Atomistic Molecular Dynamics Simulations of Drosophila Orai in a Hydrated Lipid Bilayer

Abstract

Orai channels are prominent (calcium) Ca2+ signal mediators in many cell types. In T lymphocytes, Orai1 and STIM1 form the Ca2+ release-activated Ca2+ (CRAC) channel. Following endoplasmic reticulum (ER) Ca2+ store depletion, ER membrane-resident Ca2+ sensor STIM1 physically interacts with plasma membrane-bound, pore-forming Orai1 subunits. The STIM1-Orai1 interaction results in Ca2+ influx and ultimately changes in gene expression that underlie the immune response. In 2012, the Drosophila Orai (dOrai) crystal structure was solved to 3.34 A resolution. To develop an atomistic view of Orai channel function, given the high sequence identity (73%) between dOrai and human Orai1, we constructed a complete dOrai structure by building in flexible loops missing in the crystal structure and adjusting the protonation state of several pore residues. The model was placed into a lipid bilayer with excess hydration and simulated for over 200 ns. We find shifted pKa values for several titratable residues in the pore interior, and that changing protonation states of these key residues is essential to maintain pore stability. To explore the mechanism of channel block, we also simulated the model with either Ca2+ or (gadolinium) Gd3+ in the selectivity filter. Taken together, our results help to address the stability of the closed structure in a lipid bilayer while also yielding novel atomistic insights into channel block and pH sensitivity.This work was supported by NIH grants F30CA171717 to M.L.W., P01GM86685 to D.J.T., and NS-14609 to M.D.C.1. Hou, X, et al. Science. (2012); 338; 1308-13.