演的角色时,藉由观察一群具有密集交互作用以及在多个物种上具有保守性的残基,会比只分析单一交互作用的残基还要符合生物上的意义。而在实验室之前的研究所提出的SiMMap概念,是利用大量的小分子化合物和蛋白质的交互作用,来观察蛋白质对于官能基的偏好以及在结合介面上的物化特性。因此我们将应用此概念以及跨多物种的同源蛋白交互作用家族(PPI family)来分析蛋白质交互作用介面。 在本论文中提出了一个新的概念称为区域-区域交互作用地图(site-site interaction maps),用来分析蛋白质介面上的数个微环境,此外,为了分析介面上的微环境,透过此概念可以将交互作用介面区分为数个具有保守性且较密集交互作用之区域,我们称为锚点(anchor),而这些锚点内的残基多为具有高度保守性且具有生物功能。除此之外,这些锚点可被分为两种型态,分别为:核心锚点(core anchor)表示该锚点的物化特性在PPI families间具有保守性,以及专一性锚点(specific anchor)表示该锚点在特定PPI family内会形成特定的结合环境。而为了验证我们所定义的锚点具有生物功能性,因此我们统计了29个蛋白质复合体包含332个丙胺酸点突变(alanine mutations)的资料,可以发现在锚点内的残基被突变后结合能的改变(ΔΔG)都会比锚点外的残基还要大。 在此我们利用forkhead-associated domain (FHA domain)的蛋白质家族来展现site-site interaction maps的概念,根据这些结果我们推论出在许多含有FHA domain的蛋白质中,为什么其可以与不同的下游蛋白质结合且执行不同的功能。接着利用非小细胞肺癌病患EGFR对下游蛋白质磷酸化的蛋白质体资料,包含wild type和L858R突变两种型态,发现EGFR两种突变型态对于下游蛋白质的磷酸化程度具有差异,因此藉由探讨两种形态上在蛋白质交互作介面的锚点,根据突变影响到结构部分以及专一性锚点解释两种EGFR型态对下游蛋白质的磷酸化为什么具有差异,并且利用已知结合在Mdm2家族上的抑制剂,来验证我们所定义的锚点其物化特性是否与蛋白质-抑制剂间的交互作用力相似,实验结果显示抑制剂会结合到的残基多会被我们辨识在锚点内,因此我们相信可以利用此新概念来分析蛋白质复合体的交互作用机制,以及做为未来设计多标靶或是专一性药物的参考。 Studying protein-protein interactions (PPIs) mechanisms is essential to understand cellular processes and drug discovery. Many studies of PPIs have been applied to infer the hotspots in a given protein-protein interface by considered the conservation of a single residue or pair of residues. However, these approaches often were residue-based methods and lacked of the site-site interactions for PPI interfaces. To elucidate functional and structural roles of a PPI interface, identifying a group of contact residue pairs (site-site interactions), which are usually compact region and conserved interaction across multiple species, is very important for understanding the mechanisms of PPI interfaces. In our previous studies, we have proposed a concept, SiMMap, which uses immense screening compounds to describe the relationship between the moiety preferences and physicochemical properties of the binding site. Here, we use the concept of SiMMap and the PPI family to characterize the PPI interfaces. In this thesis, we propose a new concept, called site-site interaction map to analyze the binding environments of a PPI interface. To identify binding environments of a PPI interface, the site-site map can be used to divide a PPI interface into several conserve and packing regions (called anchors), which residues are highly conserved and play a key role in certain biological functions. In addition, the anchors can be distinguished into two types, core and specific anchor. The core anchor often possesses conserved physicochemical properties between PPI families, and the specific anchor often forms a selective environment for a target PPI interface. To verify the identified anchors, we collected 29 complexes with 332 mutated residues in the database of alanine scanning mutagenesis. Our experimental results showed that the binding free energies (ΔΔG) of anchor residues are statistically significant higher than the non-anchor residues in the PPI interfaces. Here, we use the site-site interaction map to analyze FHA domain families. The results may explain why the various FHA families can recognize the different substrates and trigger different biological functions. Furthermore, we applied our method to identify the site-site interaction map of wild type (WT) and L858R EGFR, which are common mutated types of EGFR in non-small lung cancer. According to the structures and the specific anchor may explain the differently interacting mechanisms between WT and L858R EGFR. We also applied the concept to Mdm2 proteins complexes. According to the known inhibitors of the Mdm2 family, we found that PPI anchors shared similar physicochemical properties with protein-ligand interactions. In addition, the consensus interacting residues of protein-ligand are usually located on the PPI anchors. These results showed that our concept can reflect the biological functions and binding mechanisms of the Mdm2 family. We believe that site-site interaction maps can be used to explore mechanisms of PPI and design PPI inhibitors. |
URI: | http://etd.lib.nctu.edu.tw/cdrfb3/record/nctu/#GT070357211 http://hdl.handle.net/11536/139791 |
显示于类别: | Thesis |