ABCG2 is one of a few human membrane transporters which display the amazing ability to transport multiple different chemicals out of cells. These multidrug pumps, which have orthologues in all organisms, are important in humans in the context of drug pharmacokinetics, especially with respect to resistance to chemotherapy. In 2016, we presented a mini-review on ABCG2 which identified many areas of exciting research progress as well as many areas of frustrating ignorance. Just 2 years on the field has advanced, particularly with respect to structural biology as the cryo-electron microscopy revolution has brought us new insights into the structure and mechanism of ABCG2. In this update, we evaluate the degree to which new data have enhanced our understanding of the structure and mechanism of ABCG2 and whether we are now in a position to translate some of these findings into inhibitor design and development.

Introduction

It is getting close to 50 years since the first seeds in a new field of membrane protein research were sown with the identification of energy-dependent nutrient uptake systems in bacteria. The next 20–30 years of research resulted in the recognition of phenomenal functional diversity in the family of ATP-binding cassette (ABC) proteins [1,2]. Not only could they import or export substrates from cells and organelles, but the domain which powered that transport, the eponymous ABC or nucleotide-binding domain (NBD) could be subjugated into controlling chromosomal segregation, ribosome protection and cell division [3,4]. It has also become clear that ABC transporters have evolved to be able to handle an extraordinary repertoire of ‘cargo', from hydrophilic amino acids and sugars to lipids and other hydrophobic chemicals and even entire proteins. Quite how the relatively conserved architecture of an ABC transporter manages this feat remains to be resolved.

ABC proteins are one of the most widely represented classes of membrane protein in many sequenced organisms, with humans having 48 ABC protein-encoding genes. Of these, at least 3, but potentially several others, have a role in multidrug efflux, displaying the ability to transport numerous diverse chemical substrates out of cells. The current review focusses on ABCG2; this transporter remained in the shadows and lagged behind its larger cousin ABCB1/P-glycoprotein (P-gp) for many years in terms of structural and functional understanding [5]. P-gp was identified and sequenced in 1986, ABCG2 a dozen or so years later [6,7]; P-gp was purified and early structural data were obtained in the late 1990s [8,9], a full decade before ABCG2 was as well studied [10]. However, in recent years the ABCG2 research field has seen an explosion of exciting findings that have widened our understanding of its structure and mechanism (which we expand upon below) and also its wide role in physiology and pathophysiology.

ABCG2's original identification in multidrug-resistant breast cancer cell lines or placental tissues led to it having the initial abbreviations ABCP (ABC transporter of the placenta) or BCRP (breast cancer resistance protein) [7,11]. Subsequent studies have confirmed a clinical correlation between ABCG2 expression and outcome in many tumours (particularly those of haematological origin [12]). Its identification as a multidrug transporter has seen ABCG2 recognised as an important contributor to the pharmacokinetics of many prescribed drugs — and indeed all new drugs have to be tested for ABCG2-mediated transport ([13] and Table 1). The range of physiological substrates for ABCG2 and thus the range of functional roles continues to expand [21]; in recent years, protoporphyrins have been shown to be transported, leading ABCG2 to be shown to have a role in haematopoietic stem cell protection [19]; subsequently, urate was identified as a substrate after several single nucleotide polymorphisms of ABCG2 (most prominently rs2231142, Q141K) were determined as strongly predictive of serum hyperuricaemia [23]. Rs2231142 and other gout-associated polymorphisms also have a projected role in delaying onset of Parkinson's disease, where elevated serum urate levels are considered neuroprotective [24]. ABCG2's involvement in neurodegenerative diseases has also been widened to include Alzheimer's disease (AD) after the description of its ability to transport the amyloid β1–40 peptide and an association of a polymorphism to AD development [15,25]. Finally, perhaps we should not use that word given the surprises ABCG2 has had for us already, the transporter has been implicated as a regulator of autophagy [26] — the controlled process of cellular recycling — although what role ABCG2 plays here is unclear.

