Sample records for abc transporter superfamily

  1. [The ABC transporters of Saccharomyces cerevisiae].

    PubMed

    Wawrzycka, Donata

    2011-01-01

    The ABC transporters (ATP Binding Cassette) compose one of the bigest protein family with the great medical, industrial and economical impact. They are found in all organism from bacteria to man. ABC proteins are responsible for resistance of microorganism to antibiotics and fungicides and multidrug resistance of cancer cells. Mutations in ABC transporters genes cause seriuos deseases like cystic fibrosis, adrenoleucodystrophy or ataxia. Transport catalized by ABC proteins is charged with energy from the ATP hydrolysis. The ABC superfamily contains transporters, canals, receptors. Analysis of the Saccharomyces cerevisiae genome allowed to distinguish 30 potential ABC proteins which are classified into 6 subfamilies. The structural and functional similarity of the yeast and human ABC proteins allowes to use the S. cerevisiae as a model organism for ABC transporters characterisation. In this work the present state of knowleadge on yeast S. cerevisiae ABC proteins was summarised.

  2. Catalytic and transport cycles of ABC exporters.

    PubMed

    Al-Shawi, Marwan K

    2011-09-07

    ABC (ATP-binding cassette) transporters are arguably the most important family of ATP-driven transporters in biology. Despite considerable effort and advances in determining the structures and physiology of these transporters, their fundamental molecular mechanisms remain elusive and highly controversial. How does ATP hydrolysis by ABC transporters drive their transport function? Part of the problem in answering this question appears to be a perceived need to formulate a universal mechanism. Although it has been generally hoped and assumed that the whole superfamily of ABC transporters would exhibit similar conserved mechanisms, this is proving not to be the case. Structural considerations alone suggest that there are three overall types of coupling mechanisms related to ABC exporters, small ABC importers and large ABC importers. Biochemical and biophysical characterization leads us to the conclusion that, even within these three classes, the catalytic and transport mechanisms are not fully conserved, but continue to evolve. ABC transporters also exhibit unusual characteristics not observed in other primary transporters, such as uncoupled basal ATPase activity, that severely complicate mechanistic studies by established methods. In this chapter, I review these issues as related to ABC exporters in particular. A consensus view has emerged that ABC exporters follow alternating-access switch transport mechanisms. However, some biochemical data suggest that alternating catalytic site transport mechanisms are more appropriate for fully symmetrical ABC exporters. Heterodimeric and asymmetrical ABC exporters appear to conform to simple alternating-access-type mechanisms.

  3. Evolutionary Trajectories of Entomopathogenic Fungi ABC Transporters.

    PubMed

    Baral, Bikash

    2017-01-01

    The ABC protein superfamily-also called traffic ATPases-are energy-dependent ubiquitous proteins, representing one of the crucial and the largest family in the fungal genomes. The ATP-binding cassette endows a characteristic 200-250 amino acids and is omnipresent in all organisms ranging from prokaryotes to eukaryotes. Unlike in bacteria with nutrient import functions, ABC transporters in fungal entomopathogens serve as effective efflux pumps that are largely involved in the shuttle of metabolites across the biological membranes. Thus, the search for ABC proteins may prove of immense importance in elucidating the functional and molecular mechanism at the host-pathogen (insect-fungus) interface. Their sequence homology, domain topology, and functional traits led to the actual identification of nine different families in fungal entomopathogens. Evolutionary relationships within the ABC superfamily are discussed, concentrating on computational approaches for comparative identification of ABC transporters in insect-pathogenic fungi (entomopathogens) with those of animals, plants, and their bacterial orthologs. Ancestors of some fungal candidates have duplicated extensively in some phyla, while others were lost in one lineage or the other, and predictions for the cause of their duplications and/or loss in some phyla are made. ABC transporters of fungal insect-pathogens serve both defensive and offensive functions effective against land-dwelling and ground foraging voracious insects. This study may help to unravel the molecular cascades of ABC proteins to illuminate the means through which insects cope with fungal infection and fungal-related diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Transcriptome-Based Identification of ABC Transporters in the Western Tarnished Plant Bug Lygus hesperus

    PubMed Central

    Hull, J. Joe; Chaney, Kendrick; Geib, Scott M.; Fabrick, Jeffrey A.; Brent, Colin S.; Walsh, Douglas; Lavine, Laura Corley

    2014-01-01

    ATP-binding cassette (ABC) transporters are a large superfamily of proteins that mediate diverse physiological functions by coupling ATP hydrolysis with substrate transport across lipid membranes. In insects, these proteins play roles in metabolism, development, eye pigmentation, and xenobiotic clearance. While ABC transporters have been extensively studied in vertebrates, less is known concerning this superfamily in insects, particularly hemipteran pests. We used RNA-Seq transcriptome sequencing to identify 65 putative ABC transporter sequences (including 36 full-length sequences) from the eight ABC subfamilies in the western tarnished plant bug (Lygus hesperus), a polyphagous agricultural pest. Phylogenetic analyses revealed clear orthologous relationships with ABC transporters linked to insecticide/xenobiotic clearance and indicated lineage specific expansion of the L. hesperus ABCG and ABCH subfamilies. The transcriptional profile of 13 LhABCs representative of the ABCA, ABCB, ABCC, ABCG, and ABCH subfamilies was examined across L. hesperus development and within sex-specific adult tissues. All of the transcripts were amplified from both reproductively immature and mature adults and all but LhABCA8 were expressed to some degree in eggs. Expression of LhABCA8 was spatially localized to the testis and temporally timed with male reproductive development, suggesting a potential role in sexual maturation and/or spermatozoa protection. Elevated expression of LhABCC5 in Malpighian tubules suggests a possible role in xenobiotic clearance. Our results provide the first transcriptome-wide analysis of ABC transporters in an agriculturally important hemipteran pest and, because ABC transporters are known to be important mediators of insecticidal resistance, will provide the basis for future biochemical and toxicological studies on the role of this protein family in insecticide resistance in Lygus species. PMID:25401762

  5. Diversity of ABC transporter genes across the plant kingdom and their potential utility in biotechnology.

    PubMed

    Lane, Thomas S; Rempe, Caroline S; Davitt, Jack; Staton, Margaret E; Peng, Yanhui; Soltis, Douglas Edward; Melkonian, Michael; Deyholos, Michael; Leebens-Mack, James H; Chase, Mark; Rothfels, Carl J; Stevenson, Dennis; Graham, Sean W; Yu, Jun; Liu, Tao; Pires, J Chris; Edger, Patrick P; Zhang, Yong; Xie, Yinlong; Zhu, Ying; Carpenter, Eric; Wong, Gane Ka-Shu; Stewart, C Neal

    2016-05-31

    The ATP-binding cassette (ABC) transporter gene superfamily is ubiquitous among extant organisms and prominently represented in plants. ABC transporters act to transport compounds across cellular membranes and are involved in a diverse range of biological processes. Thus, the applicability to biotechnology is vast, including cancer resistance in humans, drug resistance among vertebrates, and herbicide and other xenobiotic resistance in plants. In addition, plants appear to harbor the highest diversity of ABC transporter genes compared with any other group of organisms. This study applied transcriptome analysis to survey the kingdom-wide ABC transporter diversity in plants and suggest biotechnology applications of this diversity. We utilized sequence similarity-based informatics techniques to infer the identity of ABC transporter gene candidates from 1295 phylogenetically-diverse plant transcriptomes. A total of 97,149 putative (approximately 25 % were full-length) ABC transporter gene members were identified; each RNA-Seq library (plant sample) had 88 ± 30 gene members. As expected, simpler organisms, such as algae, had fewer unique members than vascular land plants. Differences were also noted in the richness of certain ABC transporter subfamilies. Land plants had more unique ABCB, ABCC, and ABCG transporter gene members on average (p < 0.005), and green algae, red algae, and bryophytes had significantly more ABCF transporter gene members (p < 0.005). Ferns had significantly fewer ABCA transporter gene members than all other plant groups (p < 0.005). We present a transcriptomic overview of ABC transporter gene members across all major plant groups. An increase in the number of gene family members present in the ABCB, ABCC, and ABCD transporter subfamilies may indicate an expansion of the ABC transporter superfamily among green land plants, which include all crop species. The striking difference between the number of ABCA subfamily transporter

  6. Transcriptome-based identification of ABC transporters in the western tarnished plant bug lygus hesperus

    USDA-ARS?s Scientific Manuscript database

    ATP-binding cassette (ABC) transporters are a large superfamily of proteins that mediate diverse physiological functions by coupling ATP hydrolysis with substrate transport across lipid membranes. In insects, these proteins play roles in metabolism, development, eye pigmentation, and xenobiotic cle...

  7. ABC Transporters Involved in Export of Cell Surface Glycoconjugates

    PubMed Central

    Cuthbertson, Leslie; Kos, Veronica; Whitfield, Chris

    2010-01-01

    Summary: Complex glycoconjugates play critical roles in the biology of microorganisms. Despite the remarkable diversity in glycan structures and the bacteria that produce them, conserved themes are evident in the biosynthesis-export pathways. One of the primary pathways involves representatives of the ATP-binding cassette (ABC) transporter superfamily. These proteins are responsible for the export of a wide variety of cell surface oligo- and polysaccharides in both Gram-positive and Gram-negative bacteria. Recent investigations of the structure and function of ABC transporters involved in the export of lipopolysaccharide O antigens have revealed two fundamentally different strategies for coupling glycan polymerization to export. These mechanisms are distinguished by the presence (or absence) of characteristic nonreducing terminal modifications on the export substrates, which serve as chain termination and/or export signals, and by the presence (or absence) of a discrete substrate-binding domain in the nucleotide-binding domain polypeptide of the ABC transporter. A bioinformatic survey examining ABC exporters from known oligo- and polysaccharide biosynthesis loci identifies conserved nucleotide-binding domain protein families that correlate well with themes in the structures and assembly of glycans. The familial relationships among the ABC exporters generate hypotheses concerning the biosynthesis of structurally diverse oligo- and polysaccharides, which play important roles in the biology of bacteria with different lifestyles. PMID:20805402

  8. A Silent ABC Transporter Isolated from Streptomyces rochei F20 Induces Multidrug Resistance

    PubMed Central

    Fernández-Moreno, Miguel A.; Carbó, Lázaro; Cuesta, Trinidad; Vallín, Carlos; Malpartida, Francisco

    1998-01-01

    In the search for heterologous activators for actinorhodin production in Streptomyces lividans, 3.4 kb of DNA from Streptomyces rochei F20 (a streptothricin producer) were characterized. Subcloning experiments showed that the minimal DNA fragment required for activation was 0.4 kb in size. The activation is mediated by increasing the levels of transcription of the actII-ORF4 gene. Sequencing of the minimal activating fragment did not reveal any clues about its mechanism; nevertheless, it was shown to overlap the 3′ end of two convergent genes, one of whose translated products (ORF2) strongly resembles that of other genes belonging to the ABC transporter superfamily. Computer-assisted analysis of the 3.4-kb DNA sequence showed the 3′ terminus of an open reading frame (ORF), i.e., ORFA, and three complete ORFs (ORF1, ORF2, and ORFB). Searches in the databases with their respective gene products revealed similarities for ORF1 and ORF2 with ATP-binding proteins and transmembrane proteins, respectively, which are found in members of the ABC transporter superfamily. No similarities for ORFA and ORFB were found in the databases. Insertional inactivation of ORF1 and ORF2, their transcription analysis, and their cloning in heterologous hosts suggested that these genes were not expressed under our experimental conditions; however, cloning of ORF1 and ORF2 together (but not separately) under the control of an expressing promoter induced resistance to several chemically different drugs: oleandomycin, erythromycin, spiramycin, doxorubicin, and tetracycline. Thus, this genetic system, named msr, is a new bacterial multidrug ABC transporter. PMID:9696745

  9. Genome-Wide Identification, Characterization and Phylogenetic Analysis of ATP-Binding Cassette (ABC) Transporter Genes in Common Carp (Cyprinus carpio).

    PubMed

    Liu, Xiang; Li, Shangqi; Peng, Wenzhu; Feng, Shuaisheng; Feng, Jianxin; Mahboob, Shahid; Al-Ghanim, Khalid A; Xu, Peng

    2016-01-01

    The ATP-binding cassette (ABC) gene family is considered to be one of the largest gene families in all forms of prokaryotic and eukaryotic life. Although the ABC transporter genes have been annotated in some species, detailed information about the ABC superfamily and the evolutionary characterization of ABC genes in common carp (Cyprinus carpio) are still unclear. In this research, we identified 61 ABC transporter genes in the common carp genome. Phylogenetic analysis revealed that they could be classified into seven subfamilies, namely 11 ABCAs, six ABCBs, 19 ABCCs, eight ABCDs, two ABCEs, four ABCFs, and 11 ABCGs. Comparative analysis of the ABC genes in seven vertebrate species including common carp, showed that at least 10 common carp genes were retained from the third round of whole genome duplication, while 12 duplicated ABC genes may have come from the fourth round of whole genome duplication. Gene losses were also observed for 14 ABC genes. Expression profiles of the 61 ABC genes in six common carp tissues (brain, heart, spleen, kidney, intestine, and gill) revealed extensive functional divergence among the ABC genes. Different copies of some genes had tissue-specific expression patterns, which may indicate some gene function specialization. This study provides essential genomic resources for future studies in common carp.

