The family of membrane-localised ATP binding cassette transporters (ABC transporters) contains many members that are of great clinical and physiological significance. Homology within the family is weak at the level of primary structure, reflecting the diversity of functions that these transporters fulfil. This could represent a problem for structure-based modelling of ABC transporters that will be desirable as and when high resolution crystal structures become available. However, homology at the level of tertiary and quaternary structure is often found to be much higher than homology at the sequence level.
The comparison of the three-dimensional (3D) structures of a few ABC transporters is therefore of considerable interest, even at the medium resolutions offered by electron microscopy. Here we compare the 3D structures of two ABC transporters that have been obtained by electron microscopy and single particle image analysis. We also discuss a third transporter for which projection data alone have been obtained.
(i) P-glycoprotein (P-gp) is an ABC transporter that pumps hydrophobic drugs out of cells. In humans, it is associated with drug-resistant tumours and the failure of chemotherapy. The 3D structure of P-gp has been first obtained by electron microscopy coupled with single particle image analysis and later by analysis of 2D crystals (Higgins et al. 1997; Rosenberg et al. 1997). (ii) Multidrug resistance protein 1 (MRP1) is another ABC transporter that is linked to multidrug resistance and is also an efficient transporter of conjugated organic anions. Purified MRP1 has now also been studied by single particle image analysis and by electron crystallography (Rosenberg et al. 2001), yielding projection structures. (iii) TAP is a transporter associated with antigen presentation that is present in the endoplasmic reticulum (ER) membrane, and is responsible for translocation of peptides from the cytoplasm to the ER, which are then coupled to the major histocompatibility complex (MHC Class I). Unlike P-gp and MRP (which are single polypeptides), TAP consists of two homologous polypeptides, TAP1 and TAP2. TAP plays an important role in the immune response. A 3D structure for TAP has been obtained by single particle image analysis (Velarde et al. 2001).
A high degree of similarity exists for the tertiary structures of TAP and P-gp, which are characterised by a ~3 nm diameter pore or pocket on one side, which is encircled by a collar of protein of ~6-7 nm diameter. This region has been associated with the transmembrane domains (TMDs) and external loops. On the opposing side, two large globular domains appear to block the bottom of the pore. These domains have been assigned to the nucleotide binding domains (NBDs). The transporters have overall dimensions in the membrane of ~6 Ω 9 nm with a thickness estimated to be 6-7 nm. Given that the lipid bilayer is ~4 nm across, the observation of a ~6-7 nm thickness for ABC transporters implies that the globular NBDs (~4 nm diameter) will be at least partially associated with membrane-spanning regions (TMDs), with implications for the coupling of ATP hydrolysis to transport. The consistent observation of a ~3 nm diameter pore or pocket implies that this feature is probably important, although such a diameter is considerably larger than that required for passage of the substrates pumped by P-gp and TAP. All three transporters display some degree of pseudo-twofold symmetry, as would be expected for a heterodimeric protein complex (TAP) or a modular polypeptide composed of two homologous repeats (P-gp). For P-gp and TAP, a head-to-tail arrangement of the two homologous halves of the transporter is implied. In contrast to P-gp and TAP, MRP1 contains an extra transmembrane domain, hence a loss of the pseudo-twofold symmetry will be expected for the transmembrane region of the transporter. A 3D structure of MRP1 will be required in order to evaluate these differences.
The quaternary structure of the three transporters is the same in the detergent-solubilised state, i.e. monomeric (monomeric heterodimers for TAP). However, recent evidence obtained with 2D crystals of MRP suggests that this transporter might take up a different quaternary structure in the membrane (homodimeric). The crystals of MRP1, which are formed in reconstituted membranes, display p2 symmetry with two MRP1 monomers in the unit cell. It is possible that this arrangement is coincidental, although the dimer interface in the crystals appears to be substantial, being formed between one putative NBD from one MRP1 monomer and the same NBD in a second MRP1 monomer. Further reconstitution studies are required in order to fully explore this behaviour.
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