Energy Coupling Factor-Type ABC Transporters for Vitamin Uptake in Prokaryotes

Energy coupling factor (ECF) transporters are a subgroup of ATP-binding cassette (ABC) transporters involved in the uptake of vitamins and micronutrients in prokaryotes. In contrast to classical ABC importers, ECF transporters do not make use of water-soluble substrate binding proteins or domains but instead employ integral membrane proteins for substrate binding (named S-components). S-components form active translocation complexes with the ECF module, an assembly of two nucleotide-binding domains (NBDs, or EcfA) and a second transmembrane protein. In some cases, the ECF module is dedicated to a single S-component, but in many cases, the ECF module can interact with several different S-components that are unrelated in sequence and bind diverse substrates. The modular organization with exchangeable S-components on a single ECF module allows the transport of chemically different substrates via a common route. The recent determination of the crystal structures of the S-components that recognize thiamin and riboflavin has provided a first clue about the mechanism of S-component exchange. This review describes recent advances and the current views of the mechanism of transport by ECF transporters.     

A novel class of modular transporters for vitamins in prokaryotes

The specific and tightly controlled transport of numerous nutrients and metabolites across cellular membranes is crucial to all forms of life. However, many of the transporter proteins involved have yet to be identified, including the vitamin transporters in various human pathogens, whose growth depends strictly on vitamin uptake. Comparative analysis of the ever-growing collection of microbial genomes coupled with experimental validation enables the discovery of such transporters. Here, we used this approach to discover an abundant class of vitamin transporters in prokaryotes with an unprecedented architecture. These transporters have energy-coupling modules comprised of a conserved transmembrane protein and two nucleotide binding proteins similar to those of ATP binding cassette (ABC) transporters, but unlike ABC transporters, they use small integral membrane proteins to capture specific substrates. We identified 21 families of these substrate capture proteins, each with a different specificity predicted by genome context analyses. Roughly half of the substrate capture proteins (335 cases) have a dedicated energizing module, but in 459 cases distributed among almost 100 gram-positive bacteria, including numerous human pathogens, different and unrelated substrate capture proteins share the same energy-coupling module. The shared use of energy-coupling modules was experimentally confirmed for folate, thiamine, and riboflavin transporters. We propose the name energy-coupling factor transporters for the new class of membrane transporters.     

ATP binding cassette systems: structures, mechanisms, and functions

ATP-binding cassette (ABC) systems are found in all three domains of life and in some giant viruses and form one of the largest protein superfamilies. Most family members are transport proteins that couple the free energy of ATP hydrolysis to the translocation of solutes across a biological membrane. The energizing module is also used to drive non-transport processes associated, e.g., with DNA repair and protein translation. Many ABC proteins are of considerable medical importance. In humans, dysfunction of at least eighteen out of 49 ABC transporters is associated with disease, such as cystic fibrosis, Tangier disease, adrenoleukodystrophy or Stargardt’s macular degeneration. In prokaryotes, ABC proteins confer resistance to antibiotics, secrete virulence factors and envelope components, or mediate the uptake of a large variety of nutrients. Canonical ABC transporters share a common structural organization comprising two transmembrane domains (TMDs) that form the translocation pore and two nucleotide-binding domains (NBDs) that bind and hydrolyze ATP. In this Mini-Review, we summarize recent structural and biochemical data obtained from both prokaryotic and eukaryotic model systems.     

The yeast plasma membrane P4-ATPases are major transporters for lysophospholipids

The transbilayer movement of phospholipids plays an essential role in establishing and maintaining the asymmetric distribution of lipids in biological membranes. The P4-ATPase family has been implicated as the major transporters of the aminoglycerophospholipids in both surface and endomembrane systems. Historically, fluorescent lipid analogs have been used to monitor the lipid transport activity of the P4-ATPases. Recent evidence now demonstrates that lyso-phosphatidylethanolamine (lyso-PtdEtn) and lyso-phosphatidylcholine (lyso-PtdCho) are bona fide biological substrates transported by the yeast plasma membrane ATPases, Dnf1p and Dnf2p, in consort with a second protein Lem3p. Subsequent to transport, the lysophospholipids are acylated by the enzyme Ale1p to produce PtdEtn and PtdCho. The transport of the lysophospholipids occurs at rates sufficient to support all the PtdEtn and PtdCho synthesis required for rapid cell growth. The lysophospholipid transporters also utilize the anti-neoplastic and anti-parasitic ether lipid substrates related to edelfosine. The identification of biological substrates for the plasma membrane ATPases coupled with the power of yeast genetics now provides new tools to dissect the structure and function of the aminoglycerophospholipid transporters.     