  10. Genome-Wide Identification, Characterization and Phylogenetic Analysis of ATP-Binding Cassette (ABC) Transporter Genes in Common Carp (Cyprinus carpio)

    PubMed Central

    Peng, Wenzhu; Feng, Shuaisheng; Feng, Jianxin; Mahboob, Shahid; Al-Ghanim, Khalid A.

    2016-01-01

    The ATP-binding cassette (ABC) gene family is considered to be one of the largest gene families in all forms of prokaryotic and eukaryotic life. Although the ABC transporter genes have been annotated in some species, detailed information about the ABC superfamily and the evolutionary characterization of ABC genes in common carp (Cyprinus carpio) are still unclear. In this research, we identified 61 ABC transporter genes in the common carp genome. Phylogenetic analysis revealed that they could be classified into seven subfamilies, namely 11 ABCAs, six ABCBs, 19 ABCCs, eight ABCDs, two ABCEs, four ABCFs, and 11 ABCGs. Comparative analysis of the ABC genes in seven vertebrate species including common carp, showed that at least 10 common carp genes were retained from the third round of whole genome duplication, while 12 duplicated ABC genes may have come from the fourth round of whole genome duplication. Gene losses were also observed for 14 ABC genes. Expression profiles of the 61 ABC genes in six common carp tissues (brain, heart, spleen, kidney, intestine, and gill) revealed extensive functional divergence among the ABC genes. Different copies of some genes had tissue-specific expression patterns, which may indicate some gene function specialization. This study provides essential genomic resources for future studies in common carp. PMID:27058731

  11. The ABC protein turned chloride channel whose failure causes cystic fibrosis

    NASA Astrophysics Data System (ADS)

    Gadsby, David C.; Vergani, Paola; Csanády, László

    2006-03-01

    CFTR chloride channels are encoded by the gene mutated in patients with cystic fibrosis. These channels belong to the superfamily of ABC transporter ATPases. ATP-driven conformational changes, which in other ABC proteins fuel uphill substrate transport across cellular membranes, in CFTR open and close a gate to allow transmembrane flow of anions down their electrochemical gradient. New structural and biochemical information from prokaryotic ABC proteins and functional information from CFTR channels has led to a unifying mechanism explaining those ATP-driven conformational changes.

  12. Alzheimer's and ABC transporters--new opportunities for diagnostics and treatment.

    PubMed

    Pahnke, Jens; Langer, Oliver; Krohn, Markus

    2014-12-01

    Much has been said about the increasing number of demented patients and the main risk factor 'age'. Frustratingly, we do not know the precise pattern and all modulating factors that provoke the pathologic changes in the brains of affected elderly. We have to diagnose early to be able to stop the progression of diseases that irreversibly destroy brain substance. Familiar AD cases have mislead some researchers for almost 20 years, which has unfortunately narrowed the scientific understanding and has, thus, lead to insufficient funding of independent approaches. Therefore, basic researchers hardly have been able to develop causative treatments and clinicians still do not have access to prognostic and early diagnostic tools. During the recent years it became clear that insufficient Aβ export, physiologically facilitated by the ABC transporter superfamily at the brain's barriers, plays a fundamental role in disease initiation and progression. Furthermore, export mechanisms that are deficient in affected elderly are new targets for activation and, thus, treatment, but ideally also for prevention. In sporadic AD disturbed clearance of β-amyloid from the brain is so far the most important factor for its accumulation in the parenchyma and vessel walls. Here, we review findings about the contribution of ABC transporters and of the perivascular drainage/glymphatic system on β-amyloid clearance. We highlight their potential value for innovative early diagnostics using PET and describe recently described, effective ABC transporter-targeting agents as potential causative treatment for neurodegenerative proteopathies/dementias. Copyright © 2014 Elsevier Inc. All rights reserved.

  13. Alzheimer’s and ABC transporters - new opportunities for diagnostics and treatment

    PubMed Central

    Pahnke, Jens; Langer, Oliver; Krohn, Markus

    2014-01-01

    Much has been said about the increasing number of demented patients and the main risk factor ‘age’. Frustratingly, we do not know the precise pattern and all modulating factors that provoke the pathologic changes in the brains of affected elderly. We have to diagnose early to be able to stop the progression of diseases that irreversibly destroy brain substance. Familiar AD cases have mislead some researchers for almost 20 years, which has unfortunately narrowed the scientific understanding and has, thus, lead to insufficient funding of independent approaches. Therefore, basic researchers hardly have been able to develop causative treatments and clinicians still do not have access to prognostic and early diagnostic tools. During the recent years it became clear that insufficient Aβ export, physiologically facilitated by the ABC transporter superfamily at the brain’s barriers, plays a fundamental role in disease initiation and progression. Furthermore, export mechanisms that are deficient in affected elderly are new targets for activation and, thus, treatment, but ideally also for prevention. In sporadic AD disturbed clearance of β-amyloid from the brain is so far the most important factor for its accumulation in the parenchyma and vessel walls. Here, we review findings about the contribution of ABC transporters and of the perivascular drainage/glymphatic system on β-amyloid clearance. We highlight their potential value for innovative early diagnostics using PET and describe recently described, effective ABC transporter-targeting agents as potential causative treatment for neurodegenerative proteopathies/dementias. PMID:24746857

  14. ATP-binding cassette (ABC) proteins in aquatic invertebrates: Evolutionary significance and application in marine ecotoxicology.

    PubMed

    Jeong, Chang-Bum; Kim, Hui-Su; Kang, Hye-Min; Lee, Jae-Seong

    2017-04-01

    The ATP-binding cassette (ABC) protein superfamily is known to play a fundamental role in biological processes and is highly conserved across animal taxa. The ABC proteins function as active transporters for multiple substrates across the cellular membrane by ATP hydrolysis. As this superfamily is derived from a common ancestor, ABC genes have evolved via lineage-specific duplications through the process of adaptation. In this review, we summarized information about the ABC gene families in aquatic invertebrates, considering their evolution and putative functions in defense mechanisms. Phylogenetic analysis was conducted to examine the evolutionary significance of ABC gene families in aquatic invertebrates. Particularly, a massive expansion of multixenobiotic resistance (MXR)-mediated efflux transporters was identified in the absence of the ABCG2 (BCRP) gene in Ecdysozoa and Platyzoa, suggesting that a loss of Abcg2 gene occurred sporadically in these species during divergence of Protostome to Lophotrochozoa. Furthermore, in aquatic invertebrates, the ecotoxicological significance of MXR is discussed while considering the role of MXR-mediated efflux transporters in response to various environmental pollutants. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. ABC-B transporter genes in Dirofilaria immitis.

    PubMed

    Bourguinat, Catherine; Che, Hua; Mani, Thangadurai; Keller, Kathy; Prichard, Roger K

    2016-08-01

    Dirofilaria immitis is a filarial nematode causing infection and heartworm disease in dogs and other canids, cats, and occasionally in humans. Prevention with macrocyclic lactones (ML) is recommended during the mosquito transmission season. Recently, ML resistance has been reported. ABC-B transporter genes are thought to be involved in the mechanism of ML resistance in other nematodes. This study aimed to identify all the ABC-B transporter genes in D. immitis using as a reference the nDi.2.2 D. immitis whole genome, which is not completely annotated. Using bioinformatic tools and PCR amplification on pooled D. immitis genomic DNA and on pooled cDNA, nine ABC transporter genes including one pseudogene were characterized. Bioinformatic and phylogenetic analyses allowed identification of three P-glycoproteins (Pgps) (Dim-pgp-3 Dim-pgp-10, Dim-pgp-11), of two ABC-B half transporter genes (one ortholog of Cel-haf-4 and Cel-haf-9; and one ortholog of Cel-haf-1 and Cel-haf-3), of one ABC half transporter gene (ortholog of Cel-haf-5) that contained an ABC-C motif, and of one additional half transporter that would require functional study for characterization. The number of ABC-B transporter genes identified was lower than in Caenorhabditis elegans and Haemonchus contortus. Further studies are needed to understand their possible role in ML resistance in D. immitis. These ABC transporters constitute a base for ML resistance investigation in D. immitis and advance our understanding of the molecular biology of this parasite. Copyright © 2016. Published by Elsevier Ltd.

  16. Phylogenetic Characterization of Transport Protein Superfamilies: Superiority of SuperfamilyTree Programs over Those Based on Multiple Alignments

    PubMed Central

    Chen, Jonathan S.; Reddy, Vamsee; Chen, Joshua H.; Shlykov, Maksim A.; Zheng, Wei Hao; Cho, Jaehoon; Yen, Ming Ren; Saier, Milton H.

    2012-01-01

    Transport proteins function in the translocation of ions, solutes and macromolecules across cellular and organellar membranes. These integral membrane proteins fall into >600 families as tabulated in the Transporter Classification Database (www.tcdb.org). Recent studies, some of which are reported here, define distant phylogenetic relationships between families with the creation of superfamilies. Several of these are analyzed using a novel set of programs designed to allow reliable prediction of phylogenetic trees when sequence divergence is too great to allow the use of multiple alignments. These new programs, called SuperfamilyTree1 and 2 (SFT1 and 2), allow display of protein and family relationships, respectively, based on thousands of comparative BLAST scores rather than multiple alignments. Superfamilies analyzed include: (1) Aerolysins, (2) RTX Toxins, (3) Defensins, (4) Ion Transporters, (5) Bile/Arsenite/Riboflavin Transporters, (6) Cation: Proton Antiporters, and (7) the Glucose/Fructose/Lactose superfamily within the prokaryotic phosphoenol pyruvate-dependent Phosphotransferase System. In addition to defining the phylogenetic relationships of the proteins and families within these seven superfamilies, evidence is provided showing that the SFT programs outperform programs that are based on multiple alignments whenever sequence divergence of superfamily members is extensive. The SFT programs should be applicable to virtually any superfamily of proteins or nucleic acids. PMID:22286036

  17. Functional Dependence between Septal Protein SepJ from Anabaena sp. Strain PCC 7120 and an Amino Acid ABC-Type Uptake Transporter.

    PubMed

    Escudero, Leticia; Mariscal, Vicente; Flores, Enrique

    2015-08-01

    multicellular organisms. Heterocyst-forming cyanobacteria such as Anabaena represent a unique case of multicellularity, in which two cell types exchange nutrients and regulators. The SepJ protein located at the intercellular septa in the filaments of Anabaena contains a permease domain of the drug/metabolite transporter (DMT) superfamily that somehow contributes to intercellular molecular transfer. In this work, we have found that SepJ stimulates the activity of a polar amino acid uptake transporter of the ATP-binding-cassette (ABC) superfamily, which could itself affect an intercellular transfer activity related to SepJ, thus unraveling possible functional interactions between these different transporters. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  18. Placental ABC Transporters: Biological Impact and Pharmaceutical Significance.

    PubMed

    Joshi, Anand A; Vaidya, Soniya S; St-Pierre, Marie V; Mikheev, Andrei M; Desino, Kelly E; Nyandege, Abner N; Audus, Kenneth L; Unadkat, Jashvant D; Gerk, Phillip M

    2016-12-01

    The human placenta fulfills a variety of essential functions during prenatal life. Several ABC transporters are expressed in the human placenta, where they play a role in the transport of endogenous compounds and may protect the fetus from exogenous compounds such as therapeutic agents, drugs of abuse, and other xenobiotics. To date, considerable progress has been made toward understanding ABC transporters in the placenta. Recent studies on the expression and functional activities are discussed. This review discusses the placental expression and functional roles of several members of ABC transporter subfamilies B, C, and G including MDR1/P-glycoprotein, the MRPs, and BCRP, respectively. Since placental ABC transporters modulate fetal exposure to various compounds, an understanding of their functional and regulatory mechanisms will lead to more optimal medication use when necessary in pregnancy.

  19. Placental ABC Transporters: Biological Impact and Pharmaceutical Significance

    PubMed Central

    Joshi, Anand A.; Vaidya, Soniya S.; St-Pierre, Marie V.; Mikheev, Andrei M.; Desino, Kelly E.; Nyandege, Abner N.; Audus, Kenneth L.; Unadkat, Jashvant D.; Gerk, Phillip M.

    2017-01-01

    The human placenta fulfills a variety of essential functions during prenatal life. Several ABC transporters are expressed in the human placenta, where they play a role in the transport of endogenous compounds and may protect the fetus from exogenous compounds such as therapeutic agents, drugs of abuse, and other xenobiotics. To date, considerable progress has been made toward understanding ABC transporters in the placenta. Recent studies on the expression and functional activities are discussed. This review discusses the placental expression and functional roles of several members of ABC transporter subfamilies B, C, and G including MDR1/P-glycoprotein, the MRPs, and BCRP, respectively. Since placental ABC transporters modulate fetal exposure to various compounds, an understanding of their functional and regulatory mechanisms will lead to more optimal medication use when necessary in pregnancy. PMID:27644937

  20. Genome-wide identification of ATP-binding cassette (ABC) transporters and their roles in response to polycyclic aromatic hydrocarbons (PAHs) in the copepod Paracyclopina nana.

