Phosphotyrosine binding domains of Shc and insulin receptor substrate 1 recognize the NPXpY motif in a thermodynamically distinct manner
Phosphotyrosine binding domains of Shc and insulin receptor substrate 1 recognize the NPXpY motif in a thermodynamically distinct manner
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DOI：
10.1074/jbc.274.10.6114
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发表时间：
1999-03-05
影响因子：
4.8
通讯作者：
Zhou, MM
中科院分区：
文献类型：
--
作者：
Farooq, A;Plotnikova, O;Zhou, MM
适配蛋白Shc和胰岛素受体底物（IRS-1）的磷酸酪氨酸结合（PTB）结构域与一组不同的激活和酪氨酸磷酸化的细胞因子和生长因子受体相互作用，并在介导有丝分裂信号转导中发挥重要作用。我们利用等温滴定量热法研究了Shc和IRS-1 PTB结构域与已知受体结合位点衍生的酪氨酸磷酸化含npxy肽结合的热力学。结果表明，焓和熵对结合自由能的相对贡献依赖于特定的磷酸肽。Shc PTB结构域与TrkA、表皮生长因子、ErbB3和胰岛素受体酪氨酸磷酸化肽的结合是通过整体熵驱动的反应实现的。另一方面，IRS-1 PTB结构域对胰岛素和白介素-4受体磷酸肽的识别主要是一个焓驱动的过程。诱变和氨基酸替换实验表明，除了酪氨酸磷酸化的NPXY基序外，Shc和IRS-1的PTB结构域在pY-5和pY-1上分别有一个大的疏水残基（其中pY为磷酸酪氨酸）。这些结果与计算出的PTB结构域/肽结构中这两个关键肽残基的溶剂可及性一致，并支持了Shc和IRS-1的PTB结构域采用不同的功能机制来识别酪氨酸磷酸化受体的观点。
Phosphotyrosine binding (PTB) domains of the adaptor protein Shc and insulin receptor substrate (IRS-1) interact with a distinct set of activated and tyrosine-phosphorylated cytokine and growth factor receptors and play important roles in mediating mitogenic signal transduction. By using the technique of isothermal titration calorimetry, we have studied the thermodynamics of binding of the Shc and IRS-1 PTB domains to tyrosine-phosphorylated NPXY-containing peptides derived from known receptor binding sites. The results showed that relative contributions of enthalpy and entropy to the free energy of binding are dependent on specific phosphopeptides. Binding of the Shc PTB domain to tyrosine-phosphorylated peptides from TrkA, epidermal growth factor, ErbB3, and insulin receptors is achieved via an overall entropy-driven reaction. On the other hand, recognition of the phosphopeptides of insulin and interleukin-4 receptors by the IRS-1 PTB domain is predominantly an enthalpy-driven process. Mutagenesis and amino acid substitution experiments showed that in addition to the tyrosine-phosphorylated NPXY motif, the PTB domains of Shc and IRS-1 prefer a large hydrophobic residue at pY-5 and a small hydrophobic residue at pY-1, respectively (where pY is phosphotyrosine). These results agree with the calculated solvent accessibility of these two key peptide residues in the PTB domain/peptide structures and support the notion that the PTB domains of Shc and IRS-1 employ functionally distinct mechanisms to recognize tyrosine-phosphorylated receptors.