RECOGNITION OF A HIGH-AFFINITY PHOSPHOTYROSYL PEPTIDE BY THE SRC HOMOLOGY-2 DOMAIN OF P56(LCK)
RECOGNITION OF A HIGH-AFFINITY PHOSPHOTYROSYL PEPTIDE BY THE SRC HOMOLOGY-2 DOMAIN OF P56(LCK)
复制标题
DOI：
10.1038/362087a0
复制
发表时间：
1993-03-04
期刊：
影响因子：
64.8
通讯作者：
HARRISON, SC
中科院分区：
文献类型：
--
作者：
ECK, MJ;SHOELSON, SE;HARRISON, SC
Src同源性-2（SH 2）结构域是在许多细胞内信号转导蛋白中发现的约100个氨基酸残基的模块1，2。它们以高亲和力和特异性结合含磷酸酪氨酸的序列1 -4，识别紧邻多肽序列的磷酸酪氨酸3 -12。蛋白质p56 lck（Lck）是一种Src样淋巴细胞特异性酪氨酸激酶13 -15。磷酸肽库筛选最近已被用来推断Lck SH 2结构域的“最佳”结合序列16。在磷酸酪氨酸C-末端的三个位置上对残基Glu、Glu和Ile具有选择性。来自仓鼠多瘤中T抗原EPQpYEEIPIYL的11个残基磷酸肽以约1 nM的解离常数与Lck SH 2结合（参考文献17），其亲和力相当于已知最紧密的SH 2-磷酸肽复合物。我们在这里报告的高分辨率晶体学分析Lck SH 2结构域与此磷酸肽的复合物。最近，Src SH 2结构域与低亲和力肽复合的晶体学衍生结构18（不含EEI共有序列），以及未配体的Abl（参考文献19）和p85（参考文献20）SH 2结构域的NMR衍生结构揭示了SH 2结构域的保守折叠和磷酸酪氨酸结合口袋的特性。我们的高亲和力复合物显示存在第二个口袋的残基（pY + 3）的三个位置的磷酸酪氨酸（pY）的C-末端。通过将pY和pY + 3侧链插入其口袋中并通过氢键网络与肽主链锚定肽。在低亲和力磷酸肽/Src复合物18中，pY + 3残基不插入同源结合口袋中，肽主链保持从结构域表面置换。
THE Src homology-2 (SH2) domains are modules of about 100 amino-acid residues that are found in many intracellular signal-transduction proteins1,2. They bind phosphotyrosine-containing sequences with high affinity and specificity1-4, recognizing phosphotyrosine in the context of the immediately adjacent polypeptide sequence3-12. The protein p56lck (Lck) is a Src-like, lymphocyte-specific tyrosine kinase13-15. A phosphopeptide library screen has recently been used to deduce an 'optimal' binding sequence for the Lck SH2 domain16. There is selectivity for the residues Glu, Glu and Ile in the three positions C-terminal to the phosphotyrosine. An 11-residue phosphopeptide derived from the hamster polyoma middle-T antigen, EPQpYEEIPIYL, binds with an approximately 1 nM dissociation constant to the Lck SH2 (ref. 17), an affinity equivalent to that of the tightest known SH2-phosphopeptide complex. We report here the high-resolution crystallographic analysis of the Lck SH2 domain in complex with this phosphopeptide. Recent crystallographically derived structures of the Src SH2 domain in complex with low-affinity peptides18, which do not contain the EEI consensus, and NMR-derived structures of unliganded Abl (ref. 19) and p85 (ref. 20) SH2 domains have revealed the conserved fold of the SH2 domain and the properties of a phosphotyrosine binding pocket. Our high-affinity complex shows the presence of a second pocket for the residue (pY + 3) three positions C-terminal to the phosphotyrosine (pY). The peptide is anchored by insertion of the pY and pY + 3 side chains into their pockets and by a network of hydrogen bonds to the peptide main chain. In the low-affinity phosphopeptide/Src complexes18, the pY + 3 residues do not insert into the homologous binding pocket and the peptide main chain remains displaced from the surface of the domain.