Putative CaCA family member of 368 aas and 10 TMSs

Amoebozoa (Slime molds)

UP of Dictyostelium discoideum

Uncharacterized protein of 518 aas and 10 TMSs

Animals

UP of Trichoplax adhaerens (Trichoplax reptans)

The Ca²⁺:H⁺ antiporter, YfkE; homotrimer with subunit size of 451 aas.  The 3-d x-ray strcuture is known to 3.1 Å resolution (Wu et al. 2013).  The conformational transition is triggered by the rotation of the kink angles of transmembrane helices 2 and 7 and is mediated by large conformational changes in their adjacent transmembrane helices 1 and 6.  The inward facing conformation contrasts with the outward facing conformation demonstrated for NCX_Mj (TC# 2.A.19.5.3).  The inward facing conformation has a "hydrophobic seal" that closes the external exit.

Firmicutes

YfkE of Bacillus subtilis

Ca²⁺/Mg²⁺/Mn²⁺:H⁺ antiporter, CAX or CHA, of 441 aas with a 90 residue hydrophilic N-terminus followed by 11 TMSs (Wunderlich 2022).

CAX of Plasmodium falciparum

Vacuolar [Mn²⁺ or Ca²⁺]:H⁺ antiporter, Hum1 (Mn²⁺ resistance (Mnr1)) protein. Vcx1 has 11 probable TMSs with the N-terminus inside (Segarra and Thomas, 2008).  The 3-d structure has been determined at 2.3 Å resolution for the cytosolic facing, substrate bound form, favoring the alternating access mechanism of transport (Waight et al. 2013).

Yeast

Hum1 (Mnr1) of Saccharomyces cerevisiae

High affinity vacuolar (tonoplast) Ca²⁺:H⁺ antiporter (also exports Cd²⁺ and Zn²⁺; Shigaki et al., 2005) It is expressed in leaves (Cheng et al., 2005). It determines sensitivity to abscisic acid and sugars during germination and tolerance to ethylene during early seedling development (Zhao et al., 2008). BrCAX1 is involved in Ca²⁺ transport and Ca²⁺ deficiency-induced tip-burn in chinese cabbage (Brassica rapa L. ssp. pekinensis) (Cui et al. 2023).

Plants

Cax1 of Arabidopsis thaliana

Low affinity Ca²⁺:H⁺/heavy metal cation (e.g., Mn²⁺, Mg²⁺, Cd²⁺, Ca²⁺):H⁺ antiporter, Cax2.  The apple (Malus domestiga) ortholog (CAX2L-2) functions positively in modulation of Ba²⁺ tolerance (Mei et al. 2023).

Plants

Cax2 of Arabidopsis thaliana

Algae Ca²⁺: H⁺ and Na⁺:H⁺ exchanger, CAX1 (mediates stress responses to high Ca²⁺, Na⁺ and Co²⁺).

Algae

CAX1 of Chlamydomonas reinhardtii (B6ZCF4)

Ca²⁺/H⁺ antiporter, YfkE (Fujisawa et al., 2009). YfkE is a homotrimer with a subunit size of 451 aas.  The 3-d x-ray strcuture is known to 3.1 Å resolution (Wu et al. 2013).  The conformational transition is triggered by the rotation of the kink angles of transmembrane helices 2 and 7 and is mediated by large conformational changes in their adjacent transmembrane helices 1 and 6.  The inward facing conformation contrasts with the outward facing conformation demonstrated for NCX_Mj (TC# 2.A.19.5.3).  The inward facing conformation has a "hydrophobic seal" that closes the external exit (Wu et al. 2013). Intracellular Ca²⁺ regulation of this H⁺/Ca²⁺ antiporter is mediated by a Ca²⁺ mini-sensor (Lu et al. 2020).

Bacteria

YfkE of Bacillus subtilis (O34840)

The vacuolar Ca²⁺:H⁺ exchanger, CAX (Bowman et al., 2011).

