2.A.40 The Nucleobase/Ascorbate Transporter (NAT) or Nucleobase:Cation Symporter-2 (NCS2) Family

The NCS2 family, also called the nucleobase/ascorbate transporter (NAT) family (Koukaki et al. 2005; Karatza et al., 2006), consists of over 1000 sequenced proteins derived from Gram-negative and Gram-positive bacteria, archaea, fungi, plants and animals. Of the five known families of transporters that act on nucleobases, it is the only one that is widespread (;(Gournas et al. 2008; Diallinas and Gournas 2013;  (Frillingos 2012).  Many functionally characterized members are specific for nucleobases including both purines and pyrimidines, but others are purine-specific. However, two closely related rat/mouse/human members of the family, SVCT1 and SVCT2, localized to different tissues of the body, cotransport L-ascorbate and Na+ with a high degree of specificity and high affinity for the vitamin (Diallinas and Gournas 2011). Clustering of NAT/NCS2 family members on the phylogenetic tree is complex with bacterial proteins and eukaryotic proteins each falling into at least three distinct clusters. The plant and animal proteins cluster loosely together, but the fungal proteins branch from one of the three bacterial clusters (Gournas et al. 2008). E. coli possesses four distantly related paralogous members of the NCS2 family.  Evidence that this family is a member of the APC superfamily has been presented (Wong et al. 2012). Members of this family have the UraA fold (Ferrada and Superti-Furga 2022).

Proteins of the NCS2 family are 414-650 amino acyl residues in length and probably possess 14 TMSs. Lu et al. (2011) have concluded from x-ray crystallography that UraA (2.A.40.1.1) has 14 TMSs with two 7 TMS inverted repeats. A pair of antiparallel β-strands is located between TMS 3 and TMS 10 and has an important role in structural organization and substrate recognition. The structure is spatially arranged into a core domain and a gate domain. Uracil, located at the interface between the two domains, is coordinated mainly by residues from the core domain. Structural analyses and relationships to other structurally members of the APC superfamily suggest that alternating access of the substrate may be achieved through conformational changes of the gate domain (Wong et al. 2012). 

The first 3-d structure of a eukaryotic NCS2 family member to be crystalized was that of UapA (Alguel et al. 2016).  This structure is similar to UraA, but additionally revealed that NATs dimerize and that the dimer is probably the functional unit. Dimerization appeared to be critical for specificity. Subsequent publications on UraA showed that this porter is also dimeric (Yu et al. 2017). Further analyses confirmed primary sequence comparitive data showing that the NCB2 family is a member of the APC superfamily (Vastermark et al. 2014). This conclusion has been further verified (Chang and Geertsma 2017).  The 7+7 TMS inverted repeat topology of UapA/UraA is also found in several transporters of the APC suprefamily with little primary amino acid sequence similarity with NATs, such as AzgA-like purine transporters (TC# 2.A.40.7.1), plant boron transporters Bor1-3 (e.g., TC# 2.A.31.3), the human Band3 anion exchanger (TC#2.A.31.1.1), and members of SulP transporter family (TC# 2.A.53). All these may be homodimeric transporters which seem to function via the so-called “elevator mechanism” of transport.

The generalized transport reactions catalyzed by proteins of the NAT/NCS2 are:

Nucleobase (out) +  H+ (out) → Nucleobase (in) + H+ (in)

Ascorbate (out) +  Na+ (out) → Ascorbate (in) +  Na+ (in).

This family belongs to the APC Superfamily.



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TC#NameOrganismal TypeExample

Uracil permease, UraA. The crystal structure of UraA with bound uracil at 2.8 Å resolution is available (PDB: 3QE7) (Lu et al., 2011). UraA has a novel structural fold, with 14 TMSs divided into two inverted repeats. A pair of antiparallel β-strands is located between TMS3 and TMS10 and has an important role in structural organization and substrate recognition. The structure is spatially arranged into a core domain and a gate domain. Uracil, located at the interface between the two domains, is coordinated mainly by residues from the core domain. Structural analysis suggests that alternating access of the substrate may be achieved through conformational changes of the gate domain.  Multiscale molecular dynamics simulations of the UraA symporter in phospholipid bilayers revealed a closed state with 3 high affinity binding sites for cardolipin (Kalli et al. 2015).The crystal structure of UraA bound to uracil in an occluded state at 2.5 A resolution (Yu et al. 2017). UraA shows substantial motions between the core domain and the gate domain as well as intra-domain rearrangements of the gate domain. The occluded UraA forms a dimer wherein the gate domains are sandwiched by two core domains. Dimer formation is necessary for transport activity (Yu et al. 2017).


