Murphy C I, Bikel I, Livingston D M
J Virol. 1986 Sep;59(3):692-702. doi: 10.1128/JVI.59.3.692-702.1986.
When crude, radiolabeled extracts of various cells were applied to homogeneous simian virus 40 small t antigen-Sepharose adsorbents, three cell proteins (57, 32, and 20 kilodaltons [kDa]) bound specifically. Each also bound to an insoluble, truncated t derivative composed of the COOH-terminal 123 residues of the protein. The binding of these proteins was greatly inhibited after reduction and alkylation of the t ligand. Therefore, some element of native conformation, but not all of the primary structure of t, is necessary for this binding property, which may constitute a discrete, in vitro biochemical function of this protein. Results of cell fractionation experiments suggested that the 57- and 32-kDa proteins are nonnuclear cell constituents, whereas the 20-kDa protein was closely associated with a detergent-washed nuclear fraction. Specific immunoblotting and comparative partial proteolytic digestion analyses indicated that the 57-kDa protein is tubulin, a major component of the cytoskeleton. In this regard, t and tubulin were observed to coimmunoprecipitate from crude cell extracts after incubation with monospecific anti-t antibody. Therefore, it is possible that t and tubulin interact in vivo.
当将各种细胞的粗制放射性标记提取物应用于均一的猴病毒40小t抗原-琼脂糖凝胶吸附剂时，有三种细胞蛋白（57、32和20千道尔顿[kDa]）特异性结合。每种蛋白也与一种由该蛋白的COOH末端123个残基组成的不溶性截短t衍生物结合。在对t配体进行还原和烷基化后，这些蛋白的结合受到极大抑制。因此，天然构象的某些要素而非t的所有一级结构对于这种结合特性是必需的，这可能构成该蛋白一种独特的体外生化功能。细胞分级分离实验结果表明，57 kDa和32 kDa的蛋白是非核细胞成分，而20 kDa的蛋白与经去污剂洗涤的核部分紧密相关。特异性免疫印迹和比较性部分蛋白酶消化分析表明，57 kDa的蛋白是微管蛋白，是细胞骨架的主要成分。在这方面，在用单特异性抗t抗体孵育后，观察到t和微管蛋白从粗制细胞提取物中共免疫沉淀。因此，t和微管蛋白在体内相互作用是有可能的。
