Joshi Amar, Newbatt Yvette, McAndrew P Craig, Stubbs Mark, Burke Rosemary, Richards Mark W, Bhatia Chitra, Caldwell John J, McHardy Tatiana, Collins Ian, Bayliss Richard
Department of Biochemistry, University of Leicester, Leicester, United Kingdom.
Cancer Research UK Leicester Centre, University of Leicester, Leicester, United Kingdom.
Oncotarget. 2015 May 30;6(15):13019-35. doi: 10.18632/oncotarget.3864.
IRE1 transduces the unfolded protein response by splicing XBP1 through its C-terminal cytoplasmic kinase-RNase region. IRE1 autophosphorylation is coupled to RNase activity through formation of a back-to-back dimer, although the conservation of the underlying molecular mechanism is not clear from existing structures. We have crystallized human IRE1 in a back-to-back conformation only previously seen for the yeast homologue. In our structure the kinase domain appears primed for catalysis but the RNase domains are disengaged. Structure-function analysis reveals that IRE1 is autoinhibited through a Tyr-down mechanism related to that found in the unrelated Ser/Thr protein kinase Nek7. We have developed a compound that potently inhibits human IRE1 kinase activity while stimulating XBP1 splicing. A crystal structure of the inhibitor bound to IRE1 shows an increased ordering of the kinase activation loop. The structures of hIRE in apo and ligand-bound forms are consistent with a previously proposed model of IRE1 regulation in which formation of a back-to-back dimer coupled to adoption of a kinase-active conformation drive RNase activation. The structures provide opportunities for structure-guided design of IRE1 inhibitors.
肌醇需求酶1（IRE1）通过其C末端胞质激酶-核糖核酸酶区域对XBP1进行剪接，从而传导未折叠蛋白反应。IRE1的自磷酸化通过形成背对背二聚体与核糖核酸酶活性偶联，尽管从现有结构中尚不清楚潜在分子机制的保守性。我们已使人类IRE1以仅在酵母同源物中才出现过的背对背构象结晶。在我们的结构中，激酶结构域似乎已准备好进行催化，但核糖核酸酶结构域是分离的。结构-功能分析表明，IRE1通过一种与不相关的丝氨酸/苏氨酸蛋白激酶Nek7中发现的机制相关的酪氨酸向下机制进行自我抑制。我们开发了一种化合物，它能有效抑制人类IRE1激酶活性，同时刺激XBP1剪接。与IRE1结合的抑制剂的晶体结构显示激酶激活环的有序性增加。无配体和配体结合形式的hIRE结构与先前提出的IRE1调节模型一致，在该模型中，背对背二聚体的形成与激酶活性构象的采用驱动核糖核酸酶激活。这些结构为IRE1抑制剂的结构导向设计提供了机会。
