Magnesium is the most abundant divalent cation present within cells, playing an integral role in many biological processes, including protein synthesis, DNA replication, and ATP synthesis and is an essential cofactor for at least 600 enzymes. Maintenance of magnesium homeostasis is vital to normal cellular function and is tightly regulated, with dysregulation in humans linked to numerous diseases. Within the mitochondria, magnesium homeostasis has been closely linked to normal mitochondrial function with the magnesium channel MRS2, localized to the inner mitochondrial membrane, importing magnesium. The bacterial homolog, CorA, has been extensively investigated and structural studies suggest that a soluble magnesium-sensor domain is integral to the transition between open and closed states. We recently resolved the human MRS2 structure in its closed state, but despite the presence of a homologous soluble, magnesium sensor domain in MRS2, the open state structure remains elusive. Furthermore, wild-type MRS2 in vitro activity assays are unable to reconstitute an active channel, with only mammalian cell-based assays showing magnesium transport. This suggests a unique mechanism for regulation of MRS2 magnesium transport in humans. To elucidate the mechanism of human MRS2 channel regulation, channel opening, and the role of the soluble domain, homologous to the magnesium sensor in bacterial CorA, we undertook structure-function studies. Our functional studies combine mutagenesis with a magnesium transport assay in a magnesium auxotrophic E. coli strain to identify residues integral to activity in MRS2. Our structural studies utilize polymer solubilization to elucidate the putative open state structure. Intriguingly, reconstitution of MRS2 into nanodiscs with similar treatment was unable to resolve the open state, highlighting the importance of the inner mitochondrial membrane toward MRS2 dynamics, which we probed via molecular dynamics simulations. These studies provide new insight into the mechanisms of MRS2 activity regulation.
Clark et al. (Sun,) studied this question.