NMR INVESTIGATION OF HUMAN CELL SURFACE RECEPTOR CD58: IMMUNOLOGY

本研究项目的目标是确定NMR 人细胞表面粘附结构域的溶液结构 受体CD 58，研究其与反受体CD 2 并开发用于生产无聚糖形式的 糖蛋白没有丧失功能。 主要结果是 1998年的研究项目是成功的核磁共振结构 确定CD 58粘附结构域的可溶性突变形式， 通过基于结构的合理化设计， 表面残基的非极性至极性突变 功能不相关的表位。 这种基于结构的成功 这种方法通过大肠杆菌表达得到了证明。 功能性11 kDa粘附结构域提取自重 糖基化的55 kDa人CD 58胞外域。 解的结构 随后使用NOESY测定CD 58粘附结构域。 其他关键数据是从瓦里安750兆赫光谱仪获得的 在CMR，我们还通过NMR研究了与其 反受体CD 2。 已经表明，CD 58粘附结构域 采用免疫球蛋白可变结构域折叠， β链形成两个平行的β片层。 新结构 信息支持CD 2/CD 58相互作用的“握手”模型 涉及CD 2和CD 58粘附结构域的GFCCU面。 的 CD 58的CC环周围的区域最有可能负责 结合特异性，而带负电荷的残基周围的 FG环附近的凹区似乎主要有助于引导 在两个接收器对接期间的定向。 结构性 迄今为止获得的信息导致有希望的实验， 研究了两者形成的络合物的溶液结构 CD 58和CD 2的粘附结构域。 由于日益复杂的 CD 2/CD 58结合复合物结构，实验数据来自 750 Nfflz光谱仪将具有更大的意义。

The goals of this research project are to determine the NMR solution structure of the adhesion domain of human cell surface receptor CD58, to study its interaction with the counter receptor CD2 and to develop a general strategy for producing a glycan-free form of glycoproteins without loss of function. The major result of this research project during 1998 is the successful NMR structural determination of a soluble mutant form of the CD58 adhesion domain, obtained through a structure-based rationalized design employing apolar-to-polar mutations of surface residues in functionally-irrelevant epitopes. The success of this structure-based approach was demonstrated through the E-coli expression of this functional 11 kDa adhesion domain extracted from the heavily glycosylated 55 kDa human CD58 ectodomain. The solution structure of the CD58 adhesion domain was subsequently determined using the NOESY. Other critical data were obtained from the Varian 750 MHz spectrometer at CMR, where we also studied via NMR the binding to its counter-receptor CD2. It has been shown that the CD58 adhesion domain adopts the immunoglobulin variable domain fold and consists of nine beta strands forming two parallel beta sheets. The new structural information supports a "hand-shake" model of CD2/CD58 interaction involving the GFCCU faces of both CD2 and CD58 adhesion domains. The region around the CC'Ioop of CD58 is most likely responsible for binding specificity while the negatively charged residues around the concave region near FG loop appear to contribute mainly to guiding the orientation during the docking of the two receptors. The structural information obtained thus far leads to promising experiments aimed at studying the solution structure of the complex formed between the two adhesion domains of CD58 and CD2. Due to the increased complexity of the CD2/CD58 binding complex structure, the experimental data from the 750 Nfflz spectrometer will have an even greater significance.