Gulbins E, Schlottmann K, Brenner B, Lang F, Coggeshall K M
I. Institute of Physiology, University of Tuebingen, Germany.
Biochem Biophys Res Commun. 1995 Dec 26;217(3):876-85. doi: 10.1006/bbrc.1995.2853.
Vav has been shown to activate Ras (1-3) and is regulated by tyrosine phosphorylation (1) or binding of diglycerides (3) to the cysteine rich domain. In the present study employing different Ras activation assay techniques using [3H]GDP release or [32P]alpha GTP-binding from membrane-bound or soluble recombinant Ras, we demonstrate that Ras activity can be increased by tyrosine phosphorylated Vav upon cellular stimulation via the IL-2 receptor or the TCR/CD3-complex. Increase of [32P]alpha GTP-binding to Ras catalyzed by phosphorylated Vav is similar to the activity of immunoprecipitated Sos. The activity of Vav measured by binding of [32P]alpha GTP to Ras was linear with respect to the concentration of Vav protein used. To study molecular characteristics of this Vav-Ras interaction, we used several Ras mutants and demonstrate that Vav activity towards Ras depends on the integrity of the same or similar domains as Ras activation by SDC 25 or CDC 25.
已证实Vav可激活Ras（1-3），且受酪氨酸磷酸化（1）或二酰甘油与富含半胱氨酸结构域的结合（3）调控。在本研究中，我们采用不同的Ras激活检测技术，利用[3H]GDP释放或膜结合型或可溶性重组Ras的[32P]αGTP结合，证明在通过IL-2受体或TCR/CD3复合物进行细胞刺激时，酪氨酸磷酸化的Vav可增加Ras活性。磷酸化的Vav催化的[32P]αGTP与Ras的结合增加与免疫沉淀的Sos活性相似。通过[32P]αGTP与Ras的结合来测量的Vav活性与所用Vav蛋白的浓度呈线性关系。为了研究这种Vav-Ras相互作用的分子特征，我们使用了几种Ras突变体，并证明Vav对Ras的活性取决于与SDC 25或CDC 25激活Ras相同或相似结构域的完整性。
