Homes for the orphans: utilization of multiple substrate-binding proteins by ABC transporters

Acquiring nutrients from the environment is essential for all microbes, and the ATP-binding cassette (ABC) transporters are one of the major routes by which bacteria achieve it. In this issue of Molecular Microbiology , Chen et al. describe their characterization of what appeared at first glance a simple ABC transporter for acquisition of quaternary ammonium compounds (QACs) in Pseudomonas sp., but their persistence in fully determining the properties of this system led to the experimental demonstration that QAC uptake utilizes three different substrate-binding proteins (SBPs), two of which are encoded at remote locations on the genome as 'orphan' SBPs that are each able to function with a single core ABC transporter. Building on the unusual nature of this system, in which multiple SBPs with non-overlapping substrate specificities compete for the same transporter binding site, they designed elegant in vivo experiments that suggest that only substrate-bound SBPs are able to form functional complexes with the membrane domains. This new finding provides an important piece of in vivo data leading to further insight into how this ubiquitous family of transporters operates.     

High-Throughput Screening of Dipeptide Utilization Mediated by the ABC Transporter DppBCDF and Its Substrate-Binding Proteins DppA1-A5 in Pseudomonas aeruginosa

In this study, we show that the dppBCDF operon of Pseudomonas aeruginosa PA14 encodes an ABC transporter responsible for the utilization of di/tripeptides. The substrate specificity of ABC transporters is determined by its associated substrate-binding proteins (SBPs). Whereas in E. coli only one protein, DppA, determines the specificity of the transporter, five orthologous SBPs, DppA1–A5 are present in P. aeruginosa. Multiple SBPs might broaden the substrate specificity by increasing the transporter capacity. We utilized the Biolog phenotype MicroArray technology to investigate utilization of di/tripeptides in mutants lacking either the transport machinery or all of the five SBPs. This high-throughput method enabled us to screen hundreds of dipeptides with various side-chains, and subsequently, to determine the substrate profile of the dipeptide permease. The substrate spectrum of the SBPs was elucidated by complementation of a penta mutant, deficient of all five SBPs, with plasmids carrying individual SBPs. It became apparent that some dipeptides were utilized with different affinity for each SBP. We found that DppA2 shows the highest flexibility on substrate recognition and that DppA2 and DppA4 have a higher tendency to utilize tripeptides. DppA5 was not able to complement the penta mutant under our screening conditions. Phaseolotoxin, a toxic tripeptide inhibiting the enzyme ornithine carbamoyltransferase, is also transported into P. aeruginosa via the DppBCDF permease. The SBP DppA1, and with much greater extend DppA3, are responsible for delivering the toxin to the permease. Our results provide a first overview of the substrate pattern of the ABC dipeptide transport machinery in P. aeruginosa.     

Structural and functional characterization of solute binding proteins for aromatic compounds derived from lignin: p‐Coumaric acid and related aromatic acids

Lignin comprises 15–25% of plant biomass and represents a major environmental carbon source for utilization by soil microorganisms. Access to this energy resource requires the action of fungal and bacterial enzymes to break down the lignin polymer into a complex assortment of aromatic compounds that can be transported into the cells. To improve our understanding of the utilization of lignin by microorganisms, we characterized the molecular properties of solute binding proteins of ATP‐binding cassette transporter proteins that interact with these compounds. A combination of functional screens and structural studies characterized the binding specificity of the solute binding proteins for aromatic compounds derived from lignin such as p‐coumarate, 3‐phenylpropionic acid and compounds with more complex ring substitutions. A ligand screen based on thermal stabilization identified several binding protein clusters that exhibit preferences based on the size or number of aromatic ring substituents. Multiple X‐ray crystal structures of protein–ligand complexes for these clusters identified the molecular basis of the binding specificity for the lignin‐derived aromatic compounds. The screens and structural data provide new functional assignments for these solute‐binding proteins which can be used to infer their transport specificity. This knowledge of the functional roles and molecular binding specificity of these proteins will support the identification of the specific enzymes and regulatory proteins of peripheral pathways that funnel these compounds to central metabolic pathways and will improve the predictive power of sequence‐based functional annotation methods for this family of proteins.Proteins 2013; 81:1709–1726. © 2013 Wiley Periodicals, Inc.     

