PT  - JOURNAL ARTICLE
AU  - Nestor O Concha
AU  - Angela Smallwood
AU  - William Bonnette
AU  - Rachel Totoritis
AU  - Guofeng Zhang
AU  - Kelly Federowicz
AU  - Jingsong Yang
AU  - Hongwei Qi
AU  - Stephanie Chen
AU  - Nino Campobasso
AU  - Anthony E Choudhry
AU  - Leanna E Shuster
AU  - Karen A Evans
AU  - Jeff Ralph
AU  - Sharon Sweitzer
AU  - Dirk A Heerding
AU  - Carolyn A Buser
AU  - Dai-shi Su
AU  - Maurice P DeYoung
TI  - Long-Range Inhibitor-Induced Conformational Regulation of Human IRE1α Endoribonuclease Activity
AID  - 10.1124/mol.115.100917
DP  - 2015 Jan 01
TA  - Molecular Pharmacology
PG  - mol.115.100917
4099  - http://molpharm.aspetjournals.org/content/early/2015/10/05/mol.115.100917.short
4100  - http://molpharm.aspetjournals.org/content/early/2015/10/05/mol.115.100917.full
AB  - Activation of the inositol-requiring enzyme-1 alpha (IRE1α) protein caused by endoplasmic reticulum (ER) stress results in the homodimerization of the N-terminal ER luminal domains, autophosphorylation of the cytoplasmic kinase domains, and conformational changes to the cytoplasmic endoribonuclease (RNase) domains which render them functional and can lead to the splicing of XBP1 mRNA. Herein we report the first crystal structures of the cytoplasmic portion of a human phosphorylated IRE1α dimer in complex with GSK2850163, a novel, IRE1α-selective kinase inhibitor, and with staurosporine (STS), a broad spectrum kinase inhibitor. GSK2850163 inhibits both the kinase and RNAse activities of IRE1α. The inhibitor interacts with the catalytic residues Lys599 and Glu612 and displaces the kinase activation loop to the DFG-out conformation. Inactivation of IRE1α RNAse activity appears to be caused by a conformational change whereby the αC helix is displaced resulting in the rearrangement of the kinase domain-dimer interface and a rotation of the RNAse domains away from each other. In contrast, STS binds at the ATP-binding site of IRE1α resulting in a dimer consistent with RNase active yeast Ire1 dimers. Activation of IRE1α RNAse activity appears to be promoted by a network of hydrogen bond interactions between highly conserved residues across the RNAse dimer interface that place key catalytic residues poised for reaction. These data implicate that the intermolecular interactions between conserved residues in the RNase domain are required for activity, and that the disruption of these interactions can be achieved pharmacologically by small molecule kinase domain inhibitors.