Structure-function studies of the membrane-interacting domains of HIV-1 Env spike

项目摘要 HIV-1感染的第一个关键步骤是通过包膜介导的病毒和靶细胞膜的融合 糖蛋白(Env)。成熟的Env棘突[三聚体(Gp160)3，裂解为(gp120/gp41)3]是唯一的抗原 在病毒粒子表面。Gp120与受体结合时的构象变化 受体(如CCR5或CXCR4)导致gp41中一系列的折叠事件，最终导致膜 核聚变。Env的外域有大量的结构信息，但我们知道的要少得多 关于其跨膜结构域(TMD)及其膜近端(MP)区域，包括膜 近端外区(MPER)和胞浆尾部(CT)。我们最近有了一个意想不到的发现 CT域的截断极大地重塑了包膜外区的抗原性表面 膜的另一边。对物理耦合(构象和/或动力学)有深入的了解 TMD介导的CT和胞外区之间的关系可能指导基于Env的免疫原设计 广谱中和抗体(BnNAb)。因此我们假设，膜相互作用的区域 HIV-1环境蛋白(MPER、TMD和CT)采用特定的结构，这些结构对其稳定性、功能至关重要 和抗原性。我们已经完成了原子分辨率的TMD结构，使用的是状态- ART核磁共振技术。当TMD在模拟脂质双层的双分子中重组时，TMD形成有序的 三聚体主要由其C-末端附近紧密堆积的亲水性核心稳定，该核心可能是 受CT域的影响。在这项应用中，我们计划将结构生物学方法和 功能分析以阐明HIV-1env膜相互作用结构域的作用。我们会把钱花在 具体目标如下：1)我们将研究HIV和SIV Env的TMD组装以及可能的相互作用 2)我们将在以下背景下提供Env的MP区域的结构信息 我们将阐明env的膜相互作用结构域在其稳定性、功能中的作用。 和抗原性；4)我们将设计三聚体免疫原来模拟天然的和功能强大的HIV-1环境蛋白尖峰。

Project Summary The first critical step of HIV-1 infection is fusion of viral and target cell membranes mediated by envelope glycoprotein (Env). The mature Env spikes [trimeric (gp160)3, cleaved to (gp120/gp41)3] are the sole antigens on the virion surface. Conformational changes in gp120 when triggered by binding to receptor (CD4) and co- receptor (e.g., CCR5 or CXCR4) lead to a cascade of refolding events in gp41, and ultimately to membrane fusion. Vast amount of structural information is available for the ectodomain of Env, but much less is known regarding its transmembrane domain (TMD) and its membrane-proximal (MP) regions, including the membrane proximal external region (MPER) and the cytoplasmic tail (CT). We recently made an unexpected discovery that truncation of the CT domain drastically reshapes the antigenic surfaces of the Env ectodomain on the other side of the membrane. Deep understanding of the physical coupling (conformation and/or dynamics) between the CT and the ectodomain mediated by the TMD may guide Env-based immunogen design to induce broadly neutralizing antibodies (bnNabs). We thus hypothesize that the membrane-interacting domains (MPER, TMD and CT) of HIV-1 Env adopt defined structures which are critical for its stability, function and antigenicity. We have already completed a TMD structure at the atomic resolution using the state-of-the- art NMR technology. When reconstituted in bicelles that mimic lipid bilayer, the TMD forms a well-ordered trimer mainly stabilized by a tightly packed hydrophilic core near its C-terminal end that can potentially be influenced by the CT domain. In this application, we plan to combine structural biology approaches and functional assays to elucidate the roles of the membrane-interacting domains of HIV-1 Env. We will purse the following specific aims: 1) we will investigate TMD assembly of HIV and SIV Env and possible interaction with the fusion peptide; 2) we will provide structural information for the MP regions of Env in the context of membrane; 3) we will elucidate the role of the membrane-interacting domains of Env in its stability, function and antigenicity; 4) we will design trimer immunogens to mimic the native and functional HIV-1 Env spike..