Several Archaeal Homologs of Putative Oligopeptide-Binding Proteins Encoded by Thermotoga maritima Bind Sugars

The hyperthermophilic bacterium Thermotoga maritima has shared many genes with archaea through horizontal gene transfer. Several of these encode putative oligopeptide ATP binding cassette (ABC) transporters. We sought to test the hypothesis that these transporters actually transport sugars by measuring the substrate affinities of their encoded substrate-binding proteins (SBPs). This information will increase our understanding of the selective pressures that allowed this organism to retain these archaeal homologs. By measuring changes in intrinsic fluorescence of these SBPs in response to exposure to various sugars, we found that five of the eight proteins examined bind to sugars. We could not identify the ligands of the SBPs TM0460, TM1150, and TM1199. The ligands for the archaeal SBPs are TM0031 (BglE), the β-glucosides cellobiose and laminaribiose; TM0071 (XloE), xylobiose and xylotriose; TM0300 (GloE), large glucose oligosaccharides represented by xyloglucans; TM1223 (ManE), β-1,4-mannobiose; and TM1226 (ManD), β-1,4-mannobiose, β-1,4-mannotriose, β-1,4-mannotetraose, β-1,4-galactosyl mannobiose, and cellobiose. For comparison, seven bacterial putative sugar-binding proteins were examined and ligands for three (TM0595, TM0810, and TM1855) were not identified. The ligands for these bacterial SBPs are TM0114 (XylE), xylose; TM0418 (InoE), myo-inositol; TM0432 (AguE), α-1,4-digalactouronic acid; and TM0958 (RbsB), ribose. We found that T. maritima does not grow on several complex polypeptide mixtures as sole sources of carbon and nitrogen, so it is unlikely that these archaeal ABC transporters are used primarily for oligopeptide transport. Since these SBPs bind oligosaccharides with micromolar to nanomolar affinities, we propose that they are used primarily for oligosaccharide transport.     

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.     

Transcriptional Regulation,Metal Binding Properties and Structure of Pden1597, an Unusual Zinc Transport Protein from Paracoccus denitrificans

ATP-binding cassette (ABC) transporters of the cluster 9 family are ubiquitous among bacteria and essential for acquiring Zn²⁺ and Mn²⁺ from the environment or, in the case of pathogens, from the host. These rely on a substrate-binding protein (SBP) to coordinate the relevant metal with high affinity and specificity and subsequently release it to a membrane permease for translocation into the cytoplasm. Although a number of cluster 9 SBP structures have been determined, the structural attributes conferring Zn²⁺ or Mn²⁺ specificity remain ambiguous. Here we describe the gene expression profile, in vitro metal binding properties, and crystal structure of a new cluster 9 SBP from Paracoccus denitrificans we have called AztC. Although all of our results strongly indicate Zn²⁺ over Mn²⁺ specificity, the Zn²⁺ ion is coordinated by a conserved Asp residue only observed to date as a metal ligand in Mn²⁺-specific SBPs. The unusual sequence properties of this protein are shared among close homologues, including members from the human pathogens Klebsiella pneumonia and Enterobacter aerogenes, and would seem to suggest a subclass of Zn²⁺-specific transporters among the cluster 9 family. In any case, the unusual coordination environment of AztC expands the already considerable range of those available to Zn²⁺-specific SBPs and highlights the presence of a His-rich loop as the most reliable indicator of Zn²⁺ specificity.     