Fungi

CAX of Neurospora crassa (O59940)

Vacuolar cation:proton exchanger, Cax4 (transports Cd²⁺>Zn²⁺>Ca²⁺>Mn²⁺) (Cheng et al., 2002; Mei et al., 2009).  The rice orthologue, Cax4, may transport Ca²⁺, Mn²⁺ and Cu²⁺, and functions in salt stress (Yamada et al. 2014). The A. thaliana protein contributes to Cd²⁺ resistance (Liao et al. 2019).

Plants

Cax4 of Arabidopsis thaliana (Q945S5)

The 10 TMS cardiac Ca²⁺:3 Na⁺ antiporter, NCX1 (Ren and Philipson 2013).  The Ca²⁺ sensor (residues 371-508) binds cytoplasmic Ca²⁺ allosterically to activate exchange activity) (Nicoll et al., 2006; Ren et al., 2006) NCX1 forms homodimers (Ren et al., 2008). It is present in mitochondria where it catalyzes Ca²⁺ efflux. TMS packing has been analyzed by Ren et al. (2010). Cytoplasmic Ca²⁺ regulates the dimeric NCX by binding to two adjacent Ca²⁺-binding domains (CBD1 and CBD2) located in the large intracellular loop between transmembrane segments 5 and 6. John et al. (2011) showed that Ca²⁺decreases the distance between the cytoplasmic loops of NCX pairs, thereby activating transport.  Ser110 in TMS2 plays a role in both Na⁺ and Ca²⁺ transport (Ottolia and Philipson 2013). nimodipiine-sensitive NCX1, as well as mitochondrial Ca²⁺ uptake, plays an important role in clearing somatic Ca²⁺ after depolarization-induced Ca²⁺ influx in SCN neurons (Wang et al. 2015). Regulation in suprachiasmatic nucleus neurons has been studied (Cheng et al. 2018).  Genetic knockout and pharmacologic inhibitors of NCX1 attenuate hypoxia-induced pulmonary arterial hypertension (Nagata et al. 2020). The main Na⁺ influx pathway in myocardia in neonates is the NCX transporter (Oshiyama et al. 2022).  Cytosolic Ca²⁺ and Na⁺ allosterically regulate Na⁺/Ca²⁺ exchanger (NCX) proteins to vary the NCX-mediated Ca²⁺ entry/exit rates in diverse mammalian cell types (Giladi et al. 2024).

Animals

Ca²⁺ regulated Ca²⁺:Na⁺ antiporter (NCX1) of Bos taurus

Probable Ca²⁺:3Na⁺ antiporter, Calx (contains two repeat motifs Calx-α and Calxβ, between the two transmembrane domains, as is true of many Ca²⁺:Na⁺ antiporters (Schwarz and Benzer, 1997). CALX activity is inhibited by Ca²⁺ interaction within its two intracellular Ca²⁺ regulatory domains CBD1 and CBD2. The Ca²⁺ inhibition of CALX is achieved by interdomain conformational changes induced by Ca²⁺ binding at CBD1 (Wu et al., 2011). The exchanger is an essential Ca²⁺ extrusion mechanism in excitable cells. It consists of a transmembrane domain and a large intracellular loop that contains the two Ca²⁺-binding domains, CBD1 and CBD2 (de Souza Degenhardt et al. 2021). The two CBDs are adjacent to each other and form a two-domain Ca²⁺ sensor called CBD12. Binding of intracellular Ca²⁺ to CBD12 activates NCX but inhibits the NCX of Drosophila, CALX. CALX and NCX CBD12 constructs display interdomain flexibility in the apo state but assume rigid interdomain arrangements in the Ca²⁺-bound state. CALX-CBD12 preferentially samples closed conformations, whereas the wide-open interdomain arrangement, characteristic of the Ca²⁺-bound state, is less frequently sampled. These results are consistent with the view that Ca²⁺ binding shifts the CBD12 conformational ensemble toward extended conformers, which could be a key step in the NCXs' allosteric regulation mechanism (de Souza Degenhardt et al. 2021).

