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Encyclopedia of Biological Chemistry - Vol_1

Encyclopedia of Biological Chemistry - Vol_1

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 ABC Transporters
 Andre´ Goffeau, Benoı ˆ  t De Hertogh and Philippe V. Baret
Universite´ Catholique de Louvain, Louvain-la-Neuve, Belgium
The ABC proteins constitute the largest family of proteins.They are present in all living species from
. They make up to 4% of the full genome complementof bacteria such as
Escherichia coli
Bacillus subtilis
. Eacheukaryote genome contains several dozens of members (over100intheplant
).Theyarerecognizedbya consensus ATP-binding region of approximately 100 aminoacids which include the two Walker A and B motifsencompassing a linker or C region (Figure 1). The ABCproteins catalyze a wide variety of physiological functions,most (but not all) of which being related to transport. Thisarticle describes the major physiological and biochemicalfunctions as well as the structural properties of some of thebest-known ABC transporters using examples from the yeast
Saccharomyces cerevisiae
Homo sapiens
Most, but not all, ABC proteins are ABC transporters.Eachofthosemoleculescontains,orisassociatedto,oneortwocytoplasmicATP-bindingdomainsnamednucleo-tide binding domains (NBDs) (Figure 1) and one or twotransmembranedomains(TMDs) (Figure 2).Each TMDcomprises usually six
-helix spans. Association of oneTMD to one NBD results in a half-size ABC transporter;however, they are believed to function as homo- orheterodimerssothattheminimalfunctionalorganizationof an ABC transporter is considered to be TMD–NBDTMD–NBD or NBD–TMD–NBD–TMD. In eukar-yotes, two TMDs and two NBDs are often associated inone single molecule called full-sized ABC transporter.The topological relation between NBD(s) and TMDs isvariable(Figure2).InbacteriatwoNBDsoftenassociatewithtwoTMDseitherasfoursinglesubunitsencodedbythe same operon or in various combinations of fusedsubunits. Association of other proteins may occur. Themost prominent associated bacterial protein is theperiplasmic solute-binding receptor, which in gram-negative bacteria is found in the periplasm, and ingram-positive bacteria is present often as a lipoprotein,boundtotheexternalmembranesurfaceviaelectrostaticinteractions(Figure3).ThethreedomainsofthebacterialABC uptake transporters: namely the periplasmicbinding receptor, the cytoplasmic NDB, and the mem-brane TMD are believed to have arisen from a commonancestral ABC transporter in which these three proteinswere already present. However, during evolution, thesequence of the periplasmic solute-binding receptorsdiverges more rapidly than that of the TMDs, while thatof NBDs is the least divergent. Thus, all NBDsare homologous, but this is not true for the TMDs orthe receptors. Nevertheless, the phylogenetic clusteringpatterns in bacterial ABC from different species aregenerally the same for all three types of proteins, despitetheir variable rate of evolution.The topology of some eukaryotic ABC effluxers canbe complex as additional TM spans occur in somesystems (Figure 3) as well as extra cytoplasmic domainsof presumed regulatory function.
The different families of ABC proteins transport a widevariety ofsubstrates againsttheirconcentration gradientusing the energy of ATP hydrolysis carried out by NBD.In bacteria, the transported substrates are eitherimported in or exported out of the cell. In eukaryotes,only extracytoplasmic exporters (transporting sub-strates either out of the cell or into organelles) areknown up to now. Within the ABC superfamily, 61phylogenetic families have been identified so far. Thesefamilies generally correlate with substrate specificity.Their classification based both on functional andphylogenetic criteria has been carried out within thetransporter classification (TC) system developed byMilton Saier in San Diego. The TC system has recentlybeen endorsed by The International Union of theBiochemical and Molecular Biology Societies. In theTC system, prokaryotic ABC influx porters comprise 22phylogenetic families including histidine permease, thefirst ABC transporter to be cloned and sequenced in thelaboratory of Giovanna Ames in 1982. Another famousexample is the MalEFGK operon classified in TC as amaltooligosaccharide porter within “the carbohydrateuptake transporter-1 (CUT1) family,” and given the TCdigit 3.A.1.1. In this operon, MalE is the receptor, MalF
Encyclopedia of Biological Chemistry, Volume 1.
