The interaction of Jagged-1 cytoplasmic tail with afadin PDZ domain is local, folding-independent, and tuned by phosphorylation.
The interaction of Jagged-1 cytoplasmic tail with afadin PDZ domain is local, folding-independent, and tuned by phosphorylation.
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DOI：
10.1002/jmr.1042
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发表时间：
2011-03-01
期刊：
影响因子：
--
通讯作者：
Pongor, Sandor
中科院分区：
文献类型：
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作者：
Popovic, Matija;Bella, Juraj;Pongor, Sandor
jagded -1是人类5种Notch配体之一，是一种跨膜蛋白，由一个大的细胞外区域和一个带有c端PDZ识别基元(（1213)RMEYIV(1218)）的125个残基细胞质尾部组成。jagded -1细胞内区域与afadin（一种位于细胞-细胞粘附连接处的蛋白质）的PDZ结构域结合，将Notch信号与粘附系统和细胞骨架结合。利用核磁共振化学位移摄动和表面等离子体共振，研究了afadin的PDZ结构域（AF6_PDZ）与一系列包含PDZ结合基序的多肽之间的相互作用。AF6_PDZ与不同长度配体（6,24和133个残基）结合时的化学位移映射表明，相互作用是严格局部的，只涉及PDZ中的结合槽。Jagged-1的整个胞内区对应的重组蛋白J1_ic在溶液中主要是无序的，并且在AF6_PDZ存在下J1_ic的化学位移图谱显示结合不偶联折叠。对一系列不同位置磷酸化的24个残基肽的结合研究表明，pdz结合基序-2位置酪氨酸的磷酸化降低了其对AF6_PDZ的亲和力，并可能在这种相互作用的调节中发挥作用。最后，我们发现位于pdz结合基元的R1213Q突变与肝外胆道闭锁相关，增加了对AF6_PDZ的亲和力，这表明这种综合征可能是由于Notch信号与细胞骨架耦合的不平衡引起的。
Jagged-1, one of the five Notch ligands in man, is a membrane-spanning protein made of a large extracellular region and a 125-residue cytoplasmic tail bearing a C-terminal PDZ recognition motif ((1213) RMEYIV(1218) ). Binding of Jagged-1 intracellular region to the PDZ domain of afadin, a protein located at cell-cell adherens junctions, couples Notch signaling with the adhesion system and the cytoskeleton. Using NMR chemical shift perturbation and surface plasmon resonance, we studied the interaction between the PDZ domain of afadin (AF6_PDZ) and a series of polypeptides comprising the PDZ-binding motif. Chemical shift mapping of AF6_PDZ upon binding of ligands of different length (6, 24, and 133 residues) showed that the interaction is strictly local and involves only the binding groove in the PDZ. The recombinant protein corresponding to the entire intracellular region of Jagged-1, J1_ic, is mainly disordered in solution, and chemical shift mapping of J1_ic in the presence of AF6_PDZ showed that binding is not coupled to folding. Binding studies on a series of 24-residue peptides phosphorylated at different positions showed that phosphorylation of the tyrosine at position -2 of the PDZ-binding motif decreases its affinity for AF6_PDZ, and may play a role in the modulation of this interaction. Finally, we show that the R1213Q mutation located in the PDZ-binding motif and associated with extrahepatic biliary atresia increases the affinity for AF6_PDZ, suggesting that this syndrome may arise from an imbalance in the coupling of Notch signaling to the cytoskeleton.