Structure-Functional Basis of Ion Transport in Sodium–Calcium Exchanger (NCX) Proteins

Abstract

The membrane-bound sodium–calcium exchanger (NCX) proteins shape Ca2+ homeostasis in many cell types, thus participating in a wide range of physiological and pathological processes. Determination of the crystal structure of an archaeal NCX (NCX_Mj) paved the way for a thorough and systematic investigation of ion transport mechanisms in NCX proteins. Here, we review the data gathered from the X-ray crystallography, molecular dynamics simulations, hydrogen–deuterium exchange mass-spectrometry (HDX-MS), and ion-flux analyses of mutants. Strikingly, the apo NCX_Mj protein exhibits characteristic patterns in the local backbone dynamics at particular helix segments, thereby possessing characteristic HDX profiles, suggesting structure-dynamic preorganization (geometric arrangements of catalytic residues before the transition state) of conserved α1 and α2 repeats at ion-coordinating residues involved in transport activities. Moreover, dynamic preorganization of local structural entities in the apo protein predefines the status of ion-occlusion and transition states, even though Na+ or Ca2+ binding modifies the preceding backbone dynamics nearby functionally important residues. Future challenges include resolving the structural-dynamic determinants governing the ion selectivity, functional asymmetry and ion-induced alternating access. Taking into account the structural similarities of NCX_Mj with the other proteins belonging to the Ca2+/cation exchanger superfamily, the recent findings can significantly improve our understanding of ion transport mechanisms in NCX and similar proteins.

Hydrogen-Deuterium Exchange Mass-Spectrometry of Secondary Active Transporters: From Structural Dynamics to Molecular Mechanisms

“…Surprisingly, the crystal structure of NCX from Methanocaldococcus jannaschii (NCX_Mj) revealed four ion binding sites, simultaneously occupied by three Na + ions at sites termed S int , S mid , S ext , and one Ca 2+ ion at a site termed S Ca (Liao et al, 2012). Since this binding mode was inconsistent with previous studies, MD simulations and ion flux analyses of mutants were performed, suggesting that Na + ions occupy S int , S Ca , and S ext , whereas Ca 2+ occupies S Ca (Marinelli et al, 2014;Giladi et al, 2016b;Giladi et al, 2017;van Dijk et al, 2018). Thus, the Na + and Ca 2+ ions are bound in a mutually exclusive manner along the transport cycle.…”

Crosslink between calcium and sodium signalling

“…Moreover, this dynamic coupling of NCX with Na + and Ca 2+ signalling requires versatile regulatory responses of NCX to environmental changes in order to fulfil spatiotemporal requirements of coupled systems acting in functionally diverse cell types. A substantial progress has been achieved during the last years in understanding the structure-dynamic determinants underlying regulatory mechanisms belonging to NCX isoform/splice variants (Giladi et al , 2016a; Giladi et al , 2016b; Khananshvili, 2016). Below, we shortly summarize most important features of allosteric regulation of NCX variants and indicate possible contributions of specific NCX variants in Na + and Ca 2+ signalling.…”

“…In contrast, in glia the NCX1 and NCX3 variants undergo Na + -induced inactivation and only Ca 2+ binding to CBD2 in specific variants (NCX1-AD, NCX3-B or NCX3-BC) can alleviate the activity. Since NCX2 is insensitive to regulatory Na + , this isoform (forming no splice variants) is capable of retaining its activity in glia at high-amplitude and prolonged Na + -transients (Philipson & Nicoll, 2000; Lytton, 2007; Giladi et al , 2012; Giladi et al , 2015; Giladi et al , 2016a; Giladi et al , 2016b; Khananshvili, 2016; Tal et al , 2016). …”

Bond graph modelling of the cardiac action potential: implications for drift and non-unique steady states

“…The NCX was modelled using the bond graph shown in Figure S6. The reaction scheme was based on the ping-pong mechanism proposed in Giladi et al [62], with reactions r1, r2, r4 and r5 modelled by fast rate constants to approximate rapid equilibrium. We assigned voltage dependence to translocation of Na + , based on experimental findings from Hilgemann et al [63].…”