2004, Elsevier Inc. All Rights Reserved.
and MalG are distinct TMD subunits, and MalK is adouble NBD.The prokaryotic effluxers comprise 27 familiesincluding the multidrug exporter LmrA from thegram-positive
Lactococcus lactis
well studied by WillKoning and belonging to “the drug exporter-2 family”(3.A.1.117).TheeukaryoticABCscanbegroupedinonly12effluxfamilies including the famous MDR1 also named Pgp(permease-glycoprotein), discovered in 1986 by IraPastan, Michael Gottesman and colleagues, and shownto be involved in MDR of chemiotreated tumor cells.In the TC system, this ABC exporter is classified in “themultidrug resistance exporter family” (3.A.1.201).The TC system is redundant with the HumanGenome Organization (HUGO) classification adoptedby the scientific community working on mammalianobjects (mouse or man). The 45 human or mouseABC proteins, comprising efflux transporters andnontransporter proteins, have been classified in sevenfamilies named ABCA to ABCG according to topologi-cal and phylogenetical criteria that are less stringentthan those used by TC. In its present form, the HUGOnomenclature and classification are difficult to use foridentification of novel ABC from non-mammalianspecies. For instance, the
Saccharomyces cerevisiae
genome contains 32 ABC genes among which 22 (16full-size and six small-size) are associated to transmem-brane domains in four different topologies. Its largestfamily is the full-sized Pdr5p-like family identified in1996 by Anabelle Decottignies and Andre ´Goffeau andshown later to be present in all fungi and plants. Thisfamily is not detected in the animal kingdom. Con-versely, the large human and mouse ABCA subfamily isnot represented in yeast genomes. There is a necessity toadopt a consistent classification system, which combinesthe TC and HUGO nomenclatures.
Function and Diseases
The immense variety of substrates transported inbacteria is reflected by the identification of 49 phyloge-netic ABC families including 22 influx protein com-plexes and 27 efflux transporter systems. As they belongto Archaea, gram-negative and gram-positive plasmamembranes that are widely different in organization andcomposition, the number and nature of proteinsassociated to given ABC transporters are variable andtheir transport mechanisms may be partly different. Inbacterial and Archaea ABC, the variety of substrates:sugars, amino acids, lipids, ions, polysaccharides,peptides,proteins,toxins,drugs,antibiotics,xenobioticsand other metabolites is reflected by the divergence of the periplasmic sensor and that of the TMD, which mustcontrol both specificity of substrate and part of thecoupling mechanism.Even if all eukaryotic ABC transporters are effluxersthat comprise subunits in which each TMD is fused to aNBD, some of them are not directly involved in movingsubstrates. For instance, in the cystic fibrosis transmem-brane regulator CFTR, and in the sulfonylurea receptorSUR, the hydrolysis of ATP appears to be linked to theregulationofopeningandclosingofionchannelscarriedby the ABC protein itself or other proteins (Figure 3B).The conservation of NBD in all ABC transporters,however, suggests that a basic coupling mechanismexists for efflux and influx whatever the transportedsubstrate. Moreover, distantly related proteins existwhich utilize an NBD to drive diverse nontransportprocesses such as DNA repair or protein-elongation orregulation of RNAse activities.The 32 yeast ABC proteins are in principle easy tostudy, as sensitive genetic tools are available. However,only a few successful cases of overexpressions and
The consensus ATP-binding region of a typical ABCprotein is made of approximately 100 amino acids (aa), including bothWalker A and B motifs and the linker C region.
