Pore Architecture and Ion Selectivity Filter of the Mitochondrial Calcium Uniporter

Abstract

The calcium uniporter of mitochondria is a holocomplex consisting of the calcium-conducting channel, known as the MCU channel, and a score of regulatory proteins. Previous electrophysiology study found that a preeminent property of the uniporter is the high calcium selectivity and conductance and this has been shown to critically depend on the highly conserved amino acid sequence motif, DXXE, in the MCU channel. Our recent NMR data of the MCU channel indicate that the DXXE forms two parallel carboxylate rings at the channel entrance that appear to serve as the ion selectivity filter. We used paramagnetic probes to show that mutants with a single carboxylate ring can bind divalent cation specifically, while in the wild type, the two rings bind the ion cooperatively, resulting in drastically higher apparent affinity. Our new data, together with structural analysis of the DXXE motif, indicate that the double carboxylate rings at the apex of the MCU pore constitute the ion selectivity filter of the channel and that Ru360 directly blocks ion entry into the filter.