Animals

Calx of Drosophila melanogaster
(O18367)

Plasma membrane sodium:calcium exchanger, NCX3, NAC3 or SLC8A3, controlling Ca²⁺ homeostasis. Extrudes 1 Ca²⁺ for 3 extracellular Na⁺ ions.  Potent inhibitors have been identified (Secondo et al. 2015). One such inhibitor of NCX transporters, ORM-10962, exhibits high efficacy and selectivity.  Selective NCX inhibition can exert positive as well as negative inotropic effects, depending on the actual operation mode of the NCX (Kohajda et al. 2016). Bepridil is a commonly used medication for arrhythmia and heart failure. It primarily exerts hemodynamic effects by inhibiting Na⁺/K⁺ movement and regulating Na⁺/Ca²⁺ exchange (Wei et al. 2022).

Animals

SLC8A3 of Homo sapiens

Sodium/calcium exchanger 1 (Na⁺/Ca²⁺-exchange protein 1), CNC NCX1 of 973 aas and 12 TMSs in a 6 + 6 TMS arrangement.  The cardiac isoform, CAX1.1, like the archaeal homologues for which high resolution 3-d structures are available (TC#s 2.A.19.5.3 and 2.A.19.8.2), have two aqueous ion permeation channels with cavities that can face the cytoplasm or the external medium (John et al. 2013). It exchanges one Ca²⁺ ion against three to four Na⁺ ions, and thereby contributes to the regulation of cytoplasmic Ca²⁺ levels and Ca²⁺-dependent cellular processes (Komuro et al. 1992; , Van Eylen et al. 2001; Kofuji et al. 1992). It also contributes to Ca²⁺ transport during excitation-contraction coupling in muscle. In a first phase, voltage-gated channels mediate the rapid increase of cytoplasmic Ca²⁺ levels due to release of Ca²⁺ stores from the endoplasmic reticulum. SLC8A1 mediates the export of Ca²⁺ from the cell during the next phase, so that cytoplasmic Ca²⁺ levels rapidly return to baseline. It is also required for normal embryonic heart development and the onset of heart contractions. Both NCX1 and NCX2 play important roles in the motility of the gastric fundus, ileum and distal colon (Nishiyama et al. 2016). An amphipathic α-helix in the NCX1 large intracellular loop controls NCX1 palmitoylation. Thus, NCX1 palmitoylation is governed by a distal secondary structure element rather than by local primary sequence (Plain et al. 2017). The anti-aging gene NM_026333 contributes to proton-induced aging via the NCX1-pathway (Osanai et al. 2018). Dynamic palmitoylation modulates its structure, affinity for lipid-ordered domains, and inhibition by XIP (Gök et al. 2020). Na⁺/Ca²⁺ exchanger, NCX1, and canonical transient receptor potential channel 6 (TRPC6) are recruited by STIM1 to mediate Store-Operated Calcium Entry in primary cortical neurons (Tedeschi et al. 2022). SEA0400 is a potent and selective Na⁺/Ca²⁺ exchanger (NCX) inhibitor (Iwamoto et al. 2004).  Along the proximal tubule and thick ascending limb of the kidney, Ca²⁺ and Na⁺ transport occur in parallel, but those processes were dissociated in the distal convoluted tubule (Hakimi et al. 2023). Xue et al. 2023 presented cryo-EM structures of human cardiac NCX1 in both inactivated and activated states, elucidating key structural elements important for NCX ion exchange and its modulation by cytosolic Ca²⁺ and Na⁺. They showed that the interactions between the ion-transporting transmembrane (TM) domain and the cytosolic regulatory domain define the activity of NCX. In the inward-facing state with low cytosolic [Ca²⁺], a TM-associated four-stranded beta-hub mediates tight packing between the TM and cytosolic domains, resulting in the formation of a stable inactivation assembly that blocks the TM movement required for ion exchange. Ca²⁺ binding to the cytosolic second Ca²⁺-binding domain (CBD2) disrupts this inactivation assembly which releases its constraint on the TM domain, yielding an active exchanger. Thus, the NCX1 structures provide an essential framework for the mechanistic understanding of the ion transport and cellular regulation of NCX family proteins (Xue et al. 2023).  Structural insight into the allosteric inhibition of human sodium-calcium exchanger NCX1 by XIP and SEA0400 have been published (Dong et al. 2024).  The cryo-EM structure of NCX1.3 in the presence of a specific inhibitor, SEA0400 shows that conserved ion-coordinating residues are exposed on the cytoplasmic face of NCX1.3, indicating that the observed structure is stabilized in an inward-facing conformation. The regulatory calcium-binding domains (CBDs) assemble with the ion-translocation transmembrane domain (TMD). The exchanger-inhibitory peptide (XIP) is trapped within a groove between the TMD and CBD2 and is predicted to clash with gating helices TMs(1/6) at the outward-facing state, thus hindering the conformational transition and promoting inactivation of the transporter. A bound SEA0400 molecule stiffens helix TM2ab and affects conformational rearrangements of TM2ab that are associated with the ion-exchange reaction, thus allosterically attenuating Ca²⁺-uptake activity of NCX1.3 (Dong et al. 2024).