Biophysical and structural characterization of a sequence-diverse set of solute-binding proteins for aromatic compounds

Rhodopseudomonas palustris metabolizes aromatic compounds derived from lignin degradation products and has the potential for bioremediation of xenobiotic compounds. We recently identified four possible solute-binding proteins in R. palustris that demonstrated binding to aromatic lignin monomers. Characterization of these proteins in the absence and presence of the aromatic ligands will provide unprecedented insights into the specificity and mode of aromatic ligand binding in solute-binding proteins. Here, we report the thermodynamic and structural properties of the proteins with aromatic ligands using isothermal titration calorimetry, small/wide angle x-ray scattering, and theoretical predictions. The proteins exhibit high affinity for the aromatic substrates with dissociation constants in the low micromolar to nanomolar range. The global shapes of the proteins are characterized by flexible ellipsoid-like structures with maximum dimensions in the 80–90-Å range. The data demonstrate that the global shapes remained unaltered in the presence of the aromatic ligands. However, local structural changes were detected in the presence of some ligands, as judged by the observed features in the wide angle x-ray scattering regime at q ∼0.20–0.40 Å⁻¹. The theoretical models confirmed the elongated nature of the proteins and showed that they consist of two domains linked by a hinge. Evaluation of the protein-binding sites showed that the ligands were found in the hinge region and that ligand stabilization was primarily driven by hydrophobic interactions. Taken together, this study shows the capability of identifying solute-binding proteins that interact with lignin degradation products using high throughput genomic and biophysical approaches, which can be extended to other organisms.     

The CouPSTU and TarPQM Transporters in Rhodopseudomonas palustris: Redundant,Promiscuous Uptake Systems for Lignin-Derived Aromatic Substrates

The biodegradation of lignin, one of the most abundant carbon compounds on Earth, has important biotechnological applications in the derivation of useful products from lignocellulosic wastes. The purple photosynthetic bacterium Rhodopseudomonas palustris is able to grow photoheterotrophically under anaerobic conditions on a range of phenylpropeneoid lignin monomers, including coumarate, ferulate, caffeate, and cinnamate. RPA1789 (CouP) is the periplasmic binding-protein component of an ABC system (CouPSTU; RPA1789, RPA1791–1793), which has previously been implicated in the active transport of this class of aromatic substrate. Here, we show using both intrinsic tryptophan fluorescence and isothermal titration calorimetry that CouP binds a range of phenylpropeneoid ligands with K _d values in the nanomolar range. The crystal structure of CouP with ferulate as the bound ligand shows H-bond interactions between the 4-OH group of the aromatic ring with His309 and Gln305. H-bonds are also made between the carboxyl group on the ferulate side chain and Arg197, Ser222, and Thr102. An additional transport system (TarPQM; RPA1782–1784), a member of the tripartite ATP-independent periplasmic (TRAP) transporter family, is encoded at the same locus as rpa1789 and several other genes involved in coumarate metabolism. We show that the periplasmic binding-protein of this system (TarP; RPA1782) also binds coumarate, ferulate, caffeate, and cinnamate with nanomolar K _d values. Thus, we conclude that R. palustris uses two redundant but energetically distinct primary and secondary transporters that both employ high-affinity periplasmic binding-proteins to maximise the uptake of lignin-derived aromatic substrates from the environment. Our data provide a detailed thermodynamic and structural basis for understanding the interaction of lignin-derived aromatic substrates with proteins and will be of use in the further exploitation of the flexible metabolism of R. palustris for anaerobic aromatic biotransformations.     

Review. Structure and mechanism of ATP-binding cassette transporters

ATP-binding cassette (ABC) transporters constitute a large superfamily of integral membrane proteins that includes both importers and exporters. In recent years, several structures of complete ABC transporters have been determined by X-ray crystallography. These structures suggest a mechanism by which binding and hydrolysis of ATP by the cytoplasmic, nucleotide-binding domains control the conformation of the transmembrane domains and therefore which side of the membrane the translocation pathway is exposed to. A basic, conserved two-state mechanism can explain active transport of both ABC importers and ABC exporters, but various questions remain unresolved. In this article, I will review some of the crystal structures and the mechanistic insight gained from them. Future challenges for a better understanding of the mechanism of ABC transporters will be outlined.     