UraA of E. coli (P0AGM7)


High affinity uracil permease (Martinussen et al. 2001).


PyrP of Lactococcus lactis (gbCAB89870)


Pyrimidine transporter of broad specificity, RutG of 442 aas and 14 TMSs; transports both uracil and thymine with high affinity  as well as, xanthine with low efficiency (Loh et al., 2006; Kim et al. 2010; Botou et al. 2018).


RutG of E. coli (P75892)

2.A.40.1.4Uracil permease BacilliPyrP of Bacillus subtilis

TC#NameOrganismal TypeExample
2.A.40.2.1Purine permease Bacteria YcpX of Clostridium perfringens

Putative xanthine/uracil/vitamin C permease of 529 aas and 13 TMSs.

Putative permease of Methanocorpusculum labreanum


Uncharacterized putative purine permease of 427 aas and 13 or 14 TMSs. 

UP of Pyrococcus furiosus


TC#NameOrganismal TypeExample
2.A.40.3.1Xanthine permease Bacteria PbuX (XanP) of Bacillus subtilis

Uric acid permease of 449 aas and 13 TMSs, PucJ.  May function together with PucK (TC# 2.A.40.3.4) (Schultz et al. 2001).


PucJ of Bacillus subtilis


Uric acid (urate) permease YgfU. YgfU exhibits low affinity (0.5 mM) but high capacity for urate and very poor activity for xanthine.  Essential residues were identified.  Coversion of Thr-100 to ala resulted in efficient xanthine transport (Papakostas and Frillingos 2012).  


YgfU of Escherichia coli


Uric acid uptake porter of 430 aas and 13 TMSs, PucK.  May function together with PucJ (TC# 2.A.40.3.2) (Schultz et al. 2001). 

PucK of Bacillus subtilis


TC#NameOrganismal TypeExample

High affinity uric acid-xanthine permease, UapA. Functionaly critical residues in transmembrane segments 1 and 3 have been identified (Amillis et al., 2011). The substrate recognition and transport pathway have been proposed (Kosti et al., 2012; Kosti et al. 2010).  UapA oligomerization is essential for membrane trafficking and turnover and is a common theme in fungi and mammalian cells (Martzoukou et al. 2015).  Specificity is determined by the interactions of a given substrate with the TMS8-9 loop and by interactions of this loop with TMS1 and TMS12 (Papageorgiou et al. 2008). F528 and Q408 in TMS 12 are important for substrate recognition, and mutation of the former results in high efficiency uptake of several purines and pyrimidines not otherwise transported (Vlanti et al. 2006). A high resolution structure of UapA is available, and it is formed from two domains, a core domain and a gate domain, similar to the previously solved uracil transporter UraA, which belongs to the same family (Alguel et al. 2016). The structure shows UapA in an inward-facing conformation with xanthine bound to residues in the core domain. Unlike UraA, which is a monomer, UapA forms a dimer in the crystals with dimer interactions formed exclusively through the gate domain. Analysis of dominant negative mutants is consistent with dimerization playing a key role in transport. Alguel et al. 2016 postulated that UapA uses an elevator transport mechanism likely to be shared with other structurally homologous transporters including anion exchangers and prestin. Specific residues in UapA are critical for dimerization, ER-exit and function (Kourkoulou et al. 2019). Despite structural and functional differences, all elevator-type transporters use a common mechanism of substrate translocation via reversible movements of a mobile core domain (the elevator) hosting the substrate binding site along a rigid scaffold domain stably anchored in the plasma membrane via homodimerization (Dimakis et al. 2022). One of the best studied elevator transporters is the UapA uric acid-xanthine/H+ symporter of the filamentous fungus Aspergillus nidulans. TMSs 5 and 12 in UapA control, negatively or positively, the dynamics of transport as well as substrate binding affinity and specificity. Mutations in TMS5 can lead to increased rate of transport, but also to an inactive transporter due to high-affinity substrate-trapping, whereas mutations in TMS12 lead to apparently uncontrolled sliding and broadened specificity, leading in specific cases to UapA-mediated purine toxicity.  The interactome of the UapA transporter revealed putative new players in anterograde membrane cargo trafficking(Georgiou et al. 2023).



