The Inactivation Gates of BsYetJ Calcium Channel Revealed in the Endoplasmic Reticulum Environment

Abstract

The transmembrane BAX inhibitor-1-containing motif 6 (TMBIM6) is an anti-apoptotic endoplasmic reticulum (ER) membrane protein. The suppression of apoptosis by TMBIM6 is linked to its role in regulating calcium homeostasis. However, the molecular mechanisms underlying the calcium regulation in the ER remain poorly understood. Using mutagenesis and fluorescence-based functional assays, this study investigates all the highly conserved negatively charged residues in the BsYetJ (a bacterial homolog of TMBIM6) and reveals that the charged residues E49 and R205 cooperatively act as a major calcium-selectivity filter determining the channel conductance. This study identifies the conformations of BsYetJ in the ER environment using electron spin resonance and molecular dynamics. Together with the functional data, we propose a mechanism, composed of two selectivity filters, for the calcium-dependent inactivation of BsYetJ observed in the ER environment. Overall, this study provides a framework for understanding the non-canonical calcium gated membrane protein, which will facilitate structure-based development of more effective medications for TMBIM6.