Functional asymmetry of bidirectional Ca2+-movements in an archaeal sodium–calcium exchanger (NCX_Mj)

Abstract Dynamic features of Ca 2+ interactions with transport and regulatory sites control the Ca 2+ -fluxes in mammalian Na + /Ca 2+ (NCX) exchangers bearing the Ca 2+ -binding regulatory domains on the cytosolic 5L6 loop. The crystal structure of Methanococcus jannaschii NCX (NCX_Mj) may serve as a template for studying ion-transport mechanisms since NCX_Mj does not contain the regulatory domains. The turnover rate of Na + /Ca 2+ exchange ( k cat  = 0.5 ± 0.2s −1 ) in WT–NCX_Mj is 10 3 –10 4 times slower than in mammalian NCX. In NCX_Mj, the intrinsic equilibrium ( K int ) for bidirectional Ca 2+ movements (defined as the ratio between the cytosolic and extracellular K m of Ca 2+ /Ca 2+ exchange) is asymmetric, K int  = 0.15 ± 0.5. Therefore, the Ca 2+ movement from the cytosol to the extracellular side is ∼7-times faster than in the opposite direction, thereby representing a stabilization of outward-facing (extracellular) access. This intrinsic asymmetry accounts for observed differences in the cytosolic and extracellulr K m values having a physiological relevance. Bidirectional Ca 2+ movements are also asymmetric in mammalian NCX. Thus, the stabilization of the outward-facing access along the transport cycle is a common feature among NCX orthologs despite huge differences in the ion-transport kinetics. Elongation of the cytosolic 5L6 loop in NCX_Mj by 8 or 14 residues accelerates the ion transport rates ( k cat ) ∼10 fold, while increasing the K int values 100–250-fold ( K int  = 15–35). Therefore, 5L6 controls both the intrinsic equilibrium and rates of bidirectional Ca 2+ movements in NCX proteins. Some additional structural elements may shape the kinetic variances among phylogenetically distant NCX variants, although the intrinsic asymmetry ( K int ) of bidirectional Ca 2+ movements seems to be comparable among evolutionary diverged NCX variants.