UapA of Emericella (Aspergillus) nidulans

2.A.40.4.2The putative xanthine permease, YicE (Karatza and Frillingos, 2005)BacteriaYicE of E. coli (POAGM9)

The YgfO (XanQ) purine (xanthine) transporter. Residues involved in substrate binding have been identified (Georgopoulou et al., 2010). TMS3 functions in substrate recognition (Karena and Frillingos, 2011).  Many more essential residues have more recently been identified (Karena et al. 2015). An "ancestral" homolog (AncXanQ) has been constructed and proved to have broader specificity, transporting with high-affinity both xanthine and guanine, but also recognizing adenine, hypoxanthine, and a range of analogs (Tatsaki et al. 2021). AncXanQ conserves all binding-site residues of XanQ and differs substantially in only five intramembrane residues outside the binding site. Tatsaki et al. 2021 subjected both homologs to rationally designed mutagenesis and presented evidence that these five residues are linked with the specificity changes.


XanQ of E. coli (P67444)


Purine (uric acid and xanthine) permease, UapC.  Present in many Ascomycetes (Krypotou and Diallinas 2014).  Drought is an important abiotic stress that constrains the quality and quantity of tea plants. The green leaf volatiles, Z-3-hexenyl acetate (Z-3-HAC), play an essential role in stress responses (Wang et al. 2023).


UapC of Emericella nidulans


XanQ or PbuX, purine base uptake porter specific for xanthine and guanine. It is of 463 aas and 13 or 14 TMSs in a 5 + 3 or 4 +1 + 4 TMS arrangement (Tatsaki et al. 2021).

XanQ of Neisseria meningitidis


TC#NameOrganismal TypeExample

Putative purine permease, YbbY.  The ybbY gene is in an operon involved with allantoin metabolism, and is flanked by allB, encoding allantoinase, and the glxK gene, encoding glycerate kinase II. Downstream of glxK is YlbA, encoding S-uridoglycine aminohydrolase, the second enzyme involved in allantoin degradation (Moraes and Reithmeier 2012).


YbbY of E. coli


Putative purine permease of 440 aas and 13 TMSs, YwdJ.  It belongs to the HCO3_cotransp/Xan_ur_permease families in CDD.

YwkJ of Bacillus subtilis


Putative xanthine/uracil/vitamin C permease of 431 aas and 12 TMSs.

UP of Sebaldella termitidis


TC#NameOrganismal TypeExample

L-ascorbate:Na+ symporter, SVCT1. (L-ascorbate:Na+= 1:2; Mackenzie et al., 2008). Iron regulates SVCT1 in human intestinal Caco-2 cells (Scheers and Sandberg, 2011).


SVCT1 of Rattus norvegicus


Ca2+/Mg2+-dependent L-ascorbate:Na+ symporter, SVCT2; Na+:ascorbate = 2:1; binding order: Na+, ascorbate, Na+ (Na+ increases the affinity for ascorbate); Ca2+/Mg2+ are required for function) (Godoy et al., 2007; Bürzle et al. 2013). SVCT2-GLUT1-mediated ascorbic acid transport pathway in rat dental pulp has been studied, and its effects during wound healing have been described (Ohkura et al. 2023).


SLC23A2 of Homo sapiens

2.A.40.6.3High affinity (Km = 30 µM) uric acid-xanthine transporter; leaf permease protein 1, LPE1 (necessary for proper chloroplast development in maize) (Argyrou et al., 2001) PlantsLPE1 of Zea mays (Q41760)

solute carrier family 23 (nucleobase transporters), member 3, SVCT3 or SLC23A3.  Function not certain as of 1/2013 (Bürzle et al. 2013).


SLC23A3 of Homo sapiens


Solute carrier family 23 member 1 (Na+/L-ascorbic acid transporter 1; Sodium-dependent vitamin C transporter 1) (hSVCT1; Yolk sac permease-like molecule 3) (Bürzle et al. 2013). Members of the SLC23 family have the UraA fold (Ferrada and Superti-Furga 2022).