    PubMed

    Jeong, Chang-Bum; Kim, Duck-Hyun; Kang, Hye-Min; Lee, Young Hwan; Kim, Hui-Su; Kim, Il-Chan; Lee, Jae-Seong

    2017-02-01

    The ATP-binding cassette (ABC) protein superfamily is one of the largest gene families and is highly conserved in all domains. The ABC proteins play roles in several biological processes, including multi-xenobiotic resistance (MXR), by functioning as transporters in the cellular membrane. They also mediate the cellular efflux of a wide range of substrates against concentration gradients. In this study, 37 ABC genes belonging to eight distinct subfamilies were identified in the marine copepod Paracyclopina nana and annotated based on a phylogenetic analysis. Also, the functions of P-glycoproteins (P-gp) and multidrug resistance-associated proteins (MRPs), conferring MXR, were verified using fluorescent substrates and specific inhibitors. The activities of MXR-mediated ABC proteins and their transcriptional level were examined in response to polyaromatic hydrocarbons (PAHs), main components of the water-accommodated fraction. This study increases the understanding of the protective role of MXR in response to PAHs over the comparative evolution of ABC gene families. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. ATP-induced conformational changes of nucleotide-binding domains in an ABC transporter. Importance of the water-mediated entropic force.

    PubMed

    Hayashi, Tomohiko; Chiba, Shuntaro; Kaneta, Yusuke; Furuta, Tadaomi; Sakurai, Minoru

    2014-11-06

    ATP binding cassette (ABC) proteins belong to a superfamily of active transporters. Recent experimental and computational studies have shown that binding of ATP to the nucleotide binding domains (NBDs) of ABC proteins drives the dimerization of NBDs, which, in turn, causes large conformational changes within the transmembrane domains (TMDs). To elucidate the active substrate transport mechanism of ABC proteins, it is first necessary to understand how the NBD dimerization is driven by ATP binding. In this study, we selected MalKs (NBDs of a maltose transporter) as a representative NBD and calculated the free-energy change upon dimerization using molecular mechanics calculations combined with a statistical thermodynamic theory of liquids, as well as a method to calculate the translational, rotational, and vibrational entropy change. This combined method is applied to a large number of snapshot structures obtained from molecular dynamics simulations containing explicit water molecules. The results suggest that the NBD dimerization proceeds with a large gain of water entropy when ATP molecules bind to the NBDs. The energetic gain arising from direct NBD-NBD interactions is canceled by the dehydration penalty and the configurational-entropy loss. ATP hydrolysis induces a loss of the shape complementarity between the NBDs, which leads to the dissociation of the dimer, due to a decrease in the water-entropy gain and an increase in the configurational-entropy loss. This interpretation of the NBD dimerization mechanism in concert with ATP, especially focused on the water-mediated entropy force, is potentially applicable to a wide variety of the ABC transporters.

  2. Comparison of mechanistic transport cycle models of ABC exporters.

    PubMed

    Szöllősi, Dániel; Rose-Sperling, Dania; Hellmich, Ute A; Stockner, Thomas

    2018-04-01

    ABC (ATP binding cassette) transporters, ubiquitous in all kingdoms of life, carry out essential substrate transport reactions across cell membranes. Their transmembrane domains bind and translocate substrates and are connected to a pair of nucleotide binding domains, which bind and hydrolyze ATP to energize import or export of substrates. Over four decades of investigations into ABC transporters have revealed numerous details from atomic-level structural insights to their functional and physiological roles. Despite all these advances, a comprehensive understanding of the mechanistic principles of ABC transporter function remains elusive. The human multidrug resistance transporter ABCB1, also referred to as P-glycoprotein (P-gp), is one of the most intensively studied ABC exporters. Using ABCB1 as the reference point, we aim to compare the dominating mechanistic models of substrate transport and ATP hydrolysis for ABC exporters and to highlight the experimental and computational evidence in their support. In particular, we point out in silico studies that enhance and complement available biochemical data. "This article is part of a Special Issue entitled: Beyond the Structure-Function Horizon of Membrane Proteins edited by Ute Hellmich, Rupak Doshi and Benjamin McIlwain." Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  3. ABC transporters and immunity: mechanism of self-defense.

    PubMed

    Hinz, Andreas; Tampé, Robert

    2012-06-26

    The transporter associated with antigen processing (TAP) is a prototype of an asymmetric ATP-binding cassette (ABC) transporter, which uses ATP binding and hydrolysis to translocate peptides from the cytosol to the lumen of the endoplasmic reticulum (ER). Here, we review molecular details of peptide binding and ATP binding and hydrolysis as well as the resulting allosteric cross-talk between the nucleotide-binding domains and the transmembrane domains that drive translocation of the solute across the ER membrane. We also discuss the general molecular architecture of ABC transporters and demonstrate the importance of structural and functional studies for a better understanding of the role of the noncanonical site of asymmetric ABC transporters. Several aspects of peptide binding and specificity illustrate details of peptide translocation by TAP. Furthermore, this ABC transporter forms the central part of the major histocompatibility complex class I (MHC I) peptide-loading machinery. Hence, TAP is confronted with a number of viral factors, which prevent antigen translocation and MHC I loading in virally infected cells. We review how these viral factors have been used as molecular tools to decipher mechanistic aspects of solute translocation and discuss how they can help in the structural analysis of TAP.

  4. Whole-Genome Survey of the Putative ATP-Binding Cassette Transporter Family Genes in Vitis vinifera

    PubMed Central

    Çakır, Birsen; Kılıçkaya, Ozan

    2013-01-01

    The ATP-binding cassette (ABC) protein superfamily constitutes one of the largest protein families known in plants. In this report, we performed a complete inventory of ABC protein genes in Vitis vinifera, the whole genome of which has been sequenced. By comparison with ABC protein members of Arabidopsis thaliana, we identified 135 putative ABC proteins with 1 or 2 NBDs in V. vinifera. Of these, 120 encode intrinsic membrane proteins, and 15 encode proteins missing TMDs. V. vinifera ABC proteins can be divided into 13 subfamilies with 79 “full-size,” 41 “half-size,” and 15 “soluble” putative ABC proteins. The main feature of the Vitis ABC superfamily is the presence of 2 large subfamilies, ABCG (pleiotropic drug resistance and white-brown complex homolog) and ABCC (multidrug resistance-associated protein). We identified orthologs of V. vinifera putative ABC transporters in different species. This work represents the first complete inventory of ABC transporters in V. vinifera. The identification of Vitis ABC transporters and their comparative analysis with the Arabidopsis counterparts revealed a strong conservation between the 2 species. This inventory could help elucidate the biological and physiological functions of these transporters in V. vinifera. PMID:24244377

  5. Diversity in ABC transporters: Type I, II and III importers

    PubMed Central

    Rice, Austin J.; Park, Aekyung

    2014-01-01

    ATP-binding cassette transporters are multi-subunit membrane pumps that transport substrates across membranes. While significant in the transport process, transporter architecture exhibits a range of diversity that we are only beginning to recognize. This divergence may provide insight into the mechanisms of substrate transport and homeostasis. Until recently, ABC importers have been classified into two types, but with the emergence of energy-coupling factor (ECF) transporters there are potentially three types of ABC importers. In this review, we summarize an expansive body of research on the three types of importers with an emphasis on the basics that underlie ABC importers, such as structure, subunit composition and mechanism. PMID:25155087

  6. ATP binding cassette (ABC) transporters: expression and clinical value in glioblastoma.

    PubMed

    Dréan, Antonin; Rosenberg, Shai; Lejeune, François-Xavier; Goli, Larissa; Nadaradjane, Aravindan Arun; Guehennec, Jérémy; Schmitt, Charlotte; Verreault, Maïté; Bielle, Franck; Mokhtari, Karima; Sanson, Marc; Carpentier, Alexandre; Delattre, Jean-Yves; Idbaih, Ahmed

    2018-03-08

    ATP-binding cassette transporters (ABC transporters) regulate traffic of multiple compounds, including chemotherapeutic agents, through biological membranes. They are expressed by multiple cell types and have been implicated in the drug resistance of some cancer cells. Despite significant research in ABC transporters in the context of many diseases, little is known about their expression and clinical value in glioblastoma (GBM). We analyzed expression of 49 ABC transporters in both commercial and patient-derived GBM cell lines as well as from 51 human GBM tumor biopsies. Using The Cancer Genome Atlas (TCGA) cohort as a training dataset and our cohort as a validation dataset, we also investigated the prognostic value of these ABC transporters in newly diagnosed GBM patients, treated with the standard of care. In contrast to commercial GBM cell lines, GBM-patient derived cell lines (PDCL), grown as neurospheres in a serum-free medium, express ABC transporters similarly to parental tumors. Serum appeared to slightly increase resistance to temozolomide correlating with a tendency for an increased expression of ABCB1. Some differences were observed mainly due to expression of ABC transporters by microenvironmental cells. Together, our data suggest that the efficacy of chemotherapeutic agents may be misestimated in vitro if they are the targets of efflux pumps whose expression can be modulated by serum. Interestingly, several ABC transporters have prognostic value in the TCGA dataset. In our cohort of 51 GBM patients treated with radiation therapy with concurrent and adjuvant temozolomide, ABCA13 overexpression is associated with a decreased progression free survival in univariate (p < 0.01) and multivariate analyses including MGMT promoter methylation (p = 0.05) suggesting reduced sensitivity to temozolomide in ABCA13 overexpressing GBM. Expression of ABC transporters is: (i) detected in GBM and microenvironmental cells and (ii) better reproduced in GBM

  7. Detergent-free purification of ABC (ATP-binding-cassette) transporters.

    PubMed

    Gulati, Sonali; Jamshad, Mohammed; Knowles, Timothy J; Morrison, Kerrie A; Downing, Rebecca; Cant, Natasha; Collins, Richard; Koenderink, Jan B; Ford, Robert C; Overduin, Michael; Kerr, Ian D; Dafforn, Timothy R; Rothnie, Alice J

    2014-07-15

    ABC (ATP-binding-cassette) transporters carry out many vital functions and are involved in numerous diseases, but study of the structure and function of these proteins is often hampered by their large size and membrane location. Membrane protein purification usually utilizes detergents to solubilize the protein from the membrane, effectively removing it from its native lipid environment. Subsequently, lipids have to be added back and detergent removed to reconstitute the protein into a lipid bilayer. In the present study, we present the application of a new methodology for the extraction and purification of ABC transporters without the use of detergent, instead, using a copolymer, SMA (polystyrene-co-maleic acid). SMA inserts into a bilayer and assembles into discrete particles, essentially solubilizing the membrane into small discs of bilayer encircled by a polymer, termed SMALPs (SMA lipid particles). We show that this polymer can extract several eukaryotic ABC transporters, P-glycoprotein (ABCB1), MRP1 (multidrug-resistance protein 1; ABCC1), MRP4 (ABCC4), ABCG2 and CFTR (cystic fibrosis transmembrane conductance regulator; ABCC7), from a range of different expression systems. The SMALP-encapsulated ABC transporters can be purified by affinity chromatography, and are able to bind ligands comparably with those in native membranes or detergent micelles. A greater degree of purity and enhanced stability is seen compared with detergent solubilization. The present study demonstrates that eukaryotic ABC transporters can be extracted and purified without ever being removed from their lipid bilayer environment, opening up a wide range of possibilities for the future study of their structure and function.

  8. Molecular cloning and expression profile of an ATP-binding cassette (ABC) transporter gene from the hemipteran insect Nilaparvata lugens.

    PubMed

    Zha, W J; Li, S H; Zhou, L; Chen, Z J; Liu, K; Yang, G C; Hu, G; He, G C; You, A Q

    2015-03-30

    The ATP-binding cassette (ABC) transporters belong to a large superfamily of proteins that have important physiological functions in all living organisms. In insects, ABC transporters have important functions in the transport of molecules, and are also involved in insecticide resistance, metabolism, and development. In this study, the Nilaparvata lugens Stal (Hemiptera: Delphacidae) ABCG (NlABCG) gene was identified and characterized. The complete mRNA sequence of NlABCG was 2608-bp long, with an open reading frame of 2064 bp encoding a protein comprised of 687 amino acids. The conserved regions include three N-glycosylation and 34 phosphorylation sites, as well as seven transmembrane domains. The amino acid identity with the closely related species Acyrthosiphon pisum was 42.8%. Developmental expression analysis using quantitative real-time reverse transcriptase PCR suggested that the NlABCG transcript was expressed at all developmental stages of N. lugens. The lowest expression of NlABCG was in the 1st instar, and levels increased with larval growth. The transcript profiles of NlABCG were analyzed in various tissues from a 5th instar nymph, and the highest expression was observed in the midgut. These results suggest that the sequence, characteristics, and expression of NlABCG are highly conserved, and basic information is provided for its functional analysis.

