Wiederrecht G, Hung S, Chan H K, Marcy A, Martin M, Calaycay J, Boulton D, Sigal N, Kincaid R L, Siekierka J J
Department of Immunology Research, Merck Research Laboratories, Rahway, New Jersey 07065.
J Biol Chem. 1992 Oct 25;267(30):21753-60.
The immunoregulant FK-506 potently inhibits particular calcium-associated signal transduction events that occur early during T-lymphocyte activation and during IgE receptor-mediated exocytosis in mast cells. FK-506 binds to a growing family of receptors termed FK-506-binding proteins (FKBPs), the most abundant being a 12-kDa cytosolic receptor, FKBP12. To date, there is no formal evidence proving that FKBP12 is the sole receptor mediating the immunosuppressive effects or toxic side effects of FK-506. Using gel filtration chromatography as an assay for novel FK-506-binding proteins, we identified FK-506 binding activities in extracts prepared from calf brain and from JURKAT cells. Both of these new activities comigrated with apparent molecular masses of 110 kDa. However, further characterization of both binding activities revealed that the two are not identical. The 110-kDa activity observed in brain extracts appears to be the FKBP12.FK-506.calcineurin (CaN) complex previously reported (Liu, J., Farmer, J., Lane, W., Friedman, J., Weissman, I., and Schreiber, S. (1991) Cell 66, 807-815) while the 110 kDa activity observed in JURKAT cells is a novel FK-506-binding protein. Our characterization of the FKBP12.FK-506.CaN complex reveals a dependence upon calmodulin (CaM) for formation of the complex and demonstrates that the peptidyl-prolyl cis-trans isomerase (PPIase) activity of FKBP12 is not required for binding of FKBP12.FK-506 to CaN or for inhibition of CaN phosphatase activity. The novel FK-506-binding protein in JURKAT cells has been purified to homogeneity, migrates with an apparent mass of 51 kDa on denaturing gels, and has been termed FKBP51. Like FKBP12, FKBP51 has PPIase activity, but, unlike FKBP12.FK-506, FKBP51.FK-506 does not complex with or inhibit the phosphatase activity of, CaN. These results indicate that complex formation with CaN may not be a general property of the FKBPs. Peptide sequencing reveals that FKBP51 may be similar, if not identical, to hsp56, a component of non-transformed steroid receptors.
免疫调节剂FK-506能有效抑制T淋巴细胞激活早期以及肥大细胞中IgE受体介导的胞吐过程中发生的特定钙相关信号转导事件。FK-506与一类不断增加的受体结合，这类受体被称为FK-506结合蛋白（FKBPs），其中最丰富的是一种12 kDa的胞质受体FKBP12。迄今为止，尚无正式证据证明FKBP12是介导FK-506免疫抑制作用或毒副作用的唯一受体。我们使用凝胶过滤色谱法作为检测新型FK-506结合蛋白的方法，在从小牛脑和JURKAT细胞制备的提取物中鉴定出了FK-506结合活性。这两种新活性的迁移率对应的表观分子量均为110 kDa。然而，对这两种结合活性的进一步表征表明它们并不相同。在脑提取物中观察到的110 kDa活性似乎是先前报道的FKBP12.FK-506.钙调神经磷酸酶（CaN）复合物（Liu, J., Farmer, J., Lane, W., Friedman, J., Weissman, I., and Schreiber, S. (1991) Cell 66, 807 - 815），而在JURKAT细胞中观察到的110 kDa活性是一种新型FK-506结合蛋白。我们对FKBP12.FK-506.CaN复合物的表征揭示了该复合物的形成依赖于钙调蛋白（CaM），并证明FKBP12的肽基脯氨酰顺反异构酶（PPIase）活性对于FKBP12.FK-506与CaN的结合或抑制CaN磷酸酶活性并非必需。JURKAT细胞中的新型FK-506结合蛋白已被纯化至同质，在变性凝胶上的表观分子量为51 kDa，被称为FKBP51。与FKBP12一样，FKBP51具有PPIase活性，但与FKBP12.FK-506不同，FKBP51.FK-506不与CaN形成复合物，也不抑制其磷酸酶活性。这些结果表明与CaN形成复合物可能并非FKBPs的普遍特性。肽序列分析表明FKBP51可能与非转化类固醇受体的一个组分hsp56相似，甚至相同。