Table 1
Functions and substrates of ABCG2

The above is a foreshortened list of substrates and physiological roles of ABCG2. Greater detail can be found in two recent more extensive reviews [21,22].

Physiological location Function References 
Gastro-intestinal tract Reduced absorption of drugs (e.g. camptothecins, tyrosine kinase inhibitors, prazosin, statins, HIV protease inhibitors); extra-renal clearance of uric acid [14
Liver canaliculi Biliary excretion of drugs and metabolites (e.g. SN-38) [14
Blood–brain barrier Reduced penetrance of drugs (e.g. opiods) into CNS; transport of Aβ1–40 [15,16
Placenta, Mammary gland Apparent contradictory physiological roles? Protection of foetus vs. excretion of toxins and nutrients into milk [17,18
Haematopoietic system Efflux of porphyrins [19,20
Physiological location Function References 
Gastro-intestinal tract Reduced absorption of drugs (e.g. camptothecins, tyrosine kinase inhibitors, prazosin, statins, HIV protease inhibitors); extra-renal clearance of uric acid [14
Liver canaliculi Biliary excretion of drugs and metabolites (e.g. SN-38) [14
Blood–brain barrier Reduced penetrance of drugs (e.g. opiods) into CNS; transport of Aβ1–40 [15,16
Placenta, Mammary gland Apparent contradictory physiological roles? Protection of foetus vs. excretion of toxins and nutrients into milk [17,18
Haematopoietic system Efflux of porphyrins [19,20

One thing is clear: ABCG2 packs a huge punch in terms of functional diversity (see Table 1 and [5]). To better understand the molecular basis for its involvement in all these processes, we need to know more about the mechanism of the transporter, and for that we need to know the structure at high resolution. Such information has been instrumental in understanding other ABC MDR pumps, as described in an accompanying review [27].

ABCG2 structure: the hazy past

The earliest attempt to provide structural data for ABCG2 came from single-particle electron microscopy (EM) of protein purified from insect cells infected with recombinant ABCG2-expressing baculovirus [10]. The data were limited to 18 Å resolution at which it was only possible to identify that the particles were of a large oligomeric complex. Computational fitting of the electron density with Sav1866, the highest resolution ABC exporter structure of the time [28], suggested that ABCG2 was present as a tetramer of dimers. Higher resolution data from a later electron crystallographic study of human ABCG2 overexpressed in Pichia pastoris enabled a comparison of the structure in 2D crystals in the presence or absence of the drug substrate mitoxantrone [29]. This study revealed a more ordered conformation in the presence of drug substrate, and homology models based on MsbA as the template were used to rationalise the structural data, and provided evidence that ABCG2 undergoes changes in conformation from an outward facing to an inward facing conformation in the presence or absence of bound drug, respectively [29], in accordance with the power stroke model for the transporter [30]. Despite the improved resolution (up to 5 Å), this was a projection study in 2D, without 3D reconstruction, and so the number of transmembrane helices and their relative orientations remained unclear. Indeed, extension of that study to a 3D structure (resulting from tilting of 2D crystals) resulted in data to only ∼20 Å resolution, again precluding any insights into the structural details of the transport process [31].

Such was the need for structural data for ABCG2 that there were attempts to model the protein using structures that were available for the bacterial half transporters Sav1866 and MsbA [28,32], which both can function as multidrug pumps. These models were constructed despite the low primary sequence similarity between ABCG2 and prokaryotic pumps, and indeed despite the different domain organisation; i.e. that ABCG2 has an N-terminal NBD, whereas Sav1866 and MsbA have an N-terminal transmembrane domain (TMD) [33]. Unsurprisingly these models were unable to advance our understanding of the structure and molecular mechanism of ABCG2 [34,35], and subsequent higher resolution structures have revealed that these models did not accurately reflect the TMD fold of ABCG2.

