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首页> 外文期刊>Molecular and Cellular Biology >Physical and functional interactions between SH2 and SH3 domains of the Src family protein tyrosine kinase p59fyn.
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Physical and functional interactions between SH2 and SH3 domains of the Src family protein tyrosine kinase p59fyn.

机译：Src家族蛋白酪氨酸激酶p59fyn的SH2和SH3域之间的物理和功能相互作用。

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The Src family protein tyrosine kinases participate in signalling through cell surface receptors that lack intrinsic tyrosine kinase domains. All nine members of this family possess adjacent Src homology (SH2 and SH3) domains, both of which are essential for repression of the enzymatic activity. The repression is mediated by binding between the SH2 domain and a C-terminal phosphotyrosine, and the SH3 domain is required for this interaction. However, the biochemical basis of functional SH2-SH3 interaction is unclear. Here, we demonstrate that when the SH2 and SH3 domains of p59fyn (Fyn) were present as adjacent domains in a single protein, binding of phosphotyrosyl peptides and proteins to the SH2 domain was enhanced, whereas binding of a subset of cellular polypeptide ligands to the SH3 domain was decreased. An interdomain communication was further revealed by occupancy with domain-specific peptide ligands: occupancy of the SH3 domain with a proline-rich peptide enhanced phosphotyrosine binding to the linked SH2 domain, and occupancy of the SH2 domain with phosphotyrosyl peptides enhanced binding of certain SH3-specific cellular polypeptides. Second, we demonstrate a direct binding between purified SH2 and SH3 domains of Fyn and Lck Src family kinases. Heterologous binding between SH2 and SH3 domains of closely related members of the Src family, namely, Fyn, Lck, and Src, was also observed. In contrast, Grb2, Crk, Abl, p85 phosphatidylinositol 3-kinase, and GTPase-activating protein SH2 domains showed lower or no binding to Fyn or Lck SH3 domains. SH2-SH3 binding did not require an intact phosphotyrosine binding pocket on the SH2 domain; however, perturbations of the SH2 domain induced by specific high-affinity phosphotyrosyl peptide binding abrogated binding of the SH3 domain. SH3-SH2 binding was observed in the presence of proline-rich peptides or when a point mutation (W119K) was introduced in the putative ligand-binding pouch of the Fyn SH3 domain, although these treatments completely abolished the binding to p85 phosphatidylinositol 3-kinase and other SH3-specific polypeptides. These biochemical SH2-SH3 interactions suggest novel mechanisms of regulating the enzymatic activity of Src kinases and their interactions with other proteins.
机译：Src家族蛋白酪氨酸激酶通过缺乏内在的酪氨酸激酶结构域的细胞表面受体参与信号传导。该家族的所有九个成员都具有相邻的Src同源性（SH2和SH3）域，这两个域对于抑制酶活性都是必不可少的。抑制是通过SH2域和C末端磷酸酪氨酸之间的结合来介导的，而SH3域是这种相互作用所必需的。但是，尚不清楚功能性SH2-SH3相互作用的生化基础。在这里，我们证明了当p59fyn（Fyn）的SH2和SH3结构域作为单个蛋白中的相邻结构域存在时，磷酸酪氨酰肽和蛋白与SH2结构域的结合得到增强，而细胞多肽配体的一个子集与SH3域减少。通过域特异性肽配体的占据进一步揭示了域间通信：占据富含脯氨酸的肽的SH3结构域增强了磷酸酪氨酸与连接的SH2结构域的结合，占据了脯氨酸的肽的SH2结构域增强了某些SH3-的结合。特定的细胞多肽。其次，我们证明了Fyn和Lck Src家族激酶的纯化SH2和SH3域之间的直接结合。还观察到Src家族密切相关的成员Fyn，Lck和Src的SH2和SH3域之间的异源结合。相反，Grb2，Crk，Abl，p85磷脂酰肌醇3-激酶和GTPase激活蛋白SH2结构域显示与Fyn或Lck SH3结构域的结合较低或没有结合。 SH2-SH3结合不需要在SH2域上完整的磷酸酪氨酸结合口袋;然而，由特定的高亲和力磷酸酪氨酰肽结合诱导的SH2结构域的干扰消除了SH3结构域的结合。在富含脯氨酸的肽存在下或在Fyn SH3结构域的假定配体结合袋中引入点突变（W119K）时观察到SH3-SH2结合，尽管这些处理完全消除了与p85磷脂酰肌醇3激酶的结合和其他SH3特异性多肽。这些生化SH2-SH3相互作用表明调节Src激酶及其与其他蛋白质的相互作用的酶活性的新机制。

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