Animals

SLC8A1 of Homo sapiens

Sodium/calcium exchanger 2 (Na⁺/Ca²⁺-exchange protein 2; NCX2; SLC8A2) of 921 aas and 11 TMSs.  Functional inhibition of NCX2 initially causes natriuresis, and further inhibition produces hypercalciuria, suggesting that the functional significance of NCX2 lies in Na⁺ and Ca⁺ reabsorption in the kidney (Gotoh et al. 2015).  However NCX1-3 are present in the brain where they influence stroke theraputic strategies in a NCX subtype-specific fashion (Shenoda 2015). NCX1 and NCX2 play important roles in the motility of the gastric fundus, ileum and distal colon, but only NCX2 plays a role in the development of diarrhea (Nishiyama et al. 2016).

Animals

SLC8A2 of Homo sapiens

Rod photoreceptor Ca²⁺ + K⁺:4 Na⁺ antiporter, NCKX1. NCKX1 and heterologously expressed NCKX2 operate at a 4Na⁺:1Ca²⁺+1 K⁺ stoichiometry; both NCKX1 and NCKX2 are bidirectional transporters normally extruding Ca²⁺ from the cell (forward exchange), but also able to carry Ca²⁺ into the cell (reverse exchange) when the transmembrane Na⁺ gradient is reversed. Sequence changes have been observed for both NCKX1 and NCKX2 in patients with retinal diseases (Schnetkamp 2004).

Animals

Ca²⁺ + K⁺:Na⁺ antiporter (NCKX1) of Bos taurus

Sodium/potassium/calcium exchanger 3 (Na⁺/K⁺/Ca²⁺-exchange protein 3), NCKX3, or (Solute carrier family 24 member 3), SLC24A3 (Yang et al. 2013).

Animals

SLC24A3 of Homo sapiens

Putative Ca²⁺:cation exchanger of 1524 aas and an apparent duplication with 27 putative
TMSs and at leaswt 4 repeat units of 4 - 6 TMSs in the arrangement:  1 - 4, 1 - 220 aas; 5 - 9, 240 - 410 aas; 10 - 15, 460 - 650 aas; 16 - 21, 660 - 940 aas; and 22 - 27, 990 - 1200 aas.  It also has a C-terminal PAN-APP domain as in plasminogen.

Animals

Putative Ca²⁺: cation exchanger of Branchiostoma floridae

Uncharacterized protein of 623 aas.