The crystal structure of the substrate-binding protein OpuBC from Bacillus subtilis in complex with choline

Bacillus subtilis can synthesize the compatible solute glycine betaine as an osmoprotectant from an exogenous supply of the precursor choline. Import of choline is mediated by two osmotically inducible ABC transport systems: OpuB and OpuC. OpuC catalyzes the import of various osmoprotectants, whereas OpuB is highly specific for choline. OpuBC is the substrate-binding protein of the OpuB transporter, and we have analyzed the affinity of the OpuBC/choline complex by intrinsic tryptophan fluorescence and determined a K_d value of about 30 μM. The X-ray crystal structure of the OpuBC/choline complex was solved at a resolution of 1.6 Å and revealed a fold typical of class II substrate-binding proteins. The positively charged trimethylammonium head group of choline is wedged into an aromatic cage formed by four tyrosine residues and is bound via cation-pi interactions. The hydroxyl group of choline protrudes out of this aromatic cage and makes a single interaction with residue Gln19. The substitution of this residue by Ala decreases choline binding affinity by approximately 15-fold. A water network stabilizes choline within its substrate-binding site and promotes indirect interactions between the two lobes of the OpuBC protein. Disruption of this intricate water network by site-directed mutagenesis of selected residues in OpuBC either strongly reduces choline binding affinity (between 18-fold and 25-fold) or abrogates ligand binding. The crystal structure of the OpuBC/choline complex provides a rational for the observed choline specificity of the OpuB ABC importer in vivo and explains its inability to catalyze the import of glycine betaine into osmotically stressed B. subtilis cells.     

Transport capabilities of environmental Pseudomonads for sulfur compounds

Sulfur is an essential element in plant rhizospheres and microbial activity plays a key role in increasing the biological availability of sulfur in soil environments. To better understand the mechanisms facilitating the exchange of sulfur‐containing molecules in soil, we profiled the binding specificities of eight previously uncharacterized ABC transporter solute‐binding proteins from plant‐associated Pseudomonads. A high‐throughput screening procedure indicated eighteen significant organosulfur binding ligands, with at least one high‐quality screening hit for each protein target. Calorimetric and spectroscopic methods were used to validate the best ligand assignments and catalog the thermodynamic properties of the protein‐ligand interactions. Two novel high‐affinity ligand‐binding activities were identified and quantified in this set of solute‐binding proteins. Bacteria were cultured in minimal media with screening library components supplied as the sole sulfur sources, demonstrating that these organosulfur compounds can be metabolized and confirming the relevance of ligand assignments. These results expand the set of experimentally validated ligands amenable to transport by this ABC transporter family and demonstrate the complex range of protein‐ligand interactions that can be accomplished by solute‐binding proteins. Characterizing new nutrient import pathways provides insight into Pseudomonad metabolic capabilities which can be used to further interrogate bacterial survival and participation in soil and rhizosphere communities.     

Identification of various substrate-binding proteins of the hyperthermophylic archaeon <Emphasis Type="Italic">Aeropyrum pernix</Emphasis> K1

Proteins from the extracellular medium of Aeropyrum pernix K1 were separated by two-dimensional electrophoresis and identified using mass spectrometry. Six different substrate-binding proteins (SBPs) from the ATP-binding cassette (ABC) transporter family were identified: (1) ABC transporter SBP (Q9YC61); (2) Branched-chain amino-acid ABC transporter, branched-chain amino-acid-binding protein (Q9YDJ6); (3) Oligopeptide ABC transporter, oligopeptide-binding protein (Q9YBL5); (4) Probable ABC transporter SBP (Q9Y9N4); (5) ABC transporter SBP (Q9YBG7); (6) ABC transporter SBP (Q9YFD7). Based on their orthology, division into the following classes was predicted: (1) multiple sugar-transport system SBPs; (2) peptide/nickel-transport system SBPs; and (3) branched-chain amino-acid-transport system SBPs. Further bioinformatic analyses showed that the identified SBPs differ in motif and in transmembrane-domain and signal-peptide organisation. Additionally, for all of these SBPs, sequence homology was found for archaeal proteins, and homologous proteins in bacteria were also found for the ABC transporter SBP Q9YBG7 and the ABC transporter SBP Q9YFD7. This is the first study, where different ABC SBPs from the extracellular medium of A. pernix have been identified using the combined methodology of two-dimensional electrophoresis and mass spectrometry.     

