Structural Evaluation of HIV-1 Envelope Gycoprotein

关于HIV糖蛋白(gp120和gp41)的结构信息可以帮助设计基于包膜的 抗原，以及更好地了解艾滋病毒融合过程和宿主免疫反应。主要目标 项目2的目的是对HIV-1包膜糖蛋白的结构进行表征，以帮助设计HIV-1 可以诱导广泛交叉反应的中和抗体的包膜抗原，长期目标是 开发一种基于包膜的预防艾滋病毒感染的保护性疫苗。 目前已知的所有gp41结构彼此非常相似，并且代表融合后构象。 Gp41融合前中间体的结构尚不清楚。我们建议对精心设计的结构进行解决 不含N-末端七肽区的gp41片段。我们的方法包括一套精心挑选的 以及最先进的结构和计算生物学方法，包括结晶学、羟基 激进的足迹和从头开始的建模。我们目前的gp41-抗原模型解释了它的抗原性 反应性，并正在推动其工程想法。AIM 1提供无缝的gp41抗原设计-高 解析结构确定-重新设计程序内的流水线作为一个整体。 目的2研究gp120外区(Gp120OD)和OD-gp41的结构特征 结构质谱学方法。Gp120OD的糖基化信号会影响 基于gp120OD的抗原的免疫原性。因此，设计-结构表征-重新设计方法 项目2反馈给项目1团队的位置对于开发改进的基于gp120的抗原至关重要。 目前gp120的晶体结构主要来自蛋白质核心，具有可变的环区和N-和C末端 截断。解决了具有完整可变环的全长gp120的晶体结构 由于这些环段的灵活性和蛋白质的广泛糖基化，这是极其困难的。在……里面 目的3，我们提出用羟基自由基足迹、同源模建和对接等方法来表征 V1/V2可变环区在存在和不存在主要受体CD4的情况下的结构。 通过探测可接触到溶剂的蛋白质的外表面，我们将能够确定 CD4结合时可变环区的构象变化。

Structure information on HIV glycoproteins (gp120 and gp41) can facilitate the design of envelope-based antigens, and better understanding of HIV fusion process and host immune response. The major objective of Project 2 is to structurally characterize HIV-1 envelope glycoproteins to assist the design of HIV-1 envelope antigens that can elicit broadly cross-reactive neutralizing antibodies, with a long-term goal of developing an envelope-based protective vaccine against HIV infection. All currently known gp41 structures are very similar to each other and represent a post-fusion conformation. Structure of gp41 pre-fusion intermediate is not known. We propose to solve structure for carefully designed gp41 fragments without the N-terminal heptad region. Our approach includes a suite of carefully selected and state-of-the art structural and computational biology approaches, including crystallography, hydroxyl radical footprinting and ab initio modeling. Our current model of the gp41-64 antigen explains its antigenic reactivity and is driving ideas for its engineering. Aim 1 provides a seamless gp41 antigen design-high resolution structure determination-redesign pipeline within the program as a whole. Aim 2 intends to structurally characterize gp120 outer domain (gp120OD) and OD-gp41 constructs using structural mass spectrometry methods. Glycosylation signature of gp120OD would affect the immunogenicity of gp120OD-based antigens. Thus, a design-structural characterization-redesign approach where Project 2 feeds back to the Project 1 team is critical for developing improved gp120-based antigens. Present crystal structures of gp120 are mostly from protein cores, with variable loop regions and N- and Ctermini truncated. Solving a crystal structure of a full-length gp120 with intact variable loops has been extremely difficult due to the flexibility of these loop segments and extensive glycosylation of the protein. In Aim 3, we propose to use, hydroxyl radical footprinting, homology modeling, and docking to characterize the structure of the V1/V2 variable loop regions in the presence and in the absence of the primary receptor CD4. By probing the outer surfaces of proteins accessible to solvents, we will be able to determine the conformational changes in the variable loop regions upon binding of CD4.