Algae

UP of Aureococcus anophagefferens (Harmful bloom alga)

Ca²⁺:Na⁺ exchanger, NCX-9 of 651 aas and 14 TMSs.  Plays a role in developmental cell patterning and Ca²⁺ exchange in mitochondrial (Sharma et al. 2017).

NCX-9 of Caenorhabditis elegans

Putative sodium:calcium symporter of 377 aas and 10 TMSs.

Na⁺:Ca²⁺ symporter of Pseudoalteromonas phage J2-1

Uncharacterized protein of 575 aas and 10 TMSs in a 4 + 6 TMS arrangement.

UP of Entamoeba histolytica

The major neuronal Ca²⁺ + K⁺:4 Na⁺ antiporter, NCKX2. NCKX1 and NCKX2 operate with 4Na⁺:1Ca²⁺+1 K⁺ stoichiometry; both are bidirectional transporters normally extruding Ca²⁺ from the cell (forward exchange), but also able to carry Ca²⁺ into the cell (reverse exchange) when the transmembrane Na⁺ gradient is reversed. Sequence changes have been observed for both NCKX1 and NCKX2 in patients with retinal diseases (Schnetkamp 2004).

Animals

NCKX2 of Rattus norvegicus

K⁺-dependent Na⁺/Ca²⁺ antiporter, NCKX6 (Cai and Lytton, 2004). CCKX6 (NCLX) is an essential component of the mitochondrial Na⁺/Ca²⁺ exchanger (Palty et al., 2010; Drago et al., 2011).  It usually mediates mitochondrial Ca²⁺ extrusion (De Marchi et al. 2014). However, the mitochondrial calcium uniporter channel (MCU) and mitochondrial Na⁺ /Ca²⁺ exchanger, NCLX, mediate Ca²⁺ entry into and release from this organelle and couple cytosolic Ca²⁺ and Na⁺ fluctuations with cellular energetics (Verkhratsky et al. 2017). Abnormal levels occur in plasma neuron-derived extracellular vesicles of early schizophrenia and other neurodevelopmental diseases (Goetzl et al. 2022). Members of this family have the Glt fold (Ferrada and Superti-Furga 2022).  The mitochondrial calcium uniporter (MCU) is involved in an ischemic postconditioning effect against ischemic reperfusion brain injury in mice (Sasaki et al. 2024). Mitochondrial sodium/calcium exchanger (NCLX) regulates basal and starvation-induced autophagy through calcium signaling (Ramos et al. 2024).

Animals

SLC24A6 of Homo sapiens

The K⁺-dependent Na⁺/Ca²⁺ exchanger, MCKX4 (has 40x higher affinity for K⁺ than NCKX2 due to a threonine to alanine substitution at position 551 in NCKX2 (Visser et al., 2007)). NCKX4 is highly expressed and regulates Ca²⁺ transport in ameloblasts during amelogenesis (the formation of tooth enamel). In fact, MCKX4 is critical for enamel maturation (Wang et al. 2014). Residues involved in Na⁺ binding have been identified (Altimimi et al. 2010).

Animals

SLC24A4 of Homo sapiens

Trans-Golgi network K⁺-dependent Na⁺/Ca²⁺ antiporter SLC24A5 (NCKX5) (regulates melanogenesis; determines skin color variation) (Ginger et al., 2008).

Animals

SLC24A5 of Homo sapiens

Plants

CAX9 of Arabidopsis thaliana (Q9LJI2)

K⁺ uptake and Na⁺ transporter, CCX5 (CAX11) (Zhang et al., 2011).

Plants

CCX5 of Arabidopsis thaliana (O04034)

Na⁺/K⁺/Ca²⁺ exchanger-1 isoform 1, NCKX-1

Putative Ca²⁺:H⁺ or Ca²⁺:Na⁺ antiporter with two 5 TMS internal repeats (Sääf et al. 2001).