ABC transporters of the wheat pathogen<Emphasis Type="Italic"> Mycosphaerella graminicola</Emphasis> function as protectants against biotic and xenobiotic toxic compounds

We have studied the role of five ABC transporter genes (MgAtr to MgAtr5) from the wheat pathogen Mycosphaerella graminicola in multidrug resistance (MDR). Complementation of Saccharomyces cerevisiae mutants with the ABC transporter genes from M. graminicola showed that all the genes tested encode proteins that provide protection against chemically unrelated compounds, indicating that their products function as multidrug transporters with distinct but overlapping substrate specificities. Their substrate range in yeast includes fungicides, plant metabolites, antibiotics, and a mycotoxin derived from Fusarium graminearum (diacetoxyscirpenol). Transformants of M. graminicola in which individual ABC transporter genes were deleted or disrupted did not exhibit clear-cut phenotypes, probably due to the functional redundancy of transporters with overlapping substrate specificity. Independently generated MgAtr5 deletion mutants of M. graminicola showed an increase in sensitivity to the putative wheat defence compound resorcinol and to the grape phytoalexin resveratrol, suggesting a role for this transporter in protecting the fungus against plant defence compounds. Bioassays with antagonistic bacteria indicated that MgAtr2 provides protection against metabolites produced by Pseudomonas fluorescens and Burkholderia cepacia. In summary, our results show that ABC transporters from M. graminicola play a role in protection against toxic compounds of natural and artificial origin.     

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.     

PELDOR Spectroscopy Reveals Two Defined States of a Sialic Acid TRAP Transporter SBP in Solution

The tripartite ATP-independent periplasmic (TRAP) transporters are a widespread class of membrane transporters in bacteria and archaea. Typical substrates for TRAP transporters are organic acids including the sialic acid N-acetylneuraminic acid. The substrate binding proteins (SBP) of TRAP transporters are the best studied component and are responsible for initial high-affinity substrate binding. To better understand the dynamics of the ligand binding process, pulsed electron-electron double resonance (PELDOR, also known as DEER) spectroscopy was applied to study the conformational changes in the N-acetylneuraminic acid-specific SBP VcSiaP. The protein is the SBP of VcSiaPQM, a sialic acid TRAP transporter from Vibrio cholerae. Spin-labeled double-cysteine mutants of VcSiaP were analyzed in the substrate-bound and -free state and the measured distances were compared to available crystal structures. The data were compatible with two clear states only, which are consistent with the open and closed forms seen in TRAP SBP crystal structures. Substrate titration experiments demonstrated the transition of the population from one state to the other with no other observed forms. Mutants of key residues involved in ligand binding and/or proposed to be involved in domain closure were produced and the corresponding PELDOR experiments reveal important insights into the open-closed transition. The results are in excellent agreement with previous in vivo sialylation experiments. The structure of the spin-labeled Q54R1/L173R1 R125A mutant was solved at 2.1 Å resolution, revealing no significant changes in the protein structure. Thus, the loss of domain closure appears to be solely due to loss of binding. In conclusion, these data are consistent with TRAP SBPs undergoing a simple two-state transition from an open-unliganded to closed-liganded state during the transport cycle.     

ABC transporters: one,two or four extracytoplasmic substrate-binding sites?

Two families of ATP-binding cassette (ABC) transporters in which one or two extracytoplasmic substrate-binding domains are fused to either the N- or C-terminus of the translocator protein have been detected. This suggests that two, or even four, substrate-binding sites may function in the ABC transporter complex. This domain organization in ABC transporters, widely represented among microorganisms, raises new possibilities for how the substrate-binding protein(s) (SBPs) might interact with the translocator. One appealing hypothesis is that multiple substrate-binding sites in proximity to the entry site of the translocation pore enhance the transport capacity. We also discuss the implications of multiple substrate-binding sites in close proximity to the translocator in terms of broadened substrate specificity and possible cooperative interactions between SBPs and the translocator.     

