C3d/EBV Receptor Ligand Binding Sites

描述(申请人提供)：人补体受体2型(CR2/CD21)是一种~145kD的I型跨膜蛋白，作为三类配体的受体。其中包括C3裂解片段(C3d、C3dg和IC3b)、Epstein-Barr病毒gp350/220和CD23。这些配体中的每一个都与受体的氨基末端区域在16个重复元件中的2个相互作用，这些重复元件被指定为短共识重复(SCR)。与补体C3和/或C4相互作用的含有SCR的蛋白质是被称为补体激活调节因子(RCA)的家族的一部分。虽然这些蛋白质的生物学相关性已经得到了很好的证实，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作为这个家族中受体-配体相互作用的模型，最初专注于氨基末端结构域，在这里被指定为SCR1-2结构域，它与上述每个配体相互作用。使用溶液和X射线结晶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，并强烈建议了一种模型，在该模型中，SCR1-2结构域的非配体结合的自由结构和配体结合结构的构象不同。此外，最近的数据表明，CR2在SCR1-2结构域之外的SCR影响与C3d的结合。我们现在建议通过追求以下特定目标来扩展这些研究：特定目标#1：确定CR2 SCR1-2结构域的溶液相结构，以便建立非配体结合和C3d配体结合受体状态之间的物理关系。特定目标#2：建立在CR2 SCR1-2中明显的三个独特的蛋白质-蛋白质界面的相对作用：C3d共晶结构在C3d配体结合、信号转导和体内免疫反应的增强中的作用。具体目的#4：建立CR2 SCR1-15/16胞外结构域的溶液结构，并确定CR2-C3d配体-受体结合动力学改变的物理基础，将该全长受体与CR2 SCR1-2结构域进行比较。

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.