  9. Multidrug resistance in parasites: ABC transporters, P-glycoproteins and molecular modelling.

    PubMed

    Jones, P M; George, A M

    2005-04-30

    Parasitic diseases, caused by protozoa, helminths and arthropods, rank among the most important problems in human and veterinary medicine, and in agriculture, leading to debilitating sicknesses and loss of life. In the absence of vaccines and with the general failure of vector eradication programs, drugs are the main line of defence, but the newest drugs are being tracked by the emergence of resistance in parasites, sharing ominous parallels with multidrug resistance in bacterial pathogens. Any of a number of mechanisms will elicit a drug resistance phenotype in parasites, including: active efflux, reduced uptake, target modification, drug modification, drug sequestration, by-pass shunting, or substrate competition. The role of ABC transporters in parasitic multidrug resistance mechanisms is being subjected to more scrutiny, due in part to the established roles of certain ABC transporters in human diseases, and also to an increasing portfolio of ABC transporters from parasite genome sequencing projects. For example, over 100 ABC transporters have been identified in the Escherichia coli genome, but to date only about 65 in all parasitic genomes. Long established laboratory investigations are now being assisted by molecular biology, bioinformatics, and computational modelling, and it is in these areas that the role of ABC transporters in parasitic multidrug resistance mechanisms may be defined and put in perspective with that of other proteins. We discuss ABC transporters in parasites, and conclude with an example of molecular modelling that identifies a new interaction between the structural domains of a parasite P-glycoprotein.

  10. A PhoPQ-Regulated ABC Transporter System Exports Tetracycline in Pseudomonas aeruginosa.

    PubMed

    Chen, Lin; Duan, Kangmin

    2016-05-01

    Pseudomonas aeruginosa is an important human pathogen whose infections are difficult to treat due to its high intrinsic resistance to many antibiotics. Here, we show that the disruption of PA4456, encoding the ATP binding component of a putative ATP-binding cassette (ABC) transporter, increased the bacterium's susceptible to tetracycline and other antibiotics or toxic chemicals. Fluorescence spectroscopy and antibiotic accumulation tests showed that the interruption of the ABC transporter caused increased intracellular accumulation of tetracycline, demonstrating a role of the ABC transporter in tetracycline expulsion. Site-directed mutagenesis proved that the conserved residues of E170 in the Walker B motif and H203 in the H-loop, which are important for ATP hydrolysis, were essential for the function of PA4456. Through a genome-wide search, the PhoPQ two-component system was identified as a regulator of the computationally predicted PA4456-4452 operon that encodes the ABC transporter system. A >5-fold increase of the expression of this operon was observed in the phoQ mutant. The results obtained also show that the expression of the phzA1B1C1D1E1 operon and the production of pyocyanin were significantly higher in the ABC transporter mutant, signifying a connection between the ABC transporter and pyocyanin production. These results indicated that the PhoPQ-regulated ABC transporter is associated with intrinsic resistance to antibiotics and other adverse compounds in P. aeruginosa, probably by extruding them out of the cell. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  11. ABC transporters are involved in defense against permethrin insecticide in the malaria vector Anopheles stephensi.

    PubMed

    Epis, Sara; Porretta, Daniele; Mastrantonio, Valentina; Comandatore, Francesco; Sassera, Davide; Rossi, Paolo; Cafarchia, Claudia; Otranto, Domenico; Favia, Guido; Genchi, Claudio; Bandi, Claudio; Urbanelli, Sandra

    2014-07-29

    Proteins from the ABC family (ATP-binding cassette) represent the largest known group of efflux pumps, responsible for transporting specific molecules across lipid membranes in both prokaryotic and eukaryotic organisms. In arthropods they have been shown to play a role in insecticide defense/resistance. The presence of ABC transporters and their possible association with insecticide transport have not yet been investigated in the mosquito Anopheles stephensi, the major vector of human malaria in the Middle East and South Asian regions. Here we investigated the presence and role of ABCs in transport of permethrin insecticide in a susceptible strain of this mosquito species. To identify ABC transporter genes we obtained a transcriptome from untreated larvae of An. stephensi and then compared it with the annotated transcriptome of Anopheles gambiae. To analyse the association between ABC transporters and permethrin we conducted bioassays with permethrin alone and in combination with an ABC inhibitor, and then we investigated expression profiles of the identified genes in larvae exposed to permethrin. Bioassays showed an increased mortality of mosquitoes when permethrin was used in combination with the ABC-transporter inhibitor. Genes for ABC transporters were detected in the transcriptome, and five were selected (AnstABCB2, AnstABCB3, AnstABCB4, AnstABCmember6 and AnstABCG4). An increased expression in one of them (AnstABCG4) was observed in larvae exposed to the LD50 dose of permethrin. Contrary to what was found in other insect species, no up-regulation was observed in the AnstABCB genes. Our results show for the first time the involvement of ABC transporters in larval defense against permethrin in An. stephensi and, more in general, confirm the role of ABC transporters in insecticide defense. The differences observed with previous studies highlight the need of further research as, despite the growing number of studies on ABC transporters in insects, the

  12. Identification of ABC Transporter Genes of Fusarium graminearum with Roles in Azole Tolerance and/or Virulence

    PubMed Central

    Döll, Katharina; Karlovsky, Petr; Deising, Holger B.; Wirsel, Stefan G. R.

    2013-01-01

    Fusarium graminearum is a plant pathogen infecting several important cereals, resulting in substantial yield losses and mycotoxin contamination of the grain. Triazole fungicides are used to control diseases caused by this fungus on a worldwide scale. Our previous microarray study indicated that 15 ABC transporter genes were transcriptionally upregulated in response to tebuconazole treatment. Here, we deleted four ABC transporter genes in two genetic backgrounds of F. graminearum representing the DON (deoxynivalenol) and the NIV (nivalenol) trichothecene chemotypes. Deletion of FgABC3 and FgABC4 belonging to group I of ABC-G and to group V of ABC-C subfamilies of ABC transporters, respectively, considerably increased the sensitivity to the class I sterol biosynthesis inhibitors triazoles and fenarimol. Such effects were specific since they did not occur with any other fungicide class tested. Assessing the contribution of the four ABC transporters to virulence of F. graminearum revealed that, irrespective of their chemotypes, deletion mutants of FgABC1 (ABC-C subfamily group V) and FgABC3 were impeded in virulence on wheat, barley and maize. Phylogenetic context and analyses of mycotoxin production suggests that FgABC3 may encode a transporter protecting the fungus from host-derived antifungal molecules. In contrast, FgABC1 may encode a transporter responsible for the secretion of fungal secondary metabolites alleviating defence of the host. Our results show that ABC transporters play important and diverse roles in both fungicide resistance and pathogenesis of F. graminearum. PMID:24244413

  13. Harnessing Drug Resistance: Using ABC Transporter Proteins To Target Cancer Cells

    PubMed Central

    Leitner, Heather M.; Kachadourian, Remy; Day, Brian J.

    2007-01-01

    The ATP-binding cassette (ABC) class of proteins is one of the most functionally diverse transporter families found in biological systems. Although the abundance of ABC proteins varies between species, they are highly conserved in sequence and often demonstrate similar functions across prokaryotic and eukaryotic organisms. Beginning with a brief summary of the events leading to our present day knowledge of ABC transporters, the purpose of this review is to discuss the potential for utilizing ABC transporters as a means for cellular glutathione (GSH) modulation. GSH is one of the most abundant thiol antioxidants in cells. It is involved in cellular division, protein and DNA synthesis, maintenance of cellular redox status and xenobiotic metabolism. Cellular GSH levels are often altered in many disease states including cancer. Over the past two decades there has been considerable emphasis on methods to sensitize cancer cells to chemotherapeutics and ionization radiation therapy by GSH depletion. We contend that ABC transporters, particularly multi-drug resistant proteins (MRPs), may be used as therapeutic targets for applications aimed at modulation of GSH levels. This review will emphasize MRP-mediated modulation of intracellular GSH levels as a potential alternative and adjunctive approach for cancer therapy. PMID:17585883

  14. The interaction of gut microbes with host ABC transporters

    PubMed Central

    Mercado-Lubo, Regino

    2010-01-01

    ATP binding cassette (ABC) transporters are increasingly recognized for their ability to modulate the absorption, distribution, metabolism, secretion and toxicity of xenobiotics. In addition to their essential function in drug resistance, there is also emerging evidence documenting the important role ABC transporters play in tissue defense. In this respect, the gastrointestinal tract represents a critical vanguard of defense against oral exposure of drugs while at the same time functions as a physical barrier between the lumenal contents (including bacteria) and the intestinal epithelium. Given emerging evidence suggesting that multidrug resistance protein (MDR) plays an important role in host-bacterial interactions in the gastrointestinal tract, this review will discuss the interplay between MDR of the intestinal epithelial cell barrier and gut microbes in health and disease. In particular, we will explore host-microbe interactions involving three apically restricted ABC transporters of the intestinal epithelium; P-glycoprotein (P-gp), multidrug resistance-associated protein 2 (MRP2) and cystic fibrosis transmembrane regulator (CFTR). PMID:21327038

  15. ABC multidrug transporters: target for modulation of drug pharmacokinetics and drug-drug interactions.

    PubMed

    Marquez, Béatrice; Van Bambeke, Françoise

    2011-05-01

    Nine proteins of the ABC superfamily (P-glycoprotein, 7 MRPs and BCRP) are involved in multidrug transport. Being localised at the surface of endothelial or epithelial cells, they expel drugs back to the external medium (if located at the apical side [P-glycoprotein, BCRP, MRP2, MRP4 in the kidney]) or to the blood (if located at the basolateral side [MRP1, MRP3, MRP4, MRP5]), modulating thereby their absorption, distribution, and elimination. In the CNS, most transporters are oriented to expel drugs to the blood. Transporters also cooperate with Phase I/Phase II metabolism enzymes by eliminating drug metabolites. Their major features are (i) their capacity to recognize drugs belonging to unrelated pharmacological classes, and (ii) their redundancy, a single molecule being possibly substrate for different transporters. This ensures an efficient protection of the body against invasion by xenobiotics. Competition for transport is now characterized as a mechanism of interaction between co-administered drugs, one molecule limiting the transport of the other, potentially affecting bioavailability, distribution, and/or elimination. Again, this mechanism reinforces drug interactions mediated by cytochrome P450 inhibition, as many substrates of P-glycoprotein and CYP3A4 are common. Induction of the expression of genes coding for MDR transporters is another mechanism of drug interaction, which could affect all drug substrates of the up-regulated transporter. Overexpression of MDR transporters confers resistance to anticancer agents and other therapies. All together, these data justify why studying drug active transport should be part of the evaluation of new drugs, as recently recommended by the FDA.

  16. Interaction of ABC multidrug transporters with anticancer protein kinase inhibitors: substrates and/or inhibitors?

    PubMed

    Hegedus, Csilla; Ozvegy-Laczka, Csilla; Szakács, Gergely; Sarkadi, Balázs

    2009-05-01

    Protein kinase inhibitors (PKI) are becoming key agents in modern cancer chemotherapy, and combination of PKIs with classical chemotherapeutic drugs may help to overcome currently untreatable metastatic cancers. Since chemotherapy resistance is a recurrent problem, mechanisms of resistance should be clarified in order to help further drug development. Here we suggest that in addition to PKI resistance based on altered target structures, the active removal of these therapeutic agents by the MDR-ABC transporters should also be considered as a major cause of clinical resistance. We discuss the occurring systemic and cellular mechanisms, which may hamper PKI efficiency, and document the role of selected MDR-ABC transporters in these phenomena through their interactions with these anticancer agents. Moreover, we suggest that PKI interactions with ABC transporters may modulate overall drug metabolism, including the fate of diverse, chemically or target-wise unrelated drugs. These effects are based on multiple forms of MDR-ABC transporter interaction with PKIs, as these compounds may be both substrates and/or inhibitors of an ABC transporter. We propose that these interactions should be carefully considered in clinical application, and a combined MDR-ABC transporter and PKI effect may bring a major advantage in future drug development.

  17. Structure of the MacAB-TolC ABC-type tripartite multidrug efflux pump

    PubMed Central

    Llabrés, Salomé; Neuberger, Arthur; Blaza, James N.; Bai, Xiao-chen; Okada, Ui; Murakami, Satoshi; van Veen, Hendrik W.; Zachariae, Ulrich; Scheres, Sjors H.W.; Luisi, Ben F.