Example of topological relations between NBDs andtransmembrane spans in full-sized and half-sized ABC transporters.
in vitro
UTPase or ATPase measurements of purifiedABCs have been reported. Only one purified yeast ABCtransporter, the pleiotropic drug resistance effluxerPdr5p, has been submitted to structural studies.The biochemical study of human ABC transporters isoften more advanced than that of the yeasts. The Pgpprotein responsible for multiple drug resistance (MDR)in human cells is especially well studied. One strongimpetusfor the study ofmammalianABC transportersistheir involvement in diseases. Many mendelian diseasesand complex genetic disorders are caused by ABCtransporters including cystic fibrosis, adrenoleuko-dystrophy, Stargardt disease, Tangier disease, immunedeficiencies, progressive familial intrahepatic cholesta-sis, Dublin–Johnson syndrome, Pseudoxanthoma elas-ticum, persistent hyperinsulinemic hypoglycemia of infancy due to focal adenomatous hyperplasia, X-linkedsideroblastosis and anemia, age-related maculardegeneration, familial hypoapoproteinemia, Fundusflavimaculatis, Retinitis pigmentosum, cone rod dystro-phy etc. Cell lines isolated from diseased tissues allowmolecular study of the involved ABC transporter.Moreover, a variety of drug-resistant cell lines isavailable from MDR or MDR-related protein (MRP)tissues. Basic studies of human ABC transporters wouldgreatly benefit from heterologous expression of humanABC transporter genes in yeast or other cells, but thistechnology is far from being satisfactory yet. Mean-while,knockouttechnologyin the mouse may be neededto begin to understand the molecular and physiologicalfunctions of the mammalian transporters.
Structure and Biochemical Mechanism
In 1998, the first high-resolution structure of a NBD,that from the histidine ABC importer HisP, wasreported. Five years later, about six related structureswere available and a consensus view emerged. NBDs areorganized as dimers and two molecules of ATP arebound at their interface. Each nucleotide-binding sitecomprises a Walker A motiffrom monomer 1 and the Cmotif from monomer 2. This results from a “head-to-tail” arrangement of the two interacting monomers.This is supported by biochemical arguments and iscoherent with the cooperative hydrolysis for ATPhydrolysis observed with MalK.More recently, three structures of complete dimericABC transporters comprising both NDB and TMD wereobtained: that of the presumed phospholipid flippaseMsbA from
E. coli
Vibrio cholera
and that of thevitamin B12 importer BtuCD from
E. coli
. Thestructures obtained were dissimilar, which may not betoo surprising taking into account the different con-ditions used, the different numbers of TM spans and thedifferent functions (import or export) of the proteinsanalyzed. No generalization can be made, for instance,on the angle between the TM spans and the membraneplane or on the identification of the interaction domainsbetween the TM spans. The nature of communicationbetween the NBD and TMD is variable and carried outeither through the long and complex so-called intra-cellulardomainnamed ICDinMsbA,orthroughashort
-shapedlinkerbetweenthetransmembranespans6and7inBtuCD.Thenature,thesize,theorientation,andthelocation of the so-called chamber (or water channel, orpore, or cone) presumed to be involved in substratebinding are also variable. No consensus interactionpoints between the NBDs (open orclosed conformation)were observed. Obviously, more structures are neededon several transporters carrying out similar functions,such as drug efflux, for instance, to clarify these issuesand to reach a consensus interpretation of the basicstructural elements involved in the transport and in thecoupling mechanism.In contrast, recent analyses at the electronmicroscopy level of a bacteria (BmrA or YccV from
Bacillus subtilis
) and a yeast (Pdr5p) drug efflux ABCtransporter came to a remarkably coherent set of conclusions. In both cases, the basic structural unitseems to comprise four joining NBDs (that correspondsto two full-size Pdr5p or four half-size BmrA), which are
Example of proteins associated to bacterial and mammalian ABC transporters.
OutMembraneInNBD1NBD2Periplasmic-bindingreceptorSubstrateTransmembrane domainCytoplasmic nucleotide-binding domainOutNInN-terminalextensionPoresubunitNCCNBD2NBD1