Getting In or Out: Early Segregation Between Importers and Exporters in the Evolution of ATP-Binding Cassette (ABC) Transporters

ATP-binding cassette (ABC) systems, also called traffic ATPases, are found in eukaryotes and prokaryotes and almost all participate in the transport of a wide variety of molecules. ABC systems are characterized by a highly conserved ATPase module called here the ABC module, involved in coupling transport to ATP hydrolysis. We have used the sequence of one of the first representatives of bacterial ABC transporters, the MalK protein, to collect 250 closely related sequences from a nonredundant protein sequence database. The sequences collected by this objective method are all known or putative ABC transporters. After having eliminated short protein sequences and duplicates, the 197 remaining sequences were subjected to a phylogenetic analysis based on a mutational similarity matrix. An unrooted tree for these modules was found to display two major branches, one grouping all collected uptake systems and the other all collected export systems. This remarkable disposition strongly suggests that the divergence between these two functionally different types of ABC systems occurred once in the history of these systems and probably before the differentiation of prokaryotes and eukaryotes. We discuss the implications of this finding and we propose a model accounting for the generation and the diversification of ABC systems. Received: 23 February 1997 / Accepted: 7 April 1998     

Transport of nickel and cobalt ions into bacterial cells by S components of ECF transporters

Energy-coupling factor (ECF) transporters form a distinct group of ABC-type micronutrient importers in prokaryotes that do not contain extracytoplasmic, soluble substrate-binding proteins. Instead, they consist of a transmembrane substrate-specific S component that interacts with a module composed of a moderately conserved transmembrane (T) component and ABC ATPases. The majority of S components is considered to act as high-affinity binding proteins that strictly depend on their cognate T and ATPase units for transport activity. For a fraction of biotin-specific S units, however, transport activity was demonstrated in their solitary state. Here, we compared the activities of nickel- and cobalt-specific ECF transporters in the presence and absence of their T and ATPase units. Accumulation assays with radioactive metal ions showed that the truncated transporters led to approx. 25 % of cell-bound radioactivity compared to the holotransporters. Activity of urease, an intracellular nickel-dependent enzyme, was used as a reporter and clearly indicated that the cell-bound radioactivity correlates with the cytoplasmic metal concentration. The results demonstrate that S units of metal transporters not only bind their substrates on the cell surface but mediate transport across the membrane, a finding of general importance on the way to understand the mechanism of ECF transporters.     

Lipid transport by mammalian ABC proteins

ABC (ATP-binding cassette) proteins actively transport a wide variety of substrates, including peptides, amino acids, sugars, metals, drugs, vitamins and lipids, across extracellular and intracellular membranes. Of the 49 hum an ABC proteins, a significant number are known to mediate the extrusion of lipids from membranes or the flipping of membrane lipids across the bilayer to generate and maintain membrane lipid asymmetry. Typical lipid substrates include phospholipids, sterols, sphingolipids, bile acids and related lipid conjugates. Members of the ABCA subfamily of ABC transporters and other ABC proteins such as ABCB4, ABCG1 and ABCG5/8 implicated in lipid transport play important roles in diverse biological processes such as cell signalling, membrane lipid asymmetry, removal of potentially toxic compounds and metabolites, and apoptosis. The importance of these ABC lipid transporters in cell physiology is evident from the finding that mutations in the genes encoding many of these proteins are responsible for severe inherited diseases. For example, mutations in ABCA1 cause Tangier disease associated with defective efflux of cholesterol and phosphatidylcholine from the plasma membrane to the lipid acceptor protein apoA1 (apolipoprotein AI), mutations in ABCA3 cause neonatal surfactant deficiency associated with a loss in secretion of the lipid pulmonary surfactants from lungs of newborns, mutations in ABCA4 cause Stargardt macular degeneration, a retinal degenerative disease linked to the reduced clearance of retinoid compounds from photoreceptor cells, mutations in ABCA12 cause harlequin and lamellar ichthyosis, skin diseases associated with defective lipid trafficking in keratinocytes, and mutations in ABCB4 and ABCG5/ABCG8 are responsible for progressive intrafamilial hepatic disease and sitosterolaemia associated with defective phospholipid and sterol transport respectively. This chapter highlights the involvement of various mammalian ABC transporters in lipid transport in the context of their role in cell signalling, cellular homoeostasis, apoptosis and inherited disorders.     