The cryo-EM resolution revolution

Early cryo-EM data for ABCG2 posed as many questions as it answered; this in itself was not a huge surprise. After all, until the mid-2000s cryo-EM was seen by some, quite unfairly, as a last resort for structural biologists. For other ABC transporters, even the highest quality 2D crystals, obtained after reconstitution at high protein:lipid ratios, produced structural data to medium (8 Å) resolution (see ABCB1 structure at this resolution [36]). Although such data allowed us to distinguish between different states in the catalytic cycle [37], it was certainly insufficient as a structural description of an ABC transporter. Single-particle analysis and averaging typically produced even lower resolution data and so it seemed EM would remain less favoured.

In the last decade though, developments in technical and analytical fields have synergised to provide a quantum leap forward for EM in structural biology [38]. There have been three principal developments. Firstly, the use of direct electron detectors with ultra-thin support matrices to prevent reflected electrons contributing to the signal:noise ratio, thereby dramatically increasing the sensitivity of data collection. Secondly, the digital capture of movies at high frame rates (hundreds/s) has enabled electron beam induced movements to be corrected for. This has dramatically increased the resolution of the collected data in the same way that image stabilisation in a digital camera improves the final image. Finally, there have been huge advances in computational analysis resulting in better particle classification algorithms. In totality, these advances have led to an explosion in the number of primarily large proteins (>100 kDa) or macromolecular complexes solved at near atomic resolution in non-crystalline specimens (see references within [38]).

Eukaryotic ABC transporters: structural progress made through cryo-EM

Fortunately for the ABC transporter field, our pumps have been well represented in this structural revolution, and the discovery of structural diversity within ABC exporters has provided new insights into the multifarious functionalities of these membrane proteins. A brief overview of this progress is presented in Table 2, confirming that four of the five sub-families now have at least one structure determined by cryo-EM, with only the ABCD family remaining unsolved. Despite the progress made in determining membrane protein structures using cryo-EM, it is likely that this still represents the tip of an iceberg. Further technical advances in the field will doubtless lead to higher quality structural data, from smaller amounts of protein, potentially in the native lipid environment [49]. Combined with the wealth of biochemical knowledge about trapping ABC transporters in particular conformational states, this promises a multi-step structural description of ABC transporters within their native environment [27].