    2017-01-01

    The MacA-MacB-TolC assembly of Escherichia coli is a transmembrane machine that spans the cell envelope and actively extrudes substrates, including macrolide antibiotics and polypeptide virulence factors. These transport processes are energized by the ATPase MacB, a member of the ATP-binding cassette (ABC) superfamily. We present an electron cryo-microscopy structure of the ABC-type tripartite assembly at near-atomic resolution. A hexamer of the periplasmic protein MacA bridges between a TolC trimer in the outer membrane and a MacB dimer in the inner membrane, generating a quaternary structure with a central channel for substrate translocation. A gating ring found in MacA is proposed to act as a one-way valve in substrate transport. The MacB structure features an atypical transmembrane domain (TMD) with a closely packed dimer interface and a periplasmic opening that is the likely portal for substrate entry from the periplasm, with subsequent displacement through an allosteric transport mechanism. PMID:28504659

  18. ABC transporter activity linked to radiation resistance and molecular subtype in pediatric medulloblastoma

    PubMed Central

    2013-01-01

    Background Resistance to radiation treatment remains a major clinical problem for patients with brain cancer. Medulloblastoma is the most common malignant brain tumor of childhood, and occurs in the cerebellum. Though radiation treatment has been critical in increasing survival rates in recent decades, the presence of resistant cells in a substantial number of medulloblastoma patients leads to relapse and death. Methods Using the established medulloblastoma cell lines UW228 and Daoy, we developed a novel model system to enrich for and study radiation tolerant cells early after radiation exposure. Using fluorescence-activated cell sorting, dead cells and cells that had initiated apoptosis were removed, allowing surviving cells to be investigated before extensive proliferation took place. Results Isolated surviving cells were tumorigenic in vivo and displayed elevated levels of ABCG2, an ABC transporter linked to stem cell behavior and drug resistance. Further investigation showed another family member, ABCA1, was also elevated in surviving cells in these lines, as well as in early passage cultures from pediatric medulloblastoma patients. We discovered that the multi-ABC transporter inhibitors verapamil and reserpine sensitized cells from particular patients to radiation, suggesting that ABC transporters have a functional role in cellular radiation protection. Additionally, verapamil had an intrinsic anti-proliferative effect, with transient exposure in vitro slowing subsequent in vivo tumor formation. When expression of key ABC transporter genes was assessed in medulloblastoma tissue from 34 patients, levels were frequently elevated compared with normal cerebellum. Analysis of microarray data from independent cohorts (n = 428 patients) showed expression of a number of ABC transporters to be strongly correlated with certain medulloblastoma subtypes, which in turn are associated with clinical outcome. Conclusions ABC transporter inhibitors are already being

  19. ABC Transporters and Isothiocyanates: Potential for Pharmacokinetic Diet–Drug Interactions

    PubMed Central

    Telang, Urvi; Ji, Yan; Morris, Marilyn E.

    2013-01-01

    Isothiocyanates, a class of anti-cancer agents, are derived from cruciferous vegetables such as broccoli, cabbage and watercress, and have demonstrated chemopreventive activity in a number of cancer models and epidemiologic studies. Due to public interest in cancer prevention and alternative therapies in cancer, the consumption of herbal supplements and vegetables containing these compounds is widespread and increasing. Isothiocyanates interact with ATP-binding cassette (ABC) efflux transporters such as P-glycoprotein, MRP1, MRP2 and BCRP, and may influence the pharmacokinetics of substrates of these transporters. This review discusses the pharmacokinetic properties of isothiocyanates, their interactions with ABC transporters, and presents some data describing the potential for isothiocyanate-mediated diet–drug interactions. PMID:19623673

  20. Sensitive and Specific Fluorescent Probes for Functional Analysis of the Three Major Types of Mammalian ABC Transporters

    PubMed Central

    Lebedeva, Irina V.; Pande, Praveen; Patton, Wayne F.

    2011-01-01

    An underlying mechanism for multi drug resistance (MDR) is up-regulation of the transmembrane ATP-binding cassette (ABC) transporter proteins. ABC transporters also determine the general fate and effect of pharmaceutical agents in the body. The three major types of ABC transporters are MDR1 (P-gp, P-glycoprotein, ABCB1), MRP1/2 (ABCC1/2) and BCRP/MXR (ABCG2) proteins. Flow cytometry (FCM) allows determination of the functional expression levels of ABC transporters in live cells, but most dyes used as indicators (rhodamine 123, DiOC2(3), calcein-AM) have limited applicability as they do not detect all three major types of ABC transporters. Dyes with broad coverage (such as doxorubicin, daunorubicin and mitoxantrone) lack sensitivity due to overall dimness and thus may yield a significant percentage of false negative results. We describe two novel fluorescent probes that are substrates for all three common types of ABC transporters and can serve as indicators of MDR in flow cytometry assays using live cells. The probes exhibit fast internalization, favorable uptake/efflux kinetics and high sensitivity of MDR detection, as established by multidrug resistance activity factor (MAF) values and Kolmogorov-Smirnov statistical analysis. Used in combination with general or specific inhibitors of ABC transporters, both dyes readily identify functional efflux and are capable of detecting small levels of efflux as well as defining the type of multidrug resistance. The assay can be applied to the screening of putative modulators of ABC transporters, facilitating rapid, reproducible, specific and relatively simple functional detection of ABC transporter activity, and ready implementation on widely available instruments. PMID:21799851

  1. ABCdb: an online resource for ABC transporter repertories from sequenced archaeal and bacterial genomes.

    PubMed

    Fichant, Gwennaele; Basse, Marie-Jeanne; Quentin, Yves

    2006-03-01

    The ATP-binding cassette (ABC) transporters are one of the major classes of active transporters. They are widespread in archaea, bacteria, and eukaryota, indicating that they have arisen early in evolution. They are involved in many essential physiological processes, but the majority import or export a wide variety of compounds across cellular membranes. These systems share a common architecture composed of four (exporters) or five (importers) domains. To identify and reconstruct functional ABC transporters encoded by archaeal and bacterial genomes, we have developed a bioinformatic strategy. Cross-reference to the transport classification system is used to predict the type of compound transported. A high quality of annotation is achieved by manual verification of the predictions. However, in order to face the rapid increase in the number of published genomes, we also include analyses of genomes issuing directly from the automated strategy. Querying the database (http://www-abcdb.biotoul.fr) allows to easily retrieve ABC transporter repertories and related data. Additional query tools have been developed for the analysis of the ABC family from both functional and evolutionary perspectives.

  2. ABC transporters affect the elimination and toxicity of CdTe quantum dots in liver and kidney cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chen, Mingli; Yin, Huancai; Bai, Pengli

    This paper aimed to investigate the role of adenosine triphosphate-binding cassette (ABC) transporters on the efflux and the toxicity of nanoparticles in liver and kidney cells. In this study, we synthesized CdTe quantum dots (QDs) that were monodispersed and emitted green fluorescence (maximum peak at 530 nm). Such QDs tended to accumulate in human hepatocellular carcinoma cells (HepG2), human kidney cells 2 (HK-2), and Madin-Darby canine kidney (MDCK) cells, and cause significant toxicity in all the three cell lines. Using specific inhibitors and inducers of P-glycoprotein (Pgp) and multidrug resistance associated proteins (Mrps), the cellular accumulation and subsequent toxicity ofmore » QDs in HepG2 and HK-2 cells were significantly affected, while only slight changes appeared in MDCK cells, corresponding well with the functional expressions of ABC transporters in cells. Moreover, treatment of QDs caused concentration- and time- dependent induction of ABC transporters in HepG2 and HK-2 cells, but such phenomenon was barely found in MDCK cells. Furthermore, the effects of CdTe QDs on ABC transporters were found to be greater than those of CdCl{sub 2} at equivalent concentrations of cadmium, indicating that the effects of QDs should be a combination of free Cd{sup 2+} and specific properties of QDs. Overall, these results indicated a strong dependence between the functional expressions of ABC transporters and the efflux of QDs, which could be an important reason for the modulation of QDs toxicity by ABC transporters. - Highlights: • ABC transporters contributed actively to the cellular efflux of CdTe quantum dots. • ABC transporters affected the cellular toxicity of CdTe quantum dots. • Treatment of CdTe quantum dots induced the gene expression of ABC transporters. • Free Cd{sup 2+} should be partially involved in the effects of QDs on ABC transporters. • Cellular efflux of quantum dots could be an important modulator for its toxicity.« less

  3. Structure of a Type-1 Secretion System ABC Transporter.

    PubMed

    Morgan, Jacob L W; Acheson, Justin F; Zimmer, Jochen

    2017-03-07

    Type-1 secretion systems (T1SSs) represent a widespread mode of protein secretion across the cell envelope in Gram-negative bacteria. The T1SS is composed of an inner-membrane ABC transporter, a periplasmic membrane-fusion protein, and an outer-membrane porin. These three components assemble into a complex spanning both membranes and providing a conduit for the translocation of unfolded polypeptides. We show that ATP hydrolysis and assembly of the entire T1SS complex is necessary for protein secretion. Furthermore, we present a 3.15-Å crystal structure of AaPrtD, the ABC transporter found in the Aquifex aeolicus T1SS. The structure suggests a substrate entry window just above the transporter's nucleotide binding domains. In addition, highly kinked transmembrane helices, which frame a narrow channel not observed in canonical peptide transporters, are likely involved in substrate translocation. Overall, the AaPrtD structure supports a polypeptide transport mechanism distinct from alternating access. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Beyond cellular detoxification: a plethora of physiological roles for MDR transporter homologs in plants

    PubMed Central

    Remy, Estelle; Duque, Paula

    2014-01-01

    Higher plants possess a multitude of Multiple Drug Resistance (MDR) transporter homologs that group into three distinct and ubiquitous families—the ATP-Binding Cassette (ABC) superfamily, the Major Facilitator Superfamily (MFS), and the Multidrug And Toxic compound Extrusion (MATE) family. As in other organisms, such as fungi, mammals, and bacteria, MDR transporters make a primary contribution to cellular detoxification processes in plants, mainly through the extrusion of toxic compounds from the cell or their sequestration in the central vacuole. This review aims at summarizing the currently available information on the in vivo roles of MDR transporters in plant systems. Taken together, these data clearly indicate that the biological functions of ABC, MFS, and MATE carriers are not restricted to xenobiotic and metal detoxification. Importantly, the activity of plant MDR transporters also mediates biotic stress resistance and is instrumental in numerous physiological processes essential for optimal plant growth and development, including the regulation of ion homeostasis and polar transport of the phytohormone auxin. PMID:24910617

  5. Predictive Structure and Topology of Peroxisomal ATP-Binding Cassette (ABC) Transporters

    PubMed Central

    Andreoletti, Pierre; Raas, Quentin; Gondcaille, Catherine; Cherkaoui-Malki, Mustapha; Trompier, Doriane; Savary, Stéphane

    2017-01-01

    The peroxisomal ATP-binding Cassette (ABC) transporters, which are called ABCD1, ABCD2 and ABCD3, are transmembrane proteins involved in the transport of various lipids that allow their degradation inside the organelle. Defective ABCD1 leads to the accumulation of very long-chain fatty acids and is associated with a complex and severe neurodegenerative disorder called X-linked adrenoleukodystrophy (X-ALD). Although the nucleotide-binding domain is highly conserved and characterized within the ABC transporters family, solid data are missing for the transmembrane domain (TMD) of ABCD proteins. The lack of a clear consensus on the secondary and tertiary structure of the TMDs weakens any structure-function hypothesis based on the very diverse ABCD1 mutations found in X-ALD patients. Therefore, we first reinvestigated thoroughly the structure-function data available and performed refined alignments of ABCD protein sequences. Based on the 2.85  Å resolution crystal structure of the mitochondrial ABC transporter ABCB10, here we propose a structural model of peroxisomal ABCD proteins that specifies the position of the transmembrane and coupling helices, and highlight functional motifs and putative important amino acid residues. PMID:28737695

  6. Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation

    PubMed Central

    2012-01-01

    Background The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC) Superfamily and Major Facilitator Superfamily (MFS) in S. cerevisiae were scrutinized for a role in ethanol stress resistance. Results A yeast multidrug resistance ABC transporter encoded by the PDR18 gene, proposed to play a role in the incorporation of ergosterol in the yeast plasma membrane, was found to confer resistance to growth inhibitory concentrations of ethanol. PDR18 expression was seen to contribute to decreased 3 H-ethanol intracellular concentrations and decreased plasma membrane permeabilization of yeast cells challenged with inhibitory ethanol concentrations. Given the increased tolerance to ethanol of cells expressing PDR18, the final concentration of ethanol produced during high gravity alcoholic fermentation by yeast cells devoid of PDR18 was lower than the final ethanol concentration produced by the corresponding parental strain. Moreover, an engineered yeast strain in which the PDR18 promoter was replaced in the genome by the stronger PDR5 promoter, leading to increased PDR18 mRNA levels during alcoholic fermentation, was able to attain a 6 % higher ethanol concentration and a 17 % higher ethanol production yield than the parental strain. The improved fermentative performance of yeast cells over-expressing PDR18 was found to correlate with their increased ethanol tolerance and ability to restrain plasma membrane permeabilization induced throughout high gravity fermentation. Conclusions PDR18 gene over-expression increases yeast ethanol tolerance and fermentation performance leading to the production of highly inhibitory concentrations of ethanol. PDR18 overexpression in industrial yeast strains appears to be a promising approach to improve alcoholic

  7. Opioid transport by ATP-binding cassette transporters at the blood-brain barrier: implications for neuropsychopharmacology.