SLC23A1 of Homo sapiens

2.A.40.6.6Nucleobase-ascorbate transporter 12 (AtNAT12)PlantsNAT12 of Arabidopsis thaliana

Solute carrier family 23 member 2-like protein (SLC23.A.4) of 614 aas and 13 or 14 TMSs, SNBT1.  Mutations allowed functional insertion into fungal membranes (Kourkoulou et al. 2019). It transports urate and purine nucleobases in a sodium ion-dependent process (Yamamoto et al. 2010; Yasujima et al. 2018).

SNBT1 of Rattus norvegicus, the Norway rat.


TC#NameOrganismal TypeExample

The purine (hypoxanthine/adenine/guanine) transporter, AzgA (Cecchetto et al., 2004).  Topological modeling has revealed a potential substrate binding cavity, and residues important for transport activity have been identified (Krypotou et al. 2014).


AzgA of Aspergillus (Emericella) nidulans (CAE00849)

2.A.40.7.2Hypoxanthine/guanosine uptake transporter, PbuG (Johansen et al., 2003)BacteriaPbuG of Bacillus subtilis (CAB12456)

The purine transporter Azg1 (takes up 8-azaadenine and 8-azaguanine but not other toxic nucleobase analogues; similar to Azg2 of A. thaliana (Q84MA8); (Mansfield et al. 2009). AZG1 is a cytokinin transporter that interacts with auxin transporter PIN1 (TC# 2.A.69.1.1) and regulates the root stress response (Tessi et al. 2023). The structures and mechanisms of the Arabidopsis cytokinin transporter AZG1 have been determined (Xu et al. 2024).


Azg1 of Arabidopsis thaliana (Q9SRK7)


  Adenine permease, YicO.  Also recognizes with low micromolar affinity N(6)-benzoyladenine, 2,6-diaminopurine, and purines (Papakostas et al. 2013).


YicO of Escherichia coli


Purine base permease, GhxP or YjcD.  Transports purines such as guanine, hypoxanthine, and xanthine.  Also transports mutagenic purines such as 6-N-hydroxylaminopurine (HAP), 2-amino-HAP (AHAP), 6-mercaptopurine, 6-thioguanine, 1-methylguanine, 8-azaguanine, 6-thioguanine and 2-aminopurine (Kozmin et al. 2013; Papakostas et al. 2013).  


YjcD (GhxP) of E. coli


Adenine permease, PurP.  Also recognize with low micromolar affinity N(6)-benzoyladenine, 2,6-diaminopurine, and purine (Papakostas et al. 2013).


PurP of Escherichia coli


Guanine/hypoxanthine uptake porter of 455 aas, GhxQ, YgfQ or YgfR.  Also takes up mutagens such as 1-methylguanine, 8-azaguanine, 6-thioguanine, and 6-mercaptopurine (Papakostas et al. 2013) and catalyzes dis-C3 (membrane-permeable, cationic fluorescent dye, the carbocyanine diS-C3 efflux (Jindal et al. 2019).


YgfQ of E. coli


NCS2 family permease of 429 aas and 13 apparent TMSs in a 7 + 6 TMS arrangement.

NCS2 family protein of Tissierellia bacterium


TC#NameOrganismal TypeExample

Uncharacterized protein of 552 aas and 16 TMSs.

UP of Verrucosispora sediminis


Xanthine permease of 520 aas and 16 TMSs in a 7 + 2 + 7 TMS arrangement.

Xanthine permease of Providencia stuartii


Uncharacterized protein of 534 aas and 15 apparent TMSs in a 7 + 2 + 6 TMS arrangement.

UP of Stenotrophomonas maltophilia


Uncharacterized protein of 604 aas with 14 or 16 TMSs in a 6 + 6 or 7 + 7 TMS arrangement.

UP of Cavenderia fasciculata


Uncharacterized protein of 759 aas with about 14 TMSs in an apparent 4 + 7 + 3 TMS arrangement. 

UP of Aureococcus anophagefferens


Uncharacterized protein of 520 aas and ~ 14 - 16 TMSs, possibly in a 7 + 2 + 7 TMS arrangement.

UP of Alicyclobacillus sp.


Uncharacterized permease of 562 aas and possibly 16 TMSs in a 3 + 4 + 2 + 3 + 4 TMS arrangement.

UP of Planctomycetes bacterium (freshwater metagenome)


Uncharacterized protein of 568 aas and ~ 16 TMSs in a 3 + 4 + 2 + 3 + 4 TMS arrangement.

UP of Polarella glacialis