Bacteria and Archaea

ChaB (YrbG) of E. coli

Na⁺:Ca²⁺ exchanger, NCX_Mj (3-d structure known at 1.9 Å resolution; PDB# 3V5U (Liao et al., 2012). Contains 10 TMSs with two 5 TMS repeats. Four ion binding sites near the center of the protein are present, one specific for Ca²⁺ and three probably for Na⁺. Two passageways allow for Na⁺ and Ca²⁺ access from the external side.  However see a more recent analysis reported for 2.A.19.8.2 (Nishizawa et al. 2013).  Transport of both Na⁺ and Ca²⁺ requires protonation of D240, but this side chain does not coordinate either ion, implying that the ion exchange stoichiometry is 3:1 and that translocation of Na⁺ across the membrane is electrogenic although transport of Ca²⁺ is not (Marinelli et al. 2014). This system has been reviewed and considered to be a model protein for the entire family (Khananshvili 2021).

Archaea

NCX_Mj of Methanococcus (Methanocaldococcus) jannaschii (Q57556)

Na⁺/Ca²⁺ exchanger. Transport is electrogenic with a likely stoichiometry of 3 or more Na⁺ for each Ca²⁺ but K⁺-independent (Besserer et al. 2012).

MaX1 of Methanosarcina acetivorans

Vacuolar electrogenic Mg²⁺, Zn²⁺, Fe²⁺, and possibly Cd²⁺:H⁺ antiporter, MHX (found in the vascular cylinder; may control the partitioning of Mg²⁺ and Zn²⁺ between plant organs).  MHX porters are found only in plants and probably have 9 TMSs.  Their properties have been reviewed (Gaash et al. 2013).

Plants

MHX of Arabidopsis thaliana
(O22252)

Low affinity vacuolar monovalent cation (Na⁺ (K_m=20 mM) or K⁺(K_m=80 mM)):H⁺ antiporter, Vnx1. (Ca²⁺ is not transported; plays roles in ion and pH homeostasis) (Cagnac et al., 2007)

Yeast

Vnx1 of Saccharomyces cerevisiae (P42839)

Uncharacterized protein of 739 aas.

Animals

UP of Ornithorhynchus anatinus (Duckbill platypus)

Calcium:proton exchanger, CAX(CK31).  The function was demonstrated by purification and reconstitution in liposomes (Ridilla et al. 2012).  The protein forms dimers in the membrane but can be purified as a monomer.  The dimer interface seems to involve TMSs 2 and 6 (Ridilla et al. 2012).

Bacteria

CAX(CD31) of Caulobacter sp. strain K31

Ca²⁺:H⁺ antiporter of 405 aas, CAX_Af.  The inward facing 3-d structure has been solved to 2.3 Å resolution (Nishizawa et al. 2013).  The authors compare this structure to the outward facing 1.9 Å structure of NCX_Mj (TC# 2.A.19.5.3) and suggest that Ca²⁺ or H⁺ binds to the cation-binding site mutually exclusively.  The first and sixth TMSs alternately create hydrophilic cavities on the intra- and extracellluar sides of the membrane.  The inward and outward-facing transitions are triggered by ion binding (Nishizawa et al. 2013).

Archaea

Ca²⁺:H⁺ antiporter CAX_Af of Archaeoglobus fulgidus

Mg²⁺ transporter (Mg²⁺-specific channel-like exchanger) of 550 aas (Preston and Kung 1994; Haynes et al. 2002).  Has 10 putative TMSs in a 5 + 5 TMS arrangement and exhibits properties of a channel (Haynes et al. 2002).  The mutant form is called 'eccentric' and exhibits backwards swimming behavior (Preston and Kung 1994). 

Ciliates (Alveolata)

Ca²⁺-dependent Mg²⁺ transporter of Paramecium tetraurelia

Probable Mg²⁺-specific channel-like exchanger of 625 aas.

Ciliates (Alveolata)

Probable Mg2+-specific channel-like exchanger of Tetrahymena thermophila