Albert-Wolf M, Meuer S C, Wallich R
Department of Applied Immunology, German Cancer Research Center, Heidelberg.
Int Immunol. 1991 Dec;3(12):1335-47. doi: 10.1093/intimm/3.12.1335.
To produce large quantities of recombinant CD58 (rCD58) glycoproteins for biochemical and functional studies, a cDNA clone containing the phosphatidylinositol-linked form of human CD58 was expressed in insect cells using the baculovirus system. Gel filtration showed rCD58 to form soluble oligomeric aggregates which were functionally, antigenically, and biochemically similar to their natural counterpart. Sequence analysis of the amino- and carboxy-terminal ends of released rCD58 protein revealed that the 28 amino acid signal peptide was accurately removed. In contrast, the hydrophobic C-terminal peptide was not removed. rCD58 binds to its natural ligand CD2 with a dissociation constant Kd = 5 x 10(-8) M, which is equivalent to the affinity of physiological T cell adhesion mediated by the membrane bound CD2-CD58 receptor-ligand pair. Rosette formation of human T lymphocytes with sheep and human erythrocytes was completely abrogated. In addition, the mixed lymphocyte reaction was significantly inhibited by rCD58. Moreover, cytotoxicity of human NK clones (CD2+CD3-) was inhibited by rCD58 similar to inhibition by CD58 mAbs. In contrast, rCD58 synergized with mitogenic CD2R mAbs in T cell triggering. These data demonstrate that rCD58 might serve as a biological immunomodulator which influences T cell adhesion and activation.
为了大量生产用于生化和功能研究的重组CD58（rCD58）糖蛋白，使用杆状病毒系统在昆虫细胞中表达了一个包含人CD58磷脂酰肌醇连接形式的cDNA克隆。凝胶过滤显示rCD58形成可溶性寡聚聚集体，其在功能、抗原性和生化性质上与其天然对应物相似。对释放的rCD58蛋白的氨基末端和羧基末端进行序列分析表明，28个氨基酸的信号肽被准确切除。相比之下，疏水的C末端肽未被切除。rCD58以解离常数Kd = 5×10^(-8) M与其天然配体CD2结合，这等同于由膜结合的CD2 - CD58受体 - 配体对介导的生理性T细胞黏附的亲和力。人T淋巴细胞与绵羊和人红细胞的玫瑰花结形成被完全消除。此外，rCD58显著抑制混合淋巴细胞反应。而且，rCD58对人NK克隆（CD2 + CD3 -）的细胞毒性的抑制作用与CD58单克隆抗体的抑制作用相似。相反，rCD58在T细胞触发中与促有丝分裂的CD2R单克隆抗体协同作用。这些数据表明rCD58可能作为一种生物免疫调节剂，影响T细胞黏附和激活。