Multidrug resistance ABC transporters

Clinical multidrug resistance is caused by a group of integral membrane proteins that transport hydrophobic drugs and lipids across the cell membrane. One class of these permeases, known as multidrug resistance ATP binding cassette (ABC) transporters, translocate these molecules by coupling drug/lipid efflux with energy derived from the hydrolysis of ATP. In this review, we examine both the structures and conformational changes of multidrug resistance ABC transporters. Together with the available biochemical and structural evidence, we propose a general mechanism for hydrophobic substrate transport coupled to ATP hydrolysis.     

ABC转运蛋白结构及在植物病原真菌中的功能研究进展

ABC (ATP-binding cassette)转运蛋白是最大的膜转运蛋白超家族之一,其主要功能是利用ATP水解产生的能量将底物进行逆浓度梯度运输.所有生物体都含有大量ABC蛋白. ABC蛋白位于细胞的不同空间,如细胞膜、液泡、线粒体和过氧化物酶体.通常,ABC转运蛋白由跨膜结构域(TMD)和核苷酸结合结构域(NBD)组成,分别与底物和ATP结合.NBD执行与ATP结合和水解,是ABC转运蛋白的动力引擎,TMD识别特异性配体.大多数ABC转运蛋白最初是通过研究生物体耐药性而被发现的,包括多效耐药(PDR)和多药耐药(MDR).本文对ABC转运蛋白的结构及作用机制,以及植物病原真菌中ABC转运蛋白功能的研究进展进行综述.     

ABC A-subfamily transporters: structure, function and disease

ABC transporters constitute a family of evolutionarily highly conserved multispan proteins that mediate the translocation of defined substrates across membrane barriers. Evidence has accumulated during the past years to suggest that a subgroup of 12 structurally related "full-size" transporters, referred to as ABC A-subfamily transporters, mediates the transport of a variety of physiologic lipid compounds. The emerging importance of ABC A-transporters in human disease is reflected by the fact that as yet four members of this protein family (ABCA1, ABCA3, ABCR/ABCA4, ABCA12) have been causatively linked to completely unrelated groups of monogenetic disorders including familial high-density lipoprotein (HDL) deficiency, neonatal surfactant deficiency, degenerative retinopathies and congenital keratinization disorders. Although the biological function of the remaining 8 ABC A-transporters currently awaits clarification, they represent promising candidate genes for a presumably equally heterogenous group of Mendelian diseases associated with perturbed cellular lipid transport. This review summarizes our current knowledge on the role of ABC A-subfamily transporters in physiology and disease and explores clinical entities which may be potentially associated with dysfunctional members of this gene subfamily.     

Polyamines and membrane transporters

In recent years, our understanding of the importance of membrane transporters (MTs) in the disposition of and response to drugs has increased significantly. MTs are proteins that regulate the transport of endogenous molecules and xenobiotics across the cell membrane. In mammals, two super-families have been identified: ATP-binding cassette (ABC) and solute carrier (SLC) transporters. There is evidence that MTs might mediate polyamines (PA) transport. PA are ubiquitous polycations which are found in all living cells. In mammalian cells, three major PA are synthesised: putrescine, spermidine and spermine; whilst the decarboxylated arginine (agmatine) is not produced by mammals but is synthesised by plants and bacteria. In addition, research in the PA field suggests that PA are transported into cells via a specific transporter, the polyamine transport system(s) (PTS). Although the PTS has not been fully defined, there is evidence that some of the known MTs might be involved in PA transport. In this mini review, eight SLC transporters will be reviewed and their potential to mediate PA transport in human cells discussed. These transporters are SLC22A1, SLC22A2, SLC22A3, SLC47A1, SLC7A1, SLC3A2, SLC12A8A, and SLC22A16. Preliminary data from our laboratory have revealed that SLC22A1 might be involved in the PA uptake; in addition to one member of ABC superfamily (MDR1 protein) might also mediate the efflux of polyamine like molecules.     

ABC transporters: bacterial exporters.