    PubMed

    Tournier, Nicolas; Declèves, Xavier; Saubaméa, Bruno; Scherrmann, Jean-Michel; Cisternino, Salvatore

    2011-01-01

    Some of the ATP-binding cassette (ABC) transporters like P-glycoprotein (P-gp; ABCB1, MDR1), BCRP (ABCG2) and MRPs (ABCCs) that are present at the blood-brain barrier (BBB) influence the brain pharmacokinetics (PK) of their substrates by restricting their uptake or enhancing their clearance from the brain into the blood, which has consequences for their CNS pharmacodynamics (PD). Opioid drugs have been invaluable tools for understanding the PK-PD relationships of these ABC-transporters. The effects of morphine, methadone and loperamide on the CNS are modulated by P-gp. This review examines the ways in which other opioid drugs and some of their active metabolites interact with ABC transporters and suggests new mechanisms that may be involved in the variability of the response of the CNS to these drugs like carrier-mediated system belonging to the solute carrier (SLC) superfamily. Exposure to opioids may also alter the expression of ABC transporters. P-gp can be overproduced during morphine treatment, suggesting that the drug has a direct or, more likely, an indirect action. Variations in cerebral neurotransmitters during exposure to opioids and the release of cytokines during pain could be new endogenous stimuli affecting transporter synthesis. This review concludes with an analysis of the pharmacotherapeutic and clinical impacts of the interactions between ABC transporters and opioids.

  8. Functional diversity of the superfamily of K⁺ transporters to meet various requirements.

    PubMed

    Diskowski, Marina; Mikusevic, Vedrana; Stock, Charlott; Hänelt, Inga

    2015-09-01

    The superfamily of K+ transporters unites proteins from plants, fungi, bacteria, and archaea that translocate K+ and/or Na+ across membranes. These proteins are key components in osmotic regulation, pH homeostasis, and resistance to high salinity and dryness. The members of the superfamily are closely related to K+ channels such as KcsA but also show several striking differences that are attributed to their altered functions. This review highlights these functional differences, focusing on the bacterial superfamily members KtrB, TrkH, and KdpA. The functional variations within the family and comparison to MPM-type K+ channels are discussed in light of the recently solved structures of the Ktr and Trk systems.

  9. Structural basis for amino acid export by DMT superfamily transporter YddG.

    PubMed

    Tsuchiya, Hirotoshi; Doki, Shintaro; Takemoto, Mizuki; Ikuta, Tatsuya; Higuchi, Takashi; Fukui, Keita; Usuda, Yoshihiro; Tabuchi, Eri; Nagatoishi, Satoru; Tsumoto, Kouhei; Nishizawa, Tomohiro; Ito, Koichi; Dohmae, Naoshi; Ishitani, Ryuichiro; Nureki, Osamu

    2016-06-16

    The drug/metabolite transporter (DMT) superfamily is a large group of membrane transporters ubiquitously found in eukaryotes, bacteria and archaea, and includes exporters for a remarkably wide range of substrates, such as toxic compounds and metabolites. YddG is a bacterial DMT protein that expels aromatic amino acids and exogenous toxic compounds, thereby contributing to cellular homeostasis. Here we present structural and functional analyses of YddG. Using liposome-based analyses, we show that Escherichia coli and Starkeya novella YddG export various amino acids. The crystal structure of S. novella YddG at 2.4 Å resolution reveals a new membrane transporter topology, with ten transmembrane segments in an outward-facing state. The overall structure is basket-shaped, with a large substrate-binding cavity at the centre of the molecule, and is composed of inverted structural repeats related by two-fold pseudo-symmetry. On the basis of this intramolecular symmetry, we propose a structural model for the inward-facing state and a mechanism of the conformational change for substrate transport, which we confirmed by biochemical analyses. These findings provide a structural basis for the mechanism of transport of DMT superfamily proteins.

  10. Profiling of ABC transporters ABCB5, ABCF2 and nestin-positive stem cells in nevi, in situ and invasive melanoma.

    PubMed

    Setia, Namrata; Abbas, Ossama; Sousa, Yessica; Garb, Jane L; Mahalingam, Meera

    2012-08-01

    Distinct ABCB5 forms and ABCF2, members of the ATP-binding cassette (ABC) superfamily of transporters, are normally expressed in various tissues and cells, and enhanced expression of both has been demonstrated in select cancers. In melanoma cell lines, gene expression profiling of ABC transporters has revealed enhanced expression of melanocyte-specific ABCB5 and ABCF2 proteins. Given this, our primary aim was to ascertain immunohistochemical expression of the ABC transporters ABCB5 and ABCF2 and, the stem cell marker, nestin in a spectrum of benign and malignant nevomelanocytic proliferations, including nevi (n=30), in situ (n=31) and invasive (n=24) primary cutaneous melanomas to assess their role in the stepwise development of malignancy. In addition, their expression was compared with established melanoma prognosticators to ascertain their utility as independent prognosticators. A semiquantitative scoring system was utilized by deriving a cumulative score (based on percentage positivity cells and intensity of expression) and statistical analyses was carried out using analysis of variance with linear contrasts. Mean cumulative score in nevi, in situ and invasive melanoma were as follows: 3.8, 4.4 and 5.3 for ABCB5, respectively (P<0.005 for all), and 4.6, 4.6 and 5.3 for nestin, respectively (P=not significant for all). No appreciable expression of ABCF2 was noted in any of the groups. While ulcerated lesions of melanoma demonstrated lower levels of expression of ABCB5 and nestin than non-ulcerated lesions, and nestin expression was lower in lesions with mitoses >1, after controlling for the presence of ulceration and mitotic activity, the expression of both proteins did not significantly correlate with known melanoma prognosticators. The gradual increase in the expression of ABCB5 from benign nevus to in situ to invasive melanoma suggests that it plays a role in melanomagenesis. On the basis of our findings, a prospective study with follow-up data is required to

  11. Antibiotic Resistance Mediated by the MacB ABC Transporter Family: A Structural and Functional Perspective

    PubMed Central

    Greene, Nicholas P.; Kaplan, Elise; Crow, Allister; Koronakis, Vassilis

    2018-01-01

    The MacB ABC transporter forms a tripartite efflux pump with the MacA adaptor protein and TolC outer membrane exit duct to expel antibiotics and export virulence factors from Gram-negative bacteria. Here, we review recent structural and functional data on MacB and its homologs. MacB has a fold that is distinct from other structurally characterized ABC transporters and uses a unique molecular mechanism termed mechanotransmission. Unlike other bacterial ABC transporters, MacB does not transport substrates across the inner membrane in which it is based, but instead couples cytoplasmic ATP hydrolysis with transmembrane conformational changes that are used to perform work in the extra-cytoplasmic space. In the MacAB-TolC tripartite pump, mechanotransmission drives efflux of antibiotics and export of a protein toxin from the periplasmic space via the TolC exit duct. Homologous tripartite systems from pathogenic bacteria similarly export protein-like signaling molecules, virulence factors and siderophores. In addition, many MacB-like ABC transporters do not form tripartite pumps, but instead operate in diverse cellular processes including antibiotic sensing, cell division and lipoprotein trafficking. PMID:29892271

  12. Antibiotic Resistance Mediated by the MacB ABC Transporter Family: A Structural and Functional Perspective.

    PubMed

    Greene, Nicholas P; Kaplan, Elise; Crow, Allister; Koronakis, Vassilis

    2018-01-01

    The MacB ABC transporter forms a tripartite efflux pump with the MacA adaptor protein and TolC outer membrane exit duct to expel antibiotics and export virulence factors from Gram-negative bacteria. Here, we review recent structural and functional data on MacB and its homologs. MacB has a fold that is distinct from other structurally characterized ABC transporters and uses a unique molecular mechanism termed mechanotransmission. Unlike other bacterial ABC transporters, MacB does not transport substrates across the inner membrane in which it is based, but instead couples cytoplasmic ATP hydrolysis with transmembrane conformational changes that are used to perform work in the extra-cytoplasmic space. In the MacAB-TolC tripartite pump, mechanotransmission drives efflux of antibiotics and export of a protein toxin from the periplasmic space via the TolC exit duct. Homologous tripartite systems from pathogenic bacteria similarly export protein-like signaling molecules, virulence factors and siderophores. In addition, many MacB-like ABC transporters do not form tripartite pumps, but instead operate in diverse cellular processes including antibiotic sensing, cell division and lipoprotein trafficking.

  13. Transcriptomic insights on the ABC transporter gene family in the salmon louse Caligus rogercresseyi.

    PubMed

    Valenzuela-Muñoz, Valentina; Sturm, Armin; Gallardo-Escárate, Cristian

    2015-04-09

    ATP-binding cassette (ABC) protein family encode for membrane proteins involved in the transport of various biomolecules through the cellular membrane. These proteins have been identified in all taxa and present important physiological functions, including the process of insecticide detoxification in arthropods. For that reason the ectoparasite Caligus rogercresseyi represents a model species for understanding the molecular underpinnings involved in insecticide drug resistance. llumina sequencing was performed using sea lice exposed to 2 and 3 ppb of deltamethrin and azamethiphos. Contigs obtained from de novo assembly were annotated by Blastx. RNA-Seq analysis was performed and validated by qPCR analysis. From the transcriptome database of C. rogercresseyi, 57 putative members of ABC protein sequences were identified and phylogenetically classified into the eight subfamilies described for ABC transporters in arthropods. Transcriptomic profiles for ABC proteins subfamilies were evaluated throughout C. rogercresseyi development. Moreover, RNA-Seq analysis was performed for adult male and female salmon lice exposed to the delousing drugs azamethiphos and deltamethrin. High transcript levels of the ABCB and ABCC subfamilies were evidenced. Furthermore, SNPs mining was carried out for the ABC proteins sequences, revealing pivotal genomic information. The present study gives a comprehensive transcriptome analysis of ABC proteins from C. rogercresseyi, providing relevant information about transporter roles during ontogeny and in relation to delousing drug responses in salmon lice. This genomic information represents a valuable tool for pest management in the Chilean salmon aquaculture industry.

  14. The ABC transporter Rv1272c of Mycobacterium tuberculosis enhances the import of long-chain fatty acids in Escherichia coli.

    PubMed

    Martin, Audrey; Daniel, Jaiyanth

    2018-02-05

    Mycobacterium tuberculosis (Mtb), which causes tuberculosis, is capable of accumulating triacylglycerol (TAG) by utilizing fatty acids from host cells. ATP-binding cassette (ABC) transporters are involved in transport processes in all organisms. Among the classical ABC transporters in Mtb none have been implicated in fatty acid import. Since the transport of fatty acids from the host cell is important for dormancy-associated TAG synthesis in the pathogen, mycobacterial ABC transporter(s) could potentially be involved in this process. Based on sequence identities with a bacterial ABC transporter that mediates fatty acid import for TAG synthesis, we identified Rv1272c, a hitherto uncharacterized ABC-transporter in Mtb that also shows sequence identities with a plant ABC transporter involved in fatty acid transport. We expressed Rv1272c in E. coli and show that it enhances the import of radiolabeled fatty acids. We also show that Rv1272c causes a significant increase in the metabolic incorporation of radiolabeled long-chain fatty acids into cardiolipin, a tetra-acylated phospholipid, and phosphatidylglycerol in E. coli. This is the first report on the function of Rv1272c showing that it displays a long-chain fatty acid transport function. Copyright © 2018 Elsevier Inc. All rights reserved.

  15. A bacterial-type ABC transporter is involved in aluminum tolerance in rice.

    PubMed

    Huang, Chao Feng; Yamaji, Naoki; Mitani, Namiki; Yano, Masahiro; Nagamura, Yoshiaki; Ma, Jian Feng

    2009-02-01

    Aluminum (Al) toxicity is a major factor limiting crop production in acidic soil, but the molecular mechanisms of Al tolerance are poorly understood. Here, we report that two genes, STAR1 (for sensitive to Al rhizotoxicity1) and STAR2, are responsible for Al tolerance in rice. STAR1 encodes a nucleotide binding domain, while STAR2 encodes a transmembrane domain, of a bacterial-type ATP binding cassette (ABC) transporter. Disruption of either gene resulted in hypersensitivity to aluminum toxicity. Both STAR1 and STAR2 are expressed mainly in the roots and are specifically induced by Al exposure. Expression in onion epidermal cells, rice protoplasts, and yeast showed that STAR1 interacts with STAR2 to form a complex that localizes to the vesicle membranes of all root cells, except for those in the epidermal layer of the mature zone. When expressed together in Xenopus laevis oocytes, STAR1/2 shows efflux transport activity specific for UDP-glucose. Furthermore, addition of exogenous UDP-glucose rescued root growth in the star1 mutant exposed to Al. These results indicate that STAR1 and STAR2 form a complex that functions as an ABC transporter, which is required for detoxification of Al in rice. The ABC transporter transports UDP-glucose, which may be used to modify the cell wall.

  16. Zinc transporters YbtX and ZnuABC are required for the virulence of Yersinia pestis in bubonic and pneumonic plague in mice.