Table 2
Mammalian ABC transporter Cryo-EM structures
Protein Resolution (Å) Organism Expression system Conformation Key findings References 
ABCA1 4.1 for the overall structure and 3.9 for the ECD Human Sf9 insect cells Outward facing in nucleotide-free state TMDs in a type II exporter configuration. ABCA-family-specific regulatory domains act as a latch to stabilise NBD1 and NBD2 interactions. ABCA-family-specific extracellular domains (ECD) 1 and 2 co-fold around an extended hydrophobic tunnel [39
ABCB1 3.4 Human BacMam Outward open TMDs in a type I exporter configuration. Mg.ATP trapped at symmetric ATPase sites. Provides structural evidence for the long-held power stroke model [40
ABCB2/3 (TAP1/2) 6.5 Human Pichia Inward open TMDs in a type I exporter configuration. TMD0 missing from both TAP1/TAP2. Structural basis of inhibition by herpes simplex virus defined by structure of ICP47-binding site in proposed TMD translocation pathway [41,42
ABCC1 (MRP1) 3.3, 3.5 Bovine BacMam Inward open; apo and LTC4 bound Type I TMD organisation. Large binding site with structural plasticity identified. TMD0 identified but not the connecting ‘lasso' motif to TMD1 [43
ABCC7 (CFTR) Average 3.9, but as low as 6 in NBDs Human BacMam Inward open, apo, dephosphory-lated Type I TMD organisation. Difference in TM7 and TM8 helices compared with ABCC1 with potential ion binding sites due to a helical break [44
ABCC8 (SUR1) 5.5 Mouse Kir6.2 and Syrian hamster SUR1 BacMam Inward open Structure represents four regulatory SUR1 subunits bound onto Kir6.2. TMD0-and lasso motif mediate Kir/SUR interaction. Glibenclamide binding within TMD1 stabilises a closed Kir channel. Limited interactions between adjacent ABCC8 molecules [45,46
ABCG2 3.1–3.8; NBDs lower in [47Human 293-EBNA Inward open Type II TMD configuration. Initial structure at low resolution in NBDs; later structure improved NBD resolution. Conformational stabilisation of inactive state with inhibitory antibody (5D3). Potential transporter-gate residue and cholesterol-binding site observed. Binding sites identified for ABCG2 inhibitors Ko143 or Tariquidar [47,48
Protein Resolution (Å) Organism Expression system Conformation Key findings References 
ABCA1 4.1 for the overall structure and 3.9 for the ECD Human Sf9 insect cells Outward facing in nucleotide-free state TMDs in a type II exporter configuration. ABCA-family-specific regulatory domains act as a latch to stabilise NBD1 and NBD2 interactions. ABCA-family-specific extracellular domains (ECD) 1 and 2 co-fold around an extended hydrophobic tunnel [39
ABCB1 3.4 Human BacMam Outward open TMDs in a type I exporter configuration. Mg.ATP trapped at symmetric ATPase sites. Provides structural evidence for the long-held power stroke model [40
ABCB2/3 (TAP1/2) 6.5 Human Pichia Inward open TMDs in a type I exporter configuration. TMD0 missing from both TAP1/TAP2. Structural basis of inhibition by herpes simplex virus defined by structure of ICP47-binding site in proposed TMD translocation pathway [41,42
ABCC1 (MRP1) 3.3, 3.5 Bovine BacMam Inward open; apo and LTC4 bound Type I TMD organisation. Large binding site with structural plasticity identified. TMD0 identified but not the connecting ‘lasso' motif to TMD1 [43
ABCC7 (CFTR) Average 3.9, but as low as 6 in NBDs Human BacMam Inward open, apo, dephosphory-lated Type I TMD organisation. Difference in TM7 and TM8 helices compared with ABCC1 with potential ion binding sites due to a helical break [44
ABCC8 (SUR1) 5.5 Mouse Kir6.2 and Syrian hamster SUR1 BacMam Inward open Structure represents four regulatory SUR1 subunits bound onto Kir6.2. TMD0-and lasso motif mediate Kir/SUR interaction. Glibenclamide binding within TMD1 stabilises a closed Kir channel. Limited interactions between adjacent ABCC8 molecules [45,46
ABCG2 3.1–3.8; NBDs lower in [47Human 293-EBNA Inward open Type II TMD configuration. Initial structure at low resolution in NBDs; later structure improved NBD resolution. Conformational stabilisation of inactive state with inhibitory antibody (5D3). Potential transporter-gate residue and cholesterol-binding site observed. Binding sites identified for ABCG2 inhibitors Ko143 or Tariquidar [47,48

ABCG2 structure: bringing binding pockets into focus

As described above, the lack of high-resolution structural data compromised our understanding of the mechanism of action for ABCG2. The X-ray crystal structure of the sterol transporter ABCG5/G8 proved to be a landmark in understanding the ABCG subfamily [50], and this was quickly followed by homology models of ABCG2 that provided a framework to interpret existing experimental data in the light of its homology to ABCG5/G8 [5153]. As these papers came to print, the group of Kaspar Locher published a structure of human ABCG2 obtained at just under 4 Å resolution using single-particle cryo-EM of ABCG2 dimers in complex with 2 Fab fragments of the inhibitory 5D3 antibody (Table 2 and [47]). The ABCG2 structure confirmed that the ABCG5/G8 fold is broadly holding true for other ABCG family members. This fold (Figure 1) reveals a more compact conformation that is more reminiscent of bacterial ABC importers, as opposed to ABCB1 [54]. The fold is also that of a Type II exporter in which the TM helices do not cross over to contact the opposite NBDs, as seen in Type I exporter structures. The NBDs, which were initially modelled on those in the ABCG5/G8 structure due to the low resolution of the data, are typical of other ABC transporter NBDs, although they remain in contact with each other even in the absence of nucleotide [47], with an interaction surface comprising a conserved motif (NPXDF) at the C-terminus of NBD. This close NBD interaction is interesting when compared with the open apo structures of other MDR pumps (see the accompanying article [27] for more details). The ∼70 amino acid linker region that then connects to the TMD is not well resolved, but the structural quality improves again beginning with a short α-helix parallel to the plane of the membrane (referred to as helix 1a in Figure 1). An intracellular linker region (ICL1) between TM helix 2 and TM helix 3 contacts an α-helix in the NBD that includes residue Q141 and provides a structural basis for understanding the effects of the gout-associated polymorphism Q141K [23,47]. The final noteworthy feature of the TMD's secondary structure is a series of short α-helices (5b, 5c, 6a in Figure 1) connecting the membrane spans TM5 and TM6; these appear to provide a structural cap (‘capping helices') at the extracellular end of the TMD.

