Structural Evaluation of HIV-1 Envelope Gycoprotein

HIV糖蛋白（gp 120和gp 41）的结构信息可以促进基于酶的设计， 抗原，更好地了解艾滋病毒融合过程和宿主免疫反应。主要目的 项目2的第一个目标是从结构上表征HIV-1包膜糖蛋白，以帮助设计HIV-1 包膜抗原，可以引发广泛的交叉反应性中和抗体，长期目标是 开发一种基于疫苗的预防艾滋病毒感染的疫苗。 所有目前已知的gp 41结构彼此非常相似，并代表融合后构象。 gp 41融合前中间体的结构未知。我们建议解决精心设计的结构 没有N-末端七肽区的gp 41片段。我们的方法包括一套精心挑选的 和最先进的结构和计算生物学方法，包括晶体学，羟基 激进的足迹法和从头建模。我们目前的gp 41 -64抗原模型解释了它的抗原性。 反应性，并正在推动其工程设计的想法。Aim 1提供了一种无缝的gp 41抗原设计-高 解析结构的确定--整个程序内的重新设计流水线。 目的2旨在使用以下方法在结构上表征gp 120外部结构域（gp 120 OD）和OD-gp 41构建体： 结构质谱法。gp 120 OD的糖基化特征会影响 基于gp 120 OD的抗原的免疫原性。因此，设计-结构特性-再设计方法 其中项目2反馈给项目1团队对于开发改进的基于gp 120的抗原至关重要。 目前gp 120的晶体结构主要来自蛋白质核心，具有可变环区和N-和C-末端， 被截断了求解具有完整可变环的全长gp 120的晶体结构， 由于这些环片段的柔性和蛋白质的广泛糖基化，在 目的3，我们建议使用羟基自由基足迹，同源模建和对接来表征 在存在和不存在初级受体CD 4的情况下，V1/V2可变环区的结构。 通过探测溶剂可接近的蛋白质的外表面，我们将能够确定 结合CD 4后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.