    PubMed

    Bobrov, Alexander G; Kirillina, Olga; Fosso, Marina Y; Fetherston, Jacqueline D; Miller, M Clarke; VanCleave, Tiva T; Burlison, Joseph A; Arnold, William K; Lawrenz, Matthew B; Garneau-Tsodikova, Sylvie; Perry, Robert D

    2017-06-21

    A number of bacterial pathogens require the ZnuABC Zinc (Zn 2+ ) transporter and/or a second Zn 2+ transport system to overcome Zn 2+ sequestration by mammalian hosts. Previously we have shown that in addition to ZnuABC, Yersinia pestis possesses a second Zn 2+ transporter that involves components of the yersiniabactin (Ybt), siderophore-dependent iron transport system. Synthesis of the Ybt siderophore and YbtX, a member of the major facilitator superfamily, are both critical components of the second Zn 2+ transport system. Here we demonstrate that a ybtX znu double mutant is essentially avirulent in mouse models of bubonic and pneumonic plague while a ybtX mutant retains high virulence in both plague models. While sequestration of host Zn is a key nutritional immunity factor, excess Zn appears to have a significant antimicrobial role in controlling intracellular bacterial survival. Here, we demonstrate that ZntA, a Zn 2+ exporter, plays a role in resistance to Zn toxicity in vitro, but that a zntA zur double mutant retains high virulence in both pneumonic and bubonic plague models and survival in macrophages. We also confirm that Ybt does not directly bind Zn 2+ in vitro under the conditions tested. However, we detect a significant increase in Zn 2+ -binding ability of filtered supernatants from a Ybt + strain compared to those from a strain unable to produce the siderophore, supporting our previously published data that Ybt biosynthetic genes are involved in the production of a secreted Zn-binding molecule (zincophore). Our data suggest that Ybt or a modified Ybt participate in or promote Zn-binding activity in culture supernatants and is involved in Zn acquisition in Y. pestis.

  17. The Yeast Plasma Membrane ATP Binding Cassette (ABC) Transporter Aus1

    PubMed Central

    Marek, Magdalena; Milles, Sigrid; Schreiber, Gabriele; Daleke, David L.; Dittmar, Gunnar; Herrmann, Andreas; Müller, Peter; Pomorski, Thomas Günther

    2011-01-01

    The ATP binding cassette (ABC) transporter Aus1 is expressed under anaerobic growth conditions at the plasma membrane of the yeast Saccharomyces cerevisiae and is required for sterol uptake. These observations suggest that Aus1 promotes the translocation of sterols across membranes, but the precise transport mechanism has yet to be identified. In this study, an extraction and purification procedure was developed to characterize the Aus1 transporter. The detergent-solubilized protein was able to bind and hydrolyze ATP. Mutagenesis of the conserved lysine to methionine in the Walker A motif abolished ATP hydrolysis. Likewise, ATP hydrolysis was inhibited by classical inhibitors of ABC transporters. Upon reconstitution into proteoliposomes, the ATPase activity of Aus1 was specifically stimulated by phosphatidylserine (PS) in a stereoselective manner. We also found that Aus1-dependent sterol uptake, but not Aus1 expression and trafficking to the plasma membrane, was affected by changes in cellular PS levels. These results suggest a direct interaction between Aus1 and PS that is critical for the activity of the transporter. PMID:21521689

  18. Describing the role of Drosophila melanogaster ABC transporters in insecticide biology using CRISPR-Cas9 knockouts.

    PubMed

    Denecke, Shane; Fusetto, Roberto; Batterham, Philip

    2017-12-01

    ABC transporters have a well-established role in drug resistance, effluxing xenobiotics from cells and tissues within the organism. More recently, research has been dedicated to understanding the role insect ABC transporters play in insecticide toxicity, but progress in understanding the contribution of specific transporters has been hampered by the lack of functional genetic tools. Here, we report knockouts of three Drosophila melanogaster ABC transporter genes, Mdr49, Mdr50, and Mdr65, that are homologous to the well-studied mammalian ABCB1 (P-glycoprotein). Each knockout mutant was created in the same wild type background and tested against a panel of insecticides representing different chemical classes. Mdr65 knockouts were more susceptible to all neuroactive insecticides tested, but Mdr49 and Mdr50 knockouts showed increased susceptibility or resistance depending on the insecticide used. Mdr65 was chosen for further analysis. Calculation of LC 50 values for the Mdr65 knockout allowed the substrate specificity of this transporter to be examined. No obvious distinguishing structural features were shared among MDR65 substrates. A role for Mdr65 in insecticide transport was confirmed by testing the capacity of the knockout to synergize with the ABC inhibitor verapamil and by measuring the levels of insecticide retained in the body of knockout flies. These data unambiguously establish the influence of ABC transporters on the capacity of wild type D. melanogaster to tolerate insecticide exposure and suggest that both tissue and substrate specificity underpin this capacity. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Surface charge-specific interactions between polymer nanoparticles and ABC transporters in Caco-2 cells

    NASA Astrophysics Data System (ADS)

    Bhattacharjee, Sourav; van Opstal, Edward J.; Alink, Gerrit M.; Marcelis, Antonius T. M.; Zuilhof, Han; Rietjens, Ivonne M. C. M.

    2013-06-01

    The surface charge-dependent transport of polymeric nanoparticles (PNPs) across Caco-2 monolayers grown on transwell culture systems as an in vitro model for intestinal transport was tested. The transport of well-characterized, monodisperse, and fluorescent tri-block copolymer nanoparticles (TCNPs/size 45 nm) and polystyrene nanoparticles (PSNPs/size 50 nm), with different surface charges (positive and negative), was quantified. The positive PNPs showed a higher intracellular uptake and flux across the Caco-2 monolayers than the negative PNPs. Multidrug resistance/P-glycoprotein (MDR1/P-gp), a specific ATP-binding cassette (ABC) transporter, was found to play a major role in the cellular efflux of positive PNPs, whereas the multidrug resistance protein 1 took part in the efflux of negative PNPs from Caco-2 cells. The positive PNPs also caused an increased cellular uptake and apical to basolateral transport of the carcinogen PhIP across the Caco-2 monolayer. The flavonoid quercetin, which is known to interact with ABC transporters, promoted the intracellular uptake of different PNPs and interfered with the normal distribution patterns of PNPs in the transwell system. These results indicate that PNPs display surface charge-specific interactions with ABC transporters and can even affect the bioavailability of toxic food-borne compounds (like pro-carcinogens).

  20. Temporal dynamics of the ABC transporter response to insecticide treatment: insights from the malaria vector Anopheles stephensi

    NASA Astrophysics Data System (ADS)

    Epis, Sara; Porretta, Daniele; Mastrantonio, Valentina; Urbanelli, Sandra; Sassera, Davide; De Marco, Leone; Mereghetti, Valeria; Montagna, Matteo; Ricci, Irene; Favia, Guido; Bandi, Claudio

    2014-12-01

    In insects, ABC transporters have been shown to contribute to defence/resistance to insecticides by reducing toxic concentrations in cells/tissues. Despite the extensive studies about this detoxifying mechanism, the temporal patterns of ABC transporter activation have been poorly investigated. Using the malaria vector Anopheles stephensi as a study system, we investigated the expression profile of ABC genes belonging to different subfamilies in permethrin-treated larvae at different time points (30 min to 48 h). Our results showed that the expression of ABCB and ABCG subfamily genes was upregulated at 1 h after treatment, with the highest expression observed at 6 h. Therefore, future investigations on the temporal dynamics of ABC gene expression will allow a better implementation of insecticide treatment regimens, including the use of specific inhibitors of ABC efflux pumps.

  1. Investigation of the quaternary structure of an ABC transporter in living cells using spectrally resolved resonance energy transfer

    NASA Astrophysics Data System (ADS)

    Singh, Deo Raj

    Forster resonance energy transfer (FRET) has become an important tool to study proteins inside living cells. It has been used to explore membrane protein folding and dynamics, determine stoichiometry and geometry of protein complexes, and measure the distance between two molecules. In this dissertation, we use a method based on FRET and optical micro-spectroscopy (OptiMiS) technology, developed in our lab, to probe the structure of dynamic (as opposed to static) protein complexes in living cells. We use this method to determine the association stoichiometry and quaternary structure of an ABC transporter in living cells. Specifically, the transporter we investigate originates from the pathogen Pseudomonas aeruginosa, which is a Gram-negative bacterium with several virulence factors, lipopolysaccharides being one of them. This pathogen coexpresses two unique forms of lipopolysaccharides on its surface, the A- and B-bands. The A-band polysaccharides, synthesized in the cytoplasm, are translocated into the periplasm through an ATP-binding-cassette (ABC) transporter consisting of a transmembranar protein, Wzm, and a nucleotide-binding protein, Wzt. In P. aeruginosa, all of the biochemical studies of A-band LPS are concentrated on the stages of the synthesis and ligation of polysaccharides (PSs), leaving the export stage involving ABC transporter unexplored. The mode of PS export through ABC transporters is still unknown. This difficulty is due to the lack of information about sub-unit composition and structure of this bi-component ABC transporter. Using the FRET-OptiMiS combination method developed by our lab, we found that Wzt forms a rhombus-shaped homo-tetramer which becomes a square upon co-expression with Wzm, and that Wzm forms a square-shaped homo-tetramer both in the presence and absence of Wzt. Based on these results, we propose a structural model for the double-tetramer complex formed by the bi-component ABC transporter in living cells. An understanding of the

  2. Structural basis for lipopolysaccharide extraction by ABC transporter LptB2FG.

    PubMed

    Luo, Qingshan; Yang, Xu; Yu, Shan; Shi, Huigang; Wang, Kun; Xiao, Le; Zhu, Guangyu; Sun, Chuanqi; Li, Tingting; Li, Dianfan; Zhang, Xinzheng; Zhou, Min; Huang, Yihua

    2017-05-01

    After biosynthesis, bacterial lipopolysaccharides (LPS) are transiently anchored to the outer leaflet of the inner membrane (IM). The ATP-binding cassette (ABC) transporter LptB 2 FG extracts LPS molecules from the IM and transports them to the outer membrane. Here we report the crystal structure of nucleotide-free LptB 2 FG from Pseudomonas aeruginosa. The structure reveals that lipopolysaccharide transport proteins LptF and LptG each contain a transmembrane domain (TMD), a periplasmic β-jellyroll-like domain and a coupling helix that interacts with LptB on the cytoplasmic side. The LptF and LptG TMDs form a large outward-facing V-shaped cavity in the IM. Mutational analyses suggest that LPS may enter the central cavity laterally, via the interface of the TMD domains of LptF and LptG, and is expelled into the β-jellyroll-like domains upon ATP binding and hydrolysis by LptB. These studies suggest a mechanism for LPS extraction by LptB 2 FG that is distinct from those of classical ABC transporters that transport substrates across the IM.

  3. Toward Determining ATPase Mechanism in ABC Transporters: Development of the Reaction Path–Force Matching QM/MM Method

    PubMed Central

    Zhou, Y.; Ojeda-May, P.; Nagaraju, M.; Pu, J.

    2016-01-01

    Adenosine triphosphate (ATP)-binding cassette (ABC) transporters are ubiquitous ATP-dependent membrane proteins involved in translocations of a wide variety of substrates across cellular membranes. To understand the chemomechanical coupling mechanism as well as functional asymmetry in these systems, a quantitative description of how ABC transporters hydrolyze ATP is needed. Complementary to experimental approaches, computer simulations based on combined quantum mechanical and molecular mechanical (QM/MM) potentials have provided new insights into the catalytic mechanism in ABC transporters. Quantitatively reliable determination of the free energy requirement for enzymatic ATP hydrolysis, however, requires substantial statistical sampling on QM/MM potential. A case study shows that brute force sampling of ab initio QM/MM (AI/MM) potential energy surfaces is computationally impractical for enzyme simulations of ABC transporters. On the other hand, existing semiempirical QM/MM (SE/MM) methods, although affordable for free energy sampling, are unreliable for studying ATP hydrolysis. To close this gap, a multiscale QM/MM approach named reaction path–force matching (RP–FM) has been developed. In RP–FM, specific reaction parameters for a selected SE method are optimized against AI reference data along reaction paths by employing the force matching technique. The feasibility of the method is demonstrated for a proton transfer reaction in the gas phase and in solution. The RP–FM method may offer a general tool for simulating complex enzyme systems such as ABC transporters. PMID:27498639

  4. GxySBA ABC Transporter of Agrobacterium tumefaciens and Its Role in Sugar Utilization and vir Gene Expression

    PubMed Central

    Zhao, Jinlei

    2014-01-01

    Monosaccharides available in the extracellular milieu of Agrobacterium tumefaciens can be transported into the cytoplasm, or via the periplasmic sugar binding protein, ChvE, play a critical role in controlling virulence gene expression. The ChvE-MmsAB ABC transporter is involved in the utilization of a wide range of monosaccharide substrates but redundant transporters are likely given the ability of a chvE-mmsAB deletion strain to grow, albeit more slowly, in the presence of particular monosaccharides. In this study, a putative ABC transporter encoded by the gxySBA operon is identified and shown to be involved in the utilization of glucose, xylose, fucose, and arabinose, which are also substrates for the ChvE-MmsAB ABC transporter. Significantly, GxySBA is also shown to be the first characterized glucosamine ABC transporter. The divergently transcribed gene gxyR encodes a repressor of the gxySBA operon, the function of which can be relieved by a subset of the transported sugars, including glucose, xylose, and glucosamine, and this substrate-induced expression can be repressed by glycerol. Furthermore, deletion of the transporter can increase the sensitivity of the virulence gene expression system to certain sugars that regulate it. Collectively, the results reveal a remarkably diverse set of substrates for the GxySBA transporter and its contribution to the repression of sugar sensitivity by the virulence-controlling system, thereby facilitating the capacity of the bacterium to distinguish between the soil and plant environments. PMID:24957625

  5. The Yersinia pestis Siderophore, Yersiniabactin, and the ZnuABC system both contribute to Zinc acquisition and the development of lethal septicemic plague in mice

    PubMed Central

    Bobrov, Alexander G.; Kirillina, Olga; Fetherston, Jacqueline D.; Miller, M. Clarke; Burlison, Joseph A.; Perry, Robert D.