ABCG2 topology and structure.

Figure 1.
ABCG2 topology and structure.

(A) Schematic representation of the structural topology of ABCG2 with a unique TMD fold (within ABCG family) adopted by the TM helices 5 and 6 (raspberry and peach cylinders). Residues crucial for intra- and inter-molecular disulfide bond formation (C592, C603 and C608) and glycosylation (N596) have been marked. TM helices crucial in substrate binding and propulsion through the transporter have been highlighted (TM1a and TM1b; yellow, TM2; pale green, TM3; purple and TM6b). (B) Cartoon representation of the ABCG2 dimer (co-ordinates 6ETI, [48]). Helices are depicted in the same colours as in A. A 90° rotation enables an enhanced view of the access site described in Figure 2, surrounded by TM helices 2, 3 and 6b. The lower right panel depicts the leucine plug in red, with an enlarged view of cavity 1 (residues highlighted in bright orange) and cavity 2 encircled. Figure generated using PYMOL.

Figure 1.
ABCG2 topology and structure.

(A) Schematic representation of the structural topology of ABCG2 with a unique TMD fold (within ABCG family) adopted by the TM helices 5 and 6 (raspberry and peach cylinders). Residues crucial for intra- and inter-molecular disulfide bond formation (C592, C603 and C608) and glycosylation (N596) have been marked. TM helices crucial in substrate binding and propulsion through the transporter have been highlighted (TM1a and TM1b; yellow, TM2; pale green, TM3; purple and TM6b). (B) Cartoon representation of the ABCG2 dimer (co-ordinates 6ETI, [48]). Helices are depicted in the same colours as in A. A 90° rotation enables an enhanced view of the access site described in Figure 2, surrounded by TM helices 2, 3 and 6b. The lower right panel depicts the leucine plug in red, with an enlarged view of cavity 1 (residues highlighted in bright orange) and cavity 2 encircled. Figure generated using PYMOL.

The tertiary structure of the ABCG2 TMDs shows some features which may be relevant for understanding substrate and inhibitor binding. ABCG2 displays two cavities; the first of these (cavity 1) is accessible from the cytoplasmic side of the membrane and is separated from a second, extracellular site (cavity 2), by a conserved region (the ‘leucine plug'; residues L554–L554 of opposing monomers). This second cavity is then capped by the cluster of short helices described above. The relevance of these two cavities to ABCG2 mechanism was further revealed by a second structural study described in a 2018 paper in which novel derivatives of both the inhibitors Ko143 and tariquidar were synthesised and then localised to cavity 1 [48], lined by the transmembrane helices TM5 and TM2a of the opposite monomers. The Ko143 derivative was present at a 2:2 stoichiometry within ABCG2, whereas the tariquidar inhibitor was present at a 1:2 stoichiometry (i.e. 1 inhibitor per ABCG2 dimer), occupying structurally similar positions to a tentative cholesterol-binding pocket identified earlier [47]. The improved structure, and the construction of a panel of Ko143-derivatives, will facilitate the understanding of structure activity relationships for ABCG2 inhibitors.

