SEQUENCE REQUIREMENTS FOR BINDING OF SRC FAMILY TYROSINE KINASES TO ACTIVATED GROWTH-FACTOR RECEPTORS
SEQUENCE REQUIREMENTS FOR BINDING OF SRC FAMILY TYROSINE KINASES TO ACTIVATED GROWTH-FACTOR RECEPTORS
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DOI：
10.1074/jbc.270.17.9840
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发表时间：
1995-04-28
影响因子：
4.8
通讯作者：
COURTNEIDGE, SA
中科院分区：
文献类型：
--
作者：
ALONSO, G;KOEGL, M;COURTNEIDGE, SA
生长因子受体蛋白酪氨酸激酶的活化经常导致许多蛋白质与其酪氨酸磷酸化的胞质结构域结合。这些相互作用涉及结合蛋白的SH 2结构域和受体分子上的磷酸化酪氨酸，其中相互作用的特异性由磷酸化酪氨酸周围的氨基酸组成决定，在血小板衍生生长因子（PDGF）受体的情况下，Src家族酪氨酸激酶的主要结合位点在质膜结构域中，并且包括酪氨酸579（Mori，S.，Ronnstrand，L.，Yokote，K.，Engstrom，Angstrom，Courtneidge，S，A，Claesson Welsh，L，and Heldin，C-H.（1993）EMBO J. 12，2257-2264），为了更详细地分析位置579处磷酸化酪氨酸周围的哪些氨基酸对于与Src家族激酶的高亲和力相互作用是重要的，我们合成了一系列对应于该结合位点的磷酸肽，其中单个氨基酸被单独改变，并测试了它们与PDGF受体竞争结合的能力。Fyn，我们发现不仅磷酸化酪氨酸羧基端的三个残基是重要的，而且相对于酪氨酸的位置-1和+4的残基也是必需的，酪氨酸579和581的磷酸化显著增加了竞争效率，激活的集落刺激因子-1（CSF-1）受体，已知其与Src家族激酶相关，在其质膜区域具有与PDGF受体的Tyr 579周围序列相似的序列，并且以该序列为模型的磷酸肽在体外竞争Fyn与受体的结合。此外，突变分析表明，这些序列是Src家族激酶与活化的CSF-1受体在体内有效结合所必需的，与PDGF和CSF-1受体的Src家族结合位点相对应的磷酸肽在体外激活Src激酶活性。这些观察结果支持了一种模型，其中Src家族酪氨酸激酶的酶活性由酪氨酸磷酸化肽与激酶的SH 2结构域的分子内和分子间相互作用控制。
Activation of growth factor receptor protein tyrosine kinases frequently results in the binding of numerous proteins to their tyrosine-phosphorylated cytoplasmic domains, These interactions involve the SH2 domains of the binding proteins and phosphorylated tyrosines on the receptor molecules, with the specificity of interaction dictated by the amino acid composition surrounding the phosphorylated tyrosine, In the case of the platelet derived growth factor (PDGF) receptor, the major binding site for Src family tyrosine kinases is in the juxtamembrane domain and includes tyrosine 579 (Mori, S,, Ronnstrand, L., Yokote, K., Engstrom, Angstrom, Courtneidge, S, A, Claesson Welsh, L,, and Heldin, C-H. (1993) EMBO J. 12, 2257-2264), To analyze in more detail which amino acids surrounding the phosphorylated tyrosine at position 579 were important for high affinity interaction with Src family kinases, we synthesized a series of phosphopeptides corresponding to this binding site in which single amino acids were individually changed and tested their ability to compete with the PDGF receptor for binding of Fyn, We found that not only the three residues carboxyl-terminal to the phosphorylated tyrosine were important but that also residues at positions -1 and +4 relative to the tyrosine were required, Phosphorylation of both tyrosines 579 and 581 significantly increased competition efficiency, The activated colony stimulating factor-1 (CSF-1) receptor, which is known to associate with Src family kinases, has a sequence in its juxtamembrane region similar to that surrounding Tyr 579 of the PDGF receptor, and a phosphopeptide modeled on this sequence competed the association of Fyn with the receptor in vitro, Furthermore, mutational analysis demonstrated that these sequences were required for the efficient association of Src family kinases with the activated CSF-1 receptor in vivo, Phosphopeptides corresponding to the Src family binding sites of both PDGF and CSF-1 receptors activated Src kinase activity in vitro, These observations support a model in which the enzymatic activity of Src family tyrosine kinases is controlled by intra- and intermolecular interactions of tyrosine phosphorylated peptides with the SH2 domain of the kinases.