C3d/EBV Receptor Ligand Binding Sites

描述（由申请人提供）：人补体受体2型（CR2/CD21）是一种~145 Kd的I型跨膜蛋白，作为三种配体的受体。这些包括C3切割片段（C3d， C3dg和iC3b）， Epstein-Barr病毒gp350/220和CD23。这些配体中的每一个都与16个重复元件中的2个受体的氨基末端区域相互作用，这些重复元件被指定为短一致重复（SCRs）。与补体C3和/或C4相互作用的含有scr的蛋白质是补体激活调节剂（RCA）家族的一部分。虽然这些蛋白质的生物学相关性已经很好地建立，但在分子水平上对含有scr的蛋白质的重要结构-功能特征知之甚少。我们正在研究CR2作为该家族中受体-配体相互作用的模型，最初专注于本文指定的SCR1-2结构域的氨基末端结构域，该结构域与上述每种配体相互作用。使用溶液和x射线晶体学技术的初步研究已经确定了几个蛋白质-蛋白质界面，这些界面可能是CR2与C3d配体受体相互作用的关键，并强烈建议一个模型，其中SCR1-2结构域的非配体结合的自由结构和配体结合的结构在其构象上不同。此外，最近的数据表明，在SCR1-2结构域之外的CR2的SCRs影响与C3d的结合。我们现在建议通过追求以下具体目标来扩展这些研究：具体目标#1：确定CR2 SCR1-2结构域的溶液相结构，以建立非配体结合和c3d配体结合受体状态之间的物理关系。具体目标#2：建立CR2中三个独特的蛋白-蛋白界面的相对作用：CR2 SCR1-2中C3d共晶结构在C3d配体结合、信号转导和增强体内免疫反应中的作用。具体目标#3：确定CR2 SCR1-2结构域中C3d、gp350/200和CD23配体结合位点之间的关系。具体目标#4：建立CR2 SCR1-15/16细胞外结构域的溶液结构，并在将CR2 SCR1-2全长受体与CR2 SCR1-2结构域进行比较时，确定CR2- c3d配体-受体结合动力学改变的物理基础。

DESCRIPTION (provided by applicant): Human complement receptor type 2 (CR2/CD21) is an ~145 Kd Type I transmembrane protein that serves as a receptor for three classes of ligands. These include C3 cleavage fragments (C3d, C3dg and iC3b), Epstein-Barr virus gp350/220 and CD23. Each of these ligands interacts with the amino-terminal region of the receptor within 2 of 16 repetitive elements that have been designated short consensus repeats (SCRs). SCR-containing proteins that interact with complement C3 and/or C4 are part of a family called the Regulators of Complement Activation (RCA). While the biologic relevance of these proteins is well established, important structure-function characteristics of SCR-containing proteins are poorly understood at the molecular level. We are studying CR2 as a model for receptor-ligand interactions in this family, initially focusing on the amino terminal domain that is designated herein the SCR1-2 domain and which interacts with each of the ligands described above. Preliminary studies using solution as well as x-ray crystallographic techniques have identified several protein-protein interfaces that are likely key to receptor interactions of CR2 with the C3d ligand as well as strongly suggested a model in which the non-ligand-bound free and ligand-bound structures of the SCR1-2 domain differ in their conformation. In addition, recent data have shown that SCRs of CR2 outside of the SCR1-2 domain influence binding with C3d. We now propose to extend these studies by pursuing the following specific aims:Specific Aim #1: Determine the solution phase structure of the CR2 SCR1-2 domain in order to establish the physical relationship between the non-ligand-bound and C3d-ligand-bound receptor states.Specific Aim #2: Establish the relative roles of the three unique protein-protein interfaces apparent in the CR2 SCR1-2:C3d co-crystal structure in C3d ligand binding, signal transduction and the enhancement of in vivo immune responses.Specific Aim #3: Determine the relationship between the C3d, gp350/200 and CD23 ligand binding sites within the CR2 SCR1-2 domain.Specific Aim #4: Establish the solution structure of the CR2 SCR1-15/16 extracellular domain and determine the physical basis for the altered CR2-C3d ligand-receptor binding kinetics when comparing this full length receptor with the CR2 SCR1-2 domain.