Feilen Lukas P, Haubrich Kevin, Strecker Paul, Probst Sabine, Eggert Simone, Stier Gunter, Sinning Irmgard, Konietzko Uwe, Kins Stefan, Simon Bernd, Wild Klemens
Heidelberg University Biochemistry Center (BZH), University of HeidelbergHeidelberg, Germany.
European Molecular Biology Laboratory (EMBL), Structural and Computational BiologyHeidelberg, Germany.
Front Mol Neurosci. 2017 May 11;10:140. doi: 10.3389/fnmol.2017.00140. eCollection 2017.
Physiological function and pathology of the Alzheimer's disease causing amyloid precursor protein (APP) are correlated with its cytosolic adaptor Fe65 encompassing a WW and two phosphotyrosine-binding domains (PTBs). The C-terminal Fe65-PTB2 binds a large portion of the APP intracellular domain (AICD) including the GYENPTY internalization sequence fingerprint. AICD binding to Fe65-PTB2 opens an intra-molecular interaction causing a structural change and altering Fe65 activity. Here we show that in the absence of the AICD, Fe65-PTB2 forms a homodimer in solution and determine its crystal structure at 2.6 Å resolution. Dimerization involves the unwinding of a C-terminal α-helix that mimics binding of the AICD internalization sequence, thus shielding the hydrophobic binding pocket. Specific dimer formation is validated by nuclear magnetic resonance (NMR) techniques and cell-based analyses reveal that Fe65-PTB2 together with the WW domain are necessary and sufficient for dimerization. Together, our data demonstrate that Fe65 dimerizes via its APP interaction site, suggesting that besides intra- also intermolecular interactions between Fe65 molecules contribute to homeostatic regulation of APP mediated signaling.
引发阿尔茨海默病的淀粉样前体蛋白（APP）的生理功能和病理学与其包含一个WW结构域和两个磷酸酪氨酸结合结构域（PTB）的胞质衔接蛋白Fe65相关。Fe65的C末端PTB2结合了大部分APP细胞内结构域（AICD），包括GYENPTY内化序列指纹。AICD与Fe65-PTB2的结合开启了分子内相互作用，导致结构变化并改变Fe65活性。在此我们表明，在没有AICD的情况下，Fe65-PTB2在溶液中形成同源二聚体，并以2.6 Å的分辨率确定了其晶体结构。二聚化涉及C末端α螺旋的解旋，该螺旋模拟了AICD内化序列的结合，从而屏蔽了疏水结合口袋。通过核磁共振（NMR）技术验证了特异性二聚体的形成，基于细胞的分析表明，Fe65-PTB2与WW结构域一起对于二聚化是必要且充分的。总之，我们的数据表明Fe65通过其APP相互作用位点二聚化，这表明除了Fe65分子内相互作用外，分子间相互作用也有助于APP介导信号传导的稳态调节。