Several archaeal homologs of putative oligopeptide-binding proteins encoded by Thermotoga maritima bind sugars

The hyperthermophilic bacterium Thermotoga maritima has shared many genes with archaea through horizontal gene transfer. Several of these encode putative oligopeptide ATP binding cassette (ABC) transporters. We sought to test the hypothesis that these transporters actually transport sugars by measuring the substrate affinities of their encoded substrate-binding proteins (SBPs). This information will increase our understanding of the selective pressures that allowed this organism to retain these archaeal homologs. By measuring changes in intrinsic fluorescence of these SBPs in response to exposure to various sugars, we found that five of the eight proteins examined bind to sugars. We could not identify the ligands of the SBPs TM0460, TM1150, and TM1199. The ligands for the archaeal SBPs are TM0031 (BglE), the beta-glucosides cellobiose and laminaribiose; TM0071 (XloE), xylobiose and xylotriose; TM0300 (GloE), large glucose oligosaccharides represented by xyloglucans; TM1223 (ManE), beta-1,4-mannobiose; and TM1226 (ManD), beta-1,4-mannobiose, beta-1,4-mannotriose, beta-1,4-mannotetraose, beta-1,4-galactosyl mannobiose, and cellobiose. For comparison, seven bacterial putative sugar-binding proteins were examined and ligands for three (TM0595, TM0810, and TM1855) were not identified. The ligands for these bacterial SBPs are TM0114 (XylE), xylose; TM0418 (InoE), myo-inositol; TM0432 (AguE), alpha-1,4-digalactouronic acid; and TM0958 (RbsB), ribose. We found that T. maritima does not grow on several complex polypeptide mixtures as sole sources of carbon and nitrogen, so it is unlikely that these archaeal ABC transporters are used primarily for oligopeptide transport. Since these SBPs bind oligosaccharides with micromolar to nanomolar affinities, we propose that they are used primarily for oligosaccharide transport.     

Structural and Biochemical Characterization of the Early and Late Enzymes in the Lignin β-Aryl Ether Cleavage Pathway from Sphingobium sp. SYK-6

There has been great progress in the development of technology for the conversion of lignocellulosic biomass to sugars and subsequent fermentation to fuels. However, plant lignin remains an untapped source of materials for production of fuels or high value chemicals. Biological cleavage of lignin has been well characterized in fungi, in which enzymes that create free radical intermediates are used to degrade this material. In contrast, a catabolic pathway for the stereospecific cleavage of β-aryl ether units that are found in lignin has been identified in Sphingobium sp. SYK-6 bacteria. β-Aryl ether units are typically abundant in lignin, corresponding to 50–70% of all of the intermonomer linkages. Consequently, a comprehensive understanding of enzymatic β-aryl ether (β-ether) cleavage is important for future efforts to biologically process lignin and its breakdown products. The crystal structures and biochemical characterization of the NAD-dependent dehydrogenases (LigD, LigO, and LigL) and the glutathione-dependent lyase LigG provide new insights into the early and late enzymes in the β-ether degradation pathway. We present detailed information on the cofactor and substrate binding sites and on the catalytic mechanisms of these enzymes, comparing them with other known members of their respective families. Information on the Lig enzymes provides new insight into their catalysis mechanisms and can inform future strategies for using aromatic oligomers derived from plant lignin as a source of valuable aromatic compounds for biofuels and other bioproducts.     

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.     

Sensing preferences for prokaryotic solute binding protein families

Solute binding proteins (SBPs) are of central physiological relevance for prokaryotes. These proteins present substrates to transporters, but they also stimulate different signal transduction receptors. SBPs form a superfamily of at least 33 protein Pfam families. To assess possible links between SBP sequence and the ligand recognized, we have inspected manually all SBP three-dimensional structures deposited in the protein data bank and retrieved 748 prokaryotic structures that have been solved in complex with bound ligand. These structures were classified into 26 SBP Pfam families. The analysis of the ligands recognized revealed that most families possess a preference for a compound class. There were three families each that bind preferentially saccharides and amino acids. In addition, we identified families that bind preferentially purines, quaternary amines, iron and iron-chelating compounds, oxoanions, bivalent metal ions or phosphates. Phylogenetic analyses suggest convergent evolutionary events that lead to families that bind the same ligand. The functional link between chemotaxis and compound uptake is reflected in similarities in the ligands recognized by SBPs and chemoreceptors. Associating Pfam families with ligand profiles will be of help to design experimental strategies aimed at the identification of ligands for uncharacterized SBPs.