Mg²⁺-dependent conformational equilibria in CorA: an integrated view on transport regulation

Abstract

The CorA family of proteins regulates the homeostasis of divalent metal ions in many bacteria, archaea, and eukaryotic mitochondria, making it an important target in the investigation of the mechanisms of transport and its functional regulation. Although numerous structures of open and closed channels are now available for the CorA family, the mechanism of the transport regulation remains elusive. Here, we investigated the conformational distribution and associated dynamic behaviour of the pentameric Mg2+ channel CorA at room temperature using small-angle neutron scattering (SANS) in combination with molecular dynamics (MD) simulations and solid-state nuclear magnetic resonance spectroscopy (NMR). We find that neither the Mg2+-bound closed structure nor the Mg2+-free open forms are sufficient to explain the average conformation of CorA. Our data support the presence of conformational equilibria between multiple states, and we further find a variation in the behaviour of the backbone dynamics with and without Mg2+. We propose that CorA must be in a dynamic equilibrium between different non-conducting states, both symmetric and asymmetric, regardless of bound Mg2+ but that conducting states become more populated in Mg2+-free conditions. These properties are regulated by backbone dynamics and are key to understanding the functional regulation of CorA.

The human MRS2 magnesium binding domain is a regulatory feedback switch for channel activity

“…Mg 2+ increased α-helicity and stability of the human MRS2 NTD, consistent with past NMR data showing decreased backbone dynamics of CorA in the presence of high Mg 2+ (Johansen et al, 2022). The far-UV CD spectra reported here resemble previous data from our lab, where we found no effect by MgCl 2 , likely due to variability in protein concentration measurements (Daw et al, 2020).…”

“…Several CorA crystal and cryoelectron microscopy (cryoEM) structures have been elucidated in the presence of divalent cations, revealing a pentameric assembly (Cleverley et al , 2015; Eshaghi et al ., 2006; Guskov et al ., 2012; Johansen et al ., 2022; Matthies et al ., 2016; Nordin et al , 2013; Payandeh & Pai, 2006; Pfoh et al ., 2012). The first TM, which lines the channel pore, and second TM, position the intervening GMN motif for ion binding and selectivity at the pore entrance (Pfoh et al ., 2012).…”

“…M1 is made up of D89 and D253, while M2 is comprised of D175 and D179. Whereas earlier studies indicated that a symmetrizing of the pentameric intracellular domain assembly upon Mg 2+ binding to the NTD closes the channel (Matthies et al ., 2016; Pfoh et al ., 2012), more recent work indicates both symmetric and asymmetric assemblies are formed in the presence and absence of Mg 2+ , and channel conductance is dependent on lowered symmetric state population and coupled with a reduced energy barrier to an ensemble of open states in low Mg 2+ (Johansen et al ., 2022; Kowatz & Maguire, 2019). Nevertheless, it is evident that Mg 2+ binding increases the rigidity/decreases the dynamics of the CorA intracellular domain (Chakrabarti et al , 2010; Johansen et al ., 2022; Pfoh et al ., 2012; Rangl et al , 2019).…”

“…Although CorA and Mrs2, orthologues of MRS2 in bacteria and yeast, respectively, have been structurally resolved at high resolution (Eshaghi et al , 2006; Guskov et al , 2012; Johansen et al , 2022; Khan et al , 2013; Lunin et al , 2006; Matthies et al , 2016; Payandeh & Pai, 2006; Pfoh et al , 2012), low sequence similarity exists between CorA, Mrs2 and MRS2, especially in the NTDs (Zsurka et al , 2001). Specifically, the sequence similarity between human MRS2 and yeast ( Saccharomyces cerevisiae ) Mrs2 is 55.4% (and only 20.1% through the NTD), while the sequence similarity between human MRS2 and bacterial ( Thermotoga maritima ) CorA is 43.3% (and only 17.0% through the NTD) ( Fig.…”