Garcia G G, Evans G A, Michiel D F, Farrar W L
Biological Carcinogenesis and Development Program, NCI-Frederick Cancer Research and Development Center, MD 21702-1201.
Biochem J. 1992 Aug 1;285 ( Pt 3)(Pt 3):851-6. doi: 10.1042/bj2850851.
The IL-2 receptor complex is minimally composed of two genetically unrelated subunits of relative molecular masses 55 and 75 kDa respectively. Structural information deduced from the cDNA sequences of either subunit have not revealed significant information as to the basis of the mechanisms of IL-2 receptor signal transduction. Nevertheless, IL-2 stimulates the activation of one or more tyrosine kinases requiring the functional participation of the p75 member of the receptor complex. Here we have developed the methods to isolate the receptor complex with an associated tyrosine protein kinase. Extracts of membrane glycoproteins from activated normal human T lymphocytes and cell lines demonstrated catalytic activation of tyrosine kinase activity when stimulated with IL-2. Purification of the receptor complex with biotinylated IL-2 revealed the presence of two dominant phosphotyrosyl-proteins of approximate molecular masses 58 and 97 kDa. Denaturation gel electrophoresis followed by renaturation of proteins associated with the IL-2 receptor complex demonstrated that the 97 kDa protein had catalytic autophosphorylation activity. The results indicate that the 58 and 97 kDa phosphotyrosyl-proteins can be found to co-precipitate with the IL-2 receptor complex and that the 97 kDa protein was demonstrated to have protein kinase activity. The association of such kinases with receptors devoid of catalytic structure may represent a unique paradigm of growth-factor receptor mechanisms.
白细胞介素-2受体复合物至少由两个相对分子质量分别为55 kDa和75 kDa的基因不相关的亚基组成。从任一亚基的cDNA序列推导的结构信息尚未揭示关于白细胞介素-2受体信号转导机制基础的重要信息。然而，白细胞介素-2刺激一种或多种酪氨酸激酶的激活，这需要受体复合物中p75成员的功能参与。在此，我们开发了分离与相关酪氨酸蛋白激酶结合的受体复合物的方法。来自活化的正常人T淋巴细胞和细胞系的膜糖蛋白提取物在用白细胞介素-2刺激时显示出酪氨酸激酶活性的催化激活。用生物素化的白细胞介素-2纯化受体复合物揭示存在两种主要的磷酸化酪氨酸蛋白，其近似分子质量分别为58 kDa和97 kDa。变性凝胶电泳，随后使与白细胞介素-2受体复合物相关的蛋白质复性，结果表明97 kDa的蛋白质具有催化自磷酸化活性。结果表明，58 kDa和97 kDa的磷酸化酪氨酸蛋白可与白细胞介素-2受体复合物共沉淀，并且已证明97 kDa的蛋白质具有蛋白激酶活性。此类激酶与缺乏催化结构的受体的结合可能代表生长因子受体机制的一种独特模式。