ABCG2: a hypothetical mechanism for multidrug efflux

In the final section of this review, we propose a schematic model for substrate drug transport by ABCG2 (Figure 2), which has some parallels to other alternating access models for ABC exporters [55]. The model for ABCG2 illustrates three distinct drug interaction sites depicted in the centre of Figure 2: a surface access site (see below), a binding site (consistent with the structural ‘cavity 1') and an extrusion site (consistent with the structural ‘cavity 2'). We use the model to attempt a structure-based interpretation of several questions regarding current data:

  • Do the structural data enable us to propose models of substrate binding to ABCG2?

  • Do they offer an explanation of the transport process?

  • Can we explain why some drugs and inhibitors are not transported?

The questions are pertinent as positive answers could lead to genuine progress in targeted drug design for overcoming ABCG2-mediated pharmacological problems. Central to this discussion are the cavities described by the cryo-EM data, which are occupied by bound inhibitor(s), rather than by transported substrates. Indeed, while the cavities in the cryo-EM data do include some residues demonstrated to affect substrate specificity or transport (e.g. M549 [56] and I543 [57]), there are many other residues which have significant effects on transport function but which lie outside of cavity 1 (e.g. R482 and M496 in TM helix 3, and residues 523, 540, 548 and 640 among other TMs [5759]).

Asymmetric catalytic and transport cycle model for ABCG2.

Figure 2.
Asymmetric catalytic and transport cycle model for ABCG2.

As described in the text, we propose a model with three drug interaction sites described in the central smaller image for reference. Drug (yellow) binding to the access site (A) promotes ATP (green) binding and the formation of the classical NBD sandwich dimer (B). Two-way allosteric communication between the NBDs and TMDs results in the transfer of drug from the access site to cavity 1, and the occlusion of the access sites. ATP hydrolysis (C) promotes an outward open TMD conformation in which the leucine plug and the capping helices undergo structural changes allowing drug extrusion from cavity 2. ADP (orange) release (D) results in resetting of the transporter. For simplicity, we only show events resulting from binding to an access site on 1 half of the transporter. The homodimer nature of ABCG2 implies a second access site shown for reference.

Figure 2.
Asymmetric catalytic and transport cycle model for ABCG2.

As described in the text, we propose a model with three drug interaction sites described in the central smaller image for reference. Drug (yellow) binding to the access site (A) promotes ATP (green) binding and the formation of the classical NBD sandwich dimer (B). Two-way allosteric communication between the NBDs and TMDs results in the transfer of drug from the access site to cavity 1, and the occlusion of the access sites. ATP hydrolysis (C) promotes an outward open TMD conformation in which the leucine plug and the capping helices undergo structural changes allowing drug extrusion from cavity 2. ADP (orange) release (D) results in resetting of the transporter. For simplicity, we only show events resulting from binding to an access site on 1 half of the transporter. The homodimer nature of ABCG2 implies a second access site shown for reference.

We propose that access to cavity 1 for transported substrates is controlled by the surrounding TM helices; the mutagenesis data cited above support a model where the helical interface between the intracellular ends of helices TM2 and 3 permit access to cavity 1, and thus that mutations in the intracellular end of TM3 (e.g. R482) alter drug specificity (e.g. by allowing transport of daunorubicin). TM6, which is located on the lipid-exposed face of the transporter, partially shields the intracellular end of the TM2:3 interface from the lipid bilayer, and consistent with this, mutation of F640 to the much smaller alanine also results in gain of daunorubicin transport. We refer to these sites (as there would be two in an ABCG2 dimer) between TM2/3/6 as surface or ‘access sites' (Figure 2) and propose that binding of drug to an access site drives conformational changes at the NBDs resulting in ATP binding. The subsequent closure of the NBD dimer (Figure 2, Step B) results in further TMD:NBD communication that causes translocation of substrate drug from an access site to cavity 1. The binding of ATP therefore provides the power stroke for this conformational change in accordance with other ABC multidrug efflux pumps [30,40,60]. Recent biochemical and modelling data from Karl Kuchler's laboratory add molecular detail to this hypothesis by proposing that a series of charged and polar residues at the TMD:NBD interface (which includes the ICL1 between TM2 and TM3) act as an allosteric conduit (‘polar relay') between the NBDs and TMDs, with ICL1 acting as a molecular spring to couple the two domains [51]. We are tempted to speculate that the ‘molecular spring ‘ alters the exposure of the access site by changing the angle of kinking in TM3, consistent with the critical role identified for the kink-inducing proline 485 residue [56,61].

