Abstract
Single-particle cryo-EM was used to explore the structure and conformational landscape of ∼200-kDa magnesium ion channel CorA. CorA is the major Mg2+-uptake system in prokaryotes and can functionally complement the eukaryotic mitochondrial Mrs2 channel, which is essential for normal mitochondrial activity, and whose abnormal expression has been associated with diseases ranging from cancer to demyelination of neuronal tissue. CorA is a homo-pentamer and is gated by intracellular Mg2+. X-ray crystallographic studies of CorA show similar, rather symmetric conformations under Mg2+-bound and Mg2+-free conditions, but EPR spectroscopic studies reveal large Mg2+-driven quaternary conformational changes. Here, we determined the cryo-EM structures of CorA in the presence and absence of Mg2+. The five-fold symmetric Mg2+-bound “closed” state of detergent-solubilized CorA at an overall 3.8-A resolution reveals side-chain densities as well as densities for several magnesium ions, and is similar to known X-ray structures with only minor differences in some of the transmembrane segments, the C-terminal domain, and the periplasmic loops. Cryo-EM of Mg2+-free “open” state CorA shows dramatic differences in the conformation of the channel with large asymmetric changes in the soluble domain of the pentamer. In the absence of bound Mg2+, four of the five subunits are displaced ∼10 to ∼25 A by hinge-like motions at the stalk helix with a bending of up to 35°. Extensive 3D-classification during data-processing resulted in at least two distinct “open” state CorA structures at ∼7-A. The asymmetric structures were supported by single-particle cryo-EM of CorA in lipid nanodiscs and other biochemical and biophysical studies. Our findings suggest an unprecedented gating mechanism where the reversible loss of five-fold symmetry is the key structural signature of the transition between a single “closed” state and an ensemble of low-Mg2+, “open” conformational states.
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