The ABC transporters (also called traffic ATPases) make up a large superfamily of proteins which share a common function and a common ATP-binding domain. ABC transporters are classified into three major groups: bacterial importers (the periplasmic permeases), eukaryotic transporters, and bacterial exporters. We present a comprehensive review of the bacterial ABC exporter group, which currently includes over 40 systems. The bacterial ABC exporter systems are functionally subdivided on the basis of the type of substrate that each translocates. We describe three main groups: protein exporters, peptide exporters, and systems that transport nonprotein substrates. Prototype exporters from each group are described in detail to illustrate our current understanding of this protein family. The prototype systems include the alpha-hemolysin, colicin V, and capsular polysaccharide exporters from Escherichia coli, the protease exporter from Erwinia chrysanthemi, and the glucan exporters from Agrobacterium tumefaciens and Rhizobium meliloti. Phylogenetic analysis of the ATP-binding domains from 29 bacterial ABC exporters indicates that the bacterial ABC exporters can be divided into two primary branches. One branch contains the transport systems where the ATP-binding domain and the membrane-spanning domain are present on the same polypeptide, and the other branch contains the systems where these domains are found on separate polypeptides. Differences in substrate specificity do not correlate with evolutionary relatedness. A complete survey of the known and putative bacterial ABC exporters is included at the end of the review.     

Solitary BioY Proteins Mediate Biotin Transport into Recombinant Escherichia coli

Energy-coupling factor (ECF) transporters form a large group of vitamin uptake systems in prokaryotes. They are composed of highly diverse, substrate-specific, transmembrane proteins (S units), a ubiquitous transmembrane protein (T unit), and homo- or hetero-oligomeric ABC ATPases. Biotin transporters represent a special case of ECF-type systems. The majority of the biotin-specific S units (BioY) is known or predicted to interact with T units and ABC ATPases. About one-third of BioY proteins, however, are encoded in organisms lacking any recognizable T unit. This finding raises the question of whether these BioYs function as transporters in a solitary state, a feature ascribed to certain BioYs in the past. To address this question in living cells, an Escherichia coli K-12 derivative deficient in biotin synthesis and devoid of its endogenous high-affinity biotin transporter was constructed as a reference strain. This organism is particularly suited for this purpose because components of ECF transporters do not naturally occur in E. coli K-12. The double mutant was viable in media containing either high levels of biotin or a precursor of the downstream biosynthetic path. Importantly, it was nonviable on trace levels of biotin. Eight solitary bioY genes of proteobacterial origin were individually expressed in the reference strain. Each of the BioYs conferred biotin uptake activity on the recombinants, which was inferred from uptake assays with [³H]biotin and growth of the cells on trace levels of biotin. The results underscore that solitary BioY transports biotin across the cytoplasmic membrane.     

ATP-binding cassette transporters in Escherichia coli

ATP-binding cassette (ABC) transporters are integral membrane proteins that actively transport molecules across cell membranes. In Escherichia coli they consist primarily of import systems that involve in addition to the ABC transporter itself a substrate binding protein and outer membrane receptors or porins, and a number of transporters with varied functions. Recent crystal structures of a number of ATPase domains, substrate binding proteins, and full-length transporters have given new insight in the molecular basis of transport. Bioinformatics approaches allow an approximate identification of all ABC transporters in E. coli and their relation to other known transporters. Computational approaches involving modeling and simulation are beginning to yield insight into the dynamics of the transporters. We summarize the function of the known ABC transporters in E. coli and mechanistic insights from structural and computational studies.     

Emerging new paradigms for ABCG transporters

Every cell is separated from its external environment by a lipid membrane. Survival depends on the regulated and selective transport of nutrients, waste products and regulatory molecules across these membranes, a process that is often mediated by integral membrane proteins. The largest and most diverse of these membrane transport systems is the ATP binding cassette (ABC) family of membrane transport proteins. The ABC family is a large evolutionary conserved family of transmembrane proteins (> 250 members) present in all phyla, from bacteria to Homo sapiens, which require energy in the form of ATP hydrolysis to transport substrates against concentration gradients. In prokaryotes the majority of ABC transporters are involved in the transport of nutrients and other macromolecules into the cell. In eukaryotes, with the exception of the cystic fibrosis transmembrane conductance regulator (CFTR/ABCC7), ABC transporters mobilize substrates from the cytoplasm out of the cell or into specific intracellular organelles. This review focuses on the members of the ABCG subfamily of transporters, which are conserved through evolution in multiple taxa. As discussed below, these proteins participate in multiple cellular homeostatic processes, and functional mutations in some of them have clinical relevance in humans.     