In our model (Figure 2, Step C), ATP hydrolysis would then cause any drug molecules in cavity 1 to be shuttled to cavity 2 by conformational changes in the leucine plug [48], and that other conformational changes result in the opening of the capping helical bundle (TM5b,5c,6a) to enable drug to be released from cavity 2. Notably, two residues in this 3-helical cap have been mutated (I573, L533 [56,57]) and both result in a severe folding and transport defect, consistent with a critical role for the cap. Subsequent release of ADP causes ABCG2 resetting (Figure 2, Step D).

Does the model account for drug substrates being transported but that inhibitors are not? We propose that only transported drug substrates enter cavity 1 via the access site, a step which is essential for the resulting power stroke linked to the opening of the leucine plug. This provides a mechanism for substrate selectivity: inhibitors that bind directly to cavity 1 fail to engage the power stroke mechanism described [48] and so are neither transported themselves and also inhibit ATP hydrolysis and substrate drug transport. The model also infers that cholesterol (which is tentatively observed in cavity 1) acts as a regulatory molecule maintaining a basal ATPase activity and ensuring that transport competent conformations of the protein are available [47,52,6264].

Two final features of the schematic model are worth highlighting. Firstly, ABCG2 transports chemicals of different hydrophobicity; the frequently used test substrates in in vitro assays include the highly hydrophobic mitoxantrone (soluble in DMSO) and the water-soluble Lucifer Yellow. The access site described above is hydrophobic and it may be that different access sites exist for more hydrophilic transport substrates. Secondly, the model in Figure 2 is an asymmetric functional model compared with the structural symmetry applied to the cryo-EM data [47,48]. Binding of transport substrate at either access site induces an asymmetry in both the TMDs and the NBDs which is then maintained in the coupled catalytic and transport cycles. This asymmetry is compatible with other functional models of ABC transporters, such as those described for TAP1/2 [65].

So, 2 years has brought us a huge way in terms of the structure and mechanism of ABCG2. Indeed, the recent cryo-EM data probably bring ABCG2 right up alongside its larger cousin ABCB1 regarding mechanistic understanding of these multidrug pumps. More structural data will doubtless emerge, hopefully in the presence of transport substrate at various stages of the catalytic cycle. Coupled with biochemical data, this will enable us to refine models and we can be confident that a more complete molecular description of substrate binding and transport by ABCG2 is within our grasp. How we then use that data for drug discovery is a whole new chapter in the lively and productive field of transporter research.

Abbreviations

     
  • ABC

    ATP-binding cassette

  •  
  • ABCP

    ATP-binding cassette transporter of the placenta

  •  
  • AD

    Alzheimer's disease

  •  
  • BCRP

    breast cancer resistance protein

  •  
  • CFTR

    cystic fibrosis transmembrane conductance regulator

  •  
  • ECD

    extracellular domain

  •  
  • EM

    electron microscopy

  •  
  • LTC

    leukotriene

  •  
  • MDR

    multidrug resistance

  •  
  • MRP

    multidrug resistance-associated protein

  •  
  • NBD

    nucleotide-binding domain

  •  
  • P-gp

    P-glycoprotein

  •  
  • SUR

    sulphonylurea receptor

  •  
  • TAP

    transporter associated with antigen presentation

  •  
  • TMD

    transmembrane domain

Author Contribution

All authors contributed equally to the conception of the article and to its writing and approval.

Funding

P.K. is supported by University of Nottingham Vice Chancellors Scholarships for Research Excellence. The work of the laboratory is supported by the Biotechnology and Biological Sciences Research Council Doctoral Training Programme grant [grant number BB/J014508/1].

Competing Interests

The Authors declare that there are no competing interests associated with the manuscript.

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