IMMUNOPHILIN AND RELATED PROTEINS

免疫抑制剂药物的结合蛋白是最近发现的 与细胞质类固醇的寡聚体结构有关， 受体。结合蛋白与已知的 免疫抑制药物，如雷帕霉素、FK 5 O 6和环孢菌素A， 因为免疫系统的功能有升有降 随着年龄的增长和其他条件的系统，似乎有可能存在 一种或多种内源性免疫抑制剂，其水平可低于 各种控制。 在研究重新组装的 糖皮质激素受体非DNA结合复合物形式与纯化 我们发现了一种新的与FKBP-52（p55-59）复合的蛋白质 亲免素。 因为这种新蛋白质是用来自 不同的细胞系，它只出现在FKBP-52出现时，我们得出结论， 它可能在信号转导途径中起作用， 免疫抑制作用。 因此，我们计划开展以下工作： 目的：（1）全长、截短和突变的FKBP-52将被 在杆状病毒系统中过表达。定点诱变将是 然后是突变蛋白的过度表达， 蛋白质将被纯化。(2)过量表达和纯化的FKBP-52、hsp 90、 hsp 70和其它胞质因子将被重新构建成高水平的 分子量复合物，与细胞复合物相当， 免疫抑制药物配体对这些复合物的作用将是 评估。(3)59 kDa FKBP-52相关的新磷蛋白将被 克隆、过表达、纯化和表征。(4)中的序列 FKBP-52负责结合hsp 9 O、hsp 7 O、ATP和HSP 7 O。 59 kDa FKBP-52-相关蛋白将通过突变FKBP-52-相关蛋白来确定。 FKBP-52 cDNA，衍生蛋白的过表达和组装成 高分子量复合物。 (5)FKBP特异性抗体- 52及其相关的磷蛋白将被开发。 这些实验 将阐明信号转导过程中涉及的途径， 药物诱导的免疫抑制，并可能有助于所需的方法 发现内源性免疫抑制剂，结果将直接 影响器官和组织移植。

Binding proteins for immunosuppressant drugs have been discovered recently in connection with the oligomeric structures of cytoplasmic steroid receptors. The binding proteins form complexes with known immunosuppressant drugs, such as rapamycin, FK5O6 and cyclosporin A and because there are elevations and declines in the functioning of the immune system with aging and other conditions, it seems likely that there exist one or more endogenous immunosuppressant agents whose levels may be under various types of control. In studying the reassembly of the glucocorticoid receptor non-DNA-binding complex form with purified proteins, we discovered a new protein complexed with the FKBP-52 (p55-59) immunophilin. Because this new protein is purified with the FKBP-52 from different cell lines and it appears only when FKBP-52 appears, we conclude that it may function in the signal transduction pathway of immunosuppressant action. Consequently, we plan to pursue the following objectives: (1) Full-length, truncated and mutated FKBP-52 will be overexpressed in the baculovirus system. Site directed mutagenesis will be followed by overexpression of the mutant proteins and all of these proteins will be purified. (2) Overexpressed and purified FKBP-52, hsp90, hsp7O and other cytosolic factors will be reconstituted into high molecular weight complexes, comparable to cellular complexes, and the effects of immunosuppressant drug ligands on these complexes will be assessed. (3) The 59kDa FKBP-52-associated new phosphoprotein will be cloned, overexpressed, purified and characterized. (4) The sequences in FKBP-52 that are responsible for binding to hsp9O, hsp7O, ATP and the 59kDa FKBP-52-associated protein will be determined by mutation of the FKBP-52 cDNA, overexpression of the derivative protein and assembly into the high molecular weight complex. (5) Specific antibodies against FKBP- 52 and its associated phosphoprotein will be developed. These experiments will clarify the signal transduction process involved in the pathway of drug-induced immunosuppression and may contribute to the methods required to discover the endogenous immunosuppressants, results that will directly affect organ and tissue transplantation.