The human ATP-binding cassette transporter genes: from the bench to the bedside

ATP-binding cassette (ABC) transporter genes are ubiquitously present in most organisms from bacteria to man. This gene family is the largest one known as of yet. Still growing, the number of human ABC transporters counts currently 47 members which belong to seven subfamilies. ABC transporters share a similar molecular architecture: (1) Full-structured transporters harbor two symmetric halves each consisting of one nucleotide binding domain (NBD) and one transmembrane domain (TMD). (2) Half-transporters with one NBD and one TMD homo- or heterodimerize to functional transporter complexes. ABC transporters are "traffic ATPases" which hydrolyze ATP and which transport a wide array of molecules or conduct the transport of molecules by stimulating other translocation mechanisms. Many ABC transporters are involved in human inherited or sporadic diseases such as cystic fibrosis, adrenoleukodystrophy, Stargardt's disease, drug-resistant tumors, Dubin-Johnson syndrome, Byler's disease, progressive familiar intrahepatic cholestasis, X-linked sideroblastic anemia and ataxia, persistent hyperinsulimenic hypoglycemia of infancy, and others. The present review summarizes the current findings in basic research and the efforts for bridging the gap to clinical applications in therapy and diagnostics.     

Diversity of transport mechanisms: common structural principles

Traditionally, prokaryotic solute transport systems are classified into major groups based on the energetic requirement of the transport process. These include the secondary transporters that are driven by a proton or sodium motive force, and the ATP-binding cassette (ABC) primary transporters, which use the hydrolysis of ATP to fuel transport. These transporters are specified by entirely different architectures of polypeptides. Recently, transport systems have been discovered that are composed of combinations of distinct functional modules of both secondary and ABC transporters. These findings indicate that during evolution the combination of integral membrane transport proteins with either a periplasmic solute-binding protein or a cytosolic ATPase, or both, have resulted in distinct classes of transporters with unique architectures and properties.     

ATP-binding cassette transporters in Escherichia coli

ATP-binding cassette (ABC) transporters are integral membrane proteins that actively transport molecules across cell membranes. In Escherichia coli they consist primarily of import systems that involve in addition to the ABC transporter itself a substrate binding protein and outer membrane receptors or porins, and a number of transporters with varied functions. Recent crystal structures of a number of ATPase domains, substrate binding proteins, and full-length transporters have given new insight in the molecular basis of transport. Bioinformatics approaches allow an approximate identification of all ABC transporters in E. coli and their relation to other known transporters. Computational approaches involving modeling and simulation are beginning to yield insight into the dynamics of the transporters. We summarize the function of the known ABC transporters in E. coli and mechanistic insights from structural and computational studies.     

Peroxisomal ABC transporters: Structure, function and role in disease

ATP-binding cassette (ABC) transporters belong to one of the largest families of membrane proteins, and are present in almost all living organisms from eubacteria to mammals. They exist on plasma membranes and intracellular compartments such as the mitochondria, peroxisomes, endoplasmic reticulum, Golgi apparatus and lysosomes, and mediate the active transport of a wide variety of substrates in a variety of different cellular processes. These include the transport of amino acids, polysaccharides, peptides, lipids and xenobiotics, including drugs and toxins. Three ABC transporters belonging to subfamily D have been identified in mammalian peroxisomes. The ABC transporters are half-size and assemble mostly as a homodimer after posttranslational transport to peroxisomal membranes. ABCD1/ALDP and ABCD2/ALDRP are suggested to be involved in the transport of very long chain acyl-CoA with differences in substrate specificity, and ABCD3/PMP70 is involved in the transport of long and branched chain acyl-CoA. ABCD1 is known to be responsible for X-linked adrenoleukodystrophy (X-ALD), an inborn error of peroxisomal β-oxidation of very long chain fatty acids. Here, we summarize recent advances and important points in our advancing understanding of how these ABC transporters target and assemble to peroxisomal membranes and perform their functions in physiological and pathological processes, including the neurodegenerative disease, X-ALD. This article is part of a Special Issue entitled: Metabolic Functions and Biogenesis of Peroxisomes in Health and Disease.     

Structure and mechanism of ABC transporter proteins

ATP-binding cassette (ABC) transporters are ubiquitous membrane proteins that couple the transport of diverse substrates across cellular membranes to the hydrolysis of ATP. The crystal structures of four ABC transporters have recently been determined. They reveal similar arrangements of the conserved ATP-hydrolyzing nucleotide-binding domains, but unrelated architectures of the transmembrane domains, with the notable exception of a common 'coupling helix' that is essential for transmitting conformational changes. The structures suggest a mechanism that rationalizes ATP-driven transport: While binding of ATP appears to trigger an outward-facing conformation, dissociation of the hydrolysis products may promote an inward-facing conformation. This basic scheme can, in principle, explain nutrient import by ABC importers and drug extrusion by ABC exporters.