Ligand specificity modulated by prolyl imide bond cis/trans isomerization in the Itk SH2 domain: A quantitative NMR study
Ligand specificity modulated by prolyl imide bond cis/trans isomerization in the Itk SH2 domain: A quantitative NMR study
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DOI：
10.1021/ja0375380
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发表时间：
2003-12-24
影响因子：
15
通讯作者：
Andreotti, AH
中科院分区：
文献类型：
--
作者：
Breheny, PJ;Laederach, A;Andreotti, AH
白细胞介素-2酪氨酸激酶（Itk）的Src同源2（SH 2）结构域结合两个单独的配体：含磷酸酪氨酸的肽和Itk Src同源3（SH 3）结构域。这些配体的结合特异性通过Itk SH 2结构域中Asn 286−Pro 287酰亚胺键的顺式/反式异构化来调节。在这项研究中，我们开发了一种新的方法，分析化学位移扰动和交叉峰体积来测量每个SH 2构象的两个配体的亲和力。我们发现，顺式酰亚胺键含SH 2构象显示出3.5倍更高的亲和力的Itk SH 3结构域相比，反式构象的结合相同的配体，而反式构象结合磷酸肽的4倍更大的亲和力比顺式含SH 2构象。除了进一步了解这个系统，这里提出的方法将是一般的应用在定量确定构象异构系统，使用分子开关来调节多个不同的配体之间的结合的特异性。
The Src homology 2 (SH2) domain of interleukin-2 tyrosine kinase (Itk) binds two separate ligands: a phosphotyrosine-containing peptide and the Itk Src homology 3 (SH3) domain. Binding specificity for these ligands is regulated via cis/trans isomerization of the Asn 286−Pro 287 imide bond in the Itk SH2 domain. In this study, we develop a novel method of analyzing chemical shift perturbation and cross-peak volumes to measure the affinities of both ligands for each SH2 conformer. We find that the cis imide bond containing SH2 conformer exhibits a 3.5-fold higher affinity for the Itk SH3 domain compared with binding of the trans conformer to the same ligand, while the trans conformer binds phosphopeptide with a 4-fold greater affinity than the cis-containing SH2 conformer. In addition to furthering the understanding of this system, the method presented here will be of general application in quantitatively determining the specificities of conformationally heterogeneous systems that use a molecular switch to regulate binding between multiple distinct ligands.