    2014-01-01

    Summary Bacterial pathogens must overcome host sequestration of zinc (Zn2+), an essential micronutrient, during the infectious disease process. While the mechanisms to acquire chelated Zn2+ by bacteria are largely undefined, many pathogens rely upon the ZnuABC family of ABC transporters. Here we show that in Yersinia pestis, irp2, a gene encoding the synthetase (HMWP2) for the siderophore yersiniabactin (Ybt) is required for growth under Zn2+-deficient conditions in a strain lacking ZnuABC. Moreover, growth stimulation with exogenous, purified apo-Ybt provides evidence that Ybt may serve as a zincophore for Zn2+ acquisition. Studies with the Zn2+-dependent transcriptional reporter znuA∷lacZ indicate that the ability to synthesize Ybt affects the levels of intracellular Zn2+. However, the outer membrane receptor Psn and TonB as well as the inner membrane (IM) ABC transporter YbtPQ, that are required for Fe3+ acquisition by Ybt, are not needed for Ybt-dependent Zn2+ uptake. In contrast, the predicted IM protein YbtX, a member of the Major Facilitator Superfamily, was essential for Ybt-dependent Zn2+ uptake. Finally, we show that the ZnuABC system and the Ybt synthetase HMWP2, presumably by Ybt synthesis, both contribute to the development of a lethal infection in a septicemic plague mouse model. PMID:24979062

  6. The Yersinia pestis siderophore, yersiniabactin, and the ZnuABC system both contribute to zinc acquisition and the development of lethal septicaemic plague in mice.

    PubMed

    Bobrov, Alexander G; Kirillina, Olga; Fetherston, Jacqueline D; Miller, M Clarke; Burlison, Joseph A; Perry, Robert D

    2014-08-01

    Bacterial pathogens must overcome host sequestration of zinc (Zn(2+) ), an essential micronutrient, during the infectious disease process. While the mechanisms to acquire chelated Zn(2+) by bacteria are largely undefined, many pathogens rely upon the ZnuABC family of ABC transporters. Here we show that in Yersinia pestis, irp2, a gene encoding the synthetase (HMWP2) for the siderophore yersiniabactin (Ybt) is required for growth under Zn(2+) -deficient conditions in a strain lacking ZnuABC. Moreover, growth stimulation with exogenous, purified apo-Ybt provides evidence that Ybt may serve as a zincophore for Zn(2+) acquisition. Studies with the Zn(2+) -dependent transcriptional reporter znuA::lacZ indicate that the ability to synthesize Ybt affects the levels of intracellular Zn(2+) . However, the outer membrane receptor Psn and TonB as well as the inner membrane (IM) ABC transporter YbtPQ, which are required for Fe(3+) acquisition by Ybt, are not needed for Ybt-dependent Zn(2+) uptake. In contrast, the predicted IM protein YbtX, a member of the Major Facilitator Superfamily, was essential for Ybt-dependent Zn(2+) uptake. Finally, we show that the ZnuABC system and the Ybt synthetase HMWP2, presumably by Ybt synthesis, both contribute to the development of a lethal infection in a septicaemic plague mouse model. © 2014 John Wiley & Sons Ltd.

  7. Nucleotide-induced conformational dynamics in ABC transporters from structure-based coarse grained modelling.

    NASA Astrophysics Data System (ADS)

    Flechsig, Holger

    2016-02-01

    ATP-binding cassette (ABC) transporters are integral membrane proteins which mediate the exchange of diverse substrates across membranes powered by ATP molecules. Our understanding of their activity is still hampered since the conformational dynamics underlying the operation of such proteins cannot yet be resolved in detailed molecular dynamics studies. Here a coarse grained model which allows to mimic binding of nucleotides and follow subsequent conformational motions of full-length transporter structures in computer simulations is proposed and implemented. To justify its explanatory quality, the model is first applied to the maltose transporter system for which multiple conformations are known and we find that the model predictions agree remarkably well with the experimental data. For the MalK subunit the switching from open to the closed dimer configuration upon ATP binding is reproduced and, moreover, for the full-length maltose transporter, progression from inward-facing to the outward-facing state is correctly obtained. For the heme transporter HmuUV, for which only the free structure could yet be determined, the model was then applied to predict nucleotide-induced conformational motions. Upon binding of ATP-mimicking ligands the structure changed from a conformation in which the nucleotide-binding domains formed an open shape, to a conformation in which they were found in tight contact, while, at the same time, a pronounced rotation of the transmembrane domains was observed. This finding is supported by normal mode analysis, and, comparison with structural data of the homologous vitamin B12 transporter BtuCD suggests that the observed rotation mechanism may contribute a common functional aspect for this class of ABC transporters. Although in HmuuV noticeable rearrangement of essential transmembrane helices was detected, there are no indications from our simulations that ATP binding alone may facilitate propagation of substrate molecules in this transporter

  8. The SLCO (former SLC21) superfamily of transporters.

    PubMed

    Hagenbuch, Bruno; Stieger, Bruno

    2013-01-01

    The members of the organic anion transporting polypeptide superfamily (OATPs) are classified within the SLCO solute carrier family. All functionally well characterized members are predicted to have 12 transmembrane domains and are sodium-independent transport systems that mediate the transport of a broad range of endo- as well as xenobiotics. Substrates are mainly amphipathic organic anions with a molecular weight of more than 300Da, but some of the known transported substrates are also neutral or even positively charged. Among the well characterized substrates are numerous drugs including statins, angiotensin-converting enzyme inhibitors, angiotensin receptor blockers, antibiotics, antihistaminics, antihypertensives and anticancer drugs. Based on their amino acid sequence identities, the different OATPs cluster into families (in general with more than 40% amino acid sequence identity) and subfamilies (more than 60% amino acid identity). With the sequencing of genomes from different species and the computerized prediction of encoded proteins more than 300 OATPs can be found in the databases, however only a fraction of them have been identified in humans, rodents, and some additional species important for pharmaceutical research like the rhesus monkey (Macaca mulatta), the dog (Canis lupus familiaris) and the pig (Sus scrofa). These OATPs form 6 families (OATP1-OATP6) and 13 subfamilies. In this review we try to summarize what is currently known about OATPs with respect to endogenous substrates, tissue distribution, transport mechanisms, regulation of expression, structure-function relationship and mutations and polymorphisms. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. Sulfadiazine resistance in Toxoplasma gondii: no involvement of overexpression or polymorphisms in genes of therapeutic targets and ABC transporters

    PubMed Central

    Doliwa, Christelle; Escotte-Binet, Sandie; Aubert, Dominique; Sauvage, Virginie; Velard, Frédéric; Schmid, Aline; Villena, Isabelle

    2013-01-01

    Several treatment failures have been reported for the treatment of toxoplasmic encephalitis, chorioretinitis, and congenital toxoplasmosis. Recently we found three Toxoplasma gondii strains naturally resistant to sulfadiazine and we developed in vitro two sulfadiazine resistant strains, RH-RSDZ and ME-49-RSDZ, by gradual pressure. In Plasmodium, common mechanisms of drug resistance involve, among others, mutations and/or amplification within genes encoding the therapeutic targets dhps and dhfr and/or the ABC transporter genes family. To identify genotypic and/or phenotypic markers of resistance in T. gondii, we sequenced and analyzed the expression levels of therapeutic targets dhps and dhfr, three ABC genes, two Pgp, TgABC.B1 and TgABC.B2, and one MRP, TgABC.C1, on sensitive strains compared to sulfadiazine resistant strains. Neither polymorphism nor overexpression was identified. Contrary to Plasmodium, in which mutations and/or overexpression within gene targets and ABC transporters are involved in antimalarial resistance, T. gondii sulfadiazine resistance is not related to these toxoplasmic genes studied. PMID:23707894

  10. The ABC of Biofilm Drug Tolerance: the MerR-Like Regulator BrlR Is an Activator of ABC Transport Systems, with PA1874-77 Contributing to the Tolerance of Pseudomonas aeruginosa Biofilms to Tobramycin.

    PubMed

    Poudyal, Bandita; Sauer, Karin

    2018-02-01

    A hallmark of biofilms is their tolerance to killing by antimicrobial agents. In Pseudomonas aeruginosa , biofilm drug tolerance requires the c-di-GMP-responsive MerR transcriptional regulator BrlR. However, the mechanism by which BrlR mediates biofilm drug tolerance has not been elucidated. Here, we demonstrate that BrlR activates the expression of at least 7 ABC transport systems, including the PA1874-PA1875-PA1876-PA1877 (PA1874-77) operon, with chromatin immunoprecipitation and DNA binding assays confirming BrlR binding to the promoter region of PA1874-77. Insertional inactivation of the 7 ABC transport systems rendered P. aeruginosa PAO1 biofilms susceptible to tobramycin or norfloxacin. Susceptibility was linked to drug accumulation, with BrlR contributing to norfloxacin accumulation in a manner dependent on multidrug efflux pumps and the PA1874-77 ABC transport system. Inactivation of the respective ABC transport system, furthermore, eliminated the recalcitrance of biofilms to killing by tobramycin but not norfloxacin, indicating that drug accumulation is not linked to biofilm drug tolerance. Our findings indicate for the first time that BrlR, a MerR-type transcriptional activator, activates genes encoding several ABC transport systems, in addition to multiple multidrug efflux pump genes. Moreover, our data confirm a BrlR target contributing to drug tolerance, likely countering the prevailing dogma that biofilm tolerance arises from a multiplicity of factors. Copyright © 2018 American Society for Microbiology.

  11. Functional assignment of solute-binding proteins of ABC transporters using a fluorescence-based thermal shift assay.

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Giulliani, S. E.; Frank, A. E.; Collart, F. R.

    2008-12-08

    We have used a fluorescence-based thermal shift (FTS) assay to identify amino acids that bind to solute-binding proteins in the bacterial ABC transporter family. The assay was validated with a set of six proteins with known binding specificity and was consistently able to map proteins with their known binding ligands. The assay also identified additional candidate binding ligands for several of the amino acid-binding proteins in the validation set. We extended this approach to additional targets and demonstrated the ability of the FTS assay to unambiguously identify preferential binding for several homologues of amino acid-binding proteins with known specificity andmore » to functionally annotate proteins of unknown binding specificity. The assay is implemented in a microwell plate format and provides a rapid approach to validate an anticipated function or to screen proteins of unknown function. The ABC-type transporter family is ubiquitous and transports a variety of biological compounds, but the current annotation of the ligand-binding proteins is limited to mostly generic descriptions of function. The results illustrate the feasibility of the FTS assay to improve the functional annotation of binding proteins associated with ABC-type transporters and suggest this approach that can also be extended to other protein families.« less

  12. Intracellular Transport and Kinesin Superfamily Proteins: Structure, Function and Dynamics

    NASA Astrophysics Data System (ADS)

    Hirokawa, N.; Takemura, R.

    Using various molecular cell biological and molecular genetic approaches, we identified kinesin superfamily proteins (KIFs) and characterized their significant functions in intracellular transport, which is fundamental for cellular morphogenesis, functioning, and survival. We showed that KIFs not only transport various membranous organelles, proteins complexes and mRNAs fundamental for cellular functions but also play significant roles in higher brain functions such as memory and learning, determination of important developmental processes such as left-right asymmetry formation and brain wiring. We also elucidated that KIFs recognize and bind to their specific cargoes using scaffolding or adaptor protein complexes. Concerning the mechanism of motility, we discovered the simplest unique monomeric motor KIF1A and determined by molecular biophysics, cryoelectron microscopy and X-ray crystallography that KIF1A can move on a microtubule processively as a monomer by biased Brownian motion and by hydolyzing ATP.

  13. Proton-coupled sugar transport in the prototypical major facilitator superfamily protein XylE

    PubMed Central

    Wisedchaisri, Goragot; Park, Min-Sun; Iadanza, Matthew G.; Zheng, Hongjin; Gonen, Tamir

    2014-01-01

    The major facilitator superfamily (MFS) is the largest collection of structurally related membrane proteins that transport a wide array of substrates. The proton-coupled sugar transporter XylE is the first member of the MFS that has been structurally characterized in multiple transporting conformations, including both the outward and inward-facing states. Here we report the crystal structure of XylE in a new inward-facing open conformation, allowing us to visualize the rocker-switch movement of the N-domain against the C-domain during the transport cycle. Using molecular dynamics simulation, and functional transport assays, we describe the movement of XylE that facilitates sugar translocation across a lipid membrane and identify the likely candidate proton-coupling residues as the conserved Asp27 and Arg133. This study addresses the structural basis for proton-coupled substrate transport and release mechanism for the sugar porter family of proteins. PMID:25088546

  14. Molecular identification of an ABC transporter complex for manganese: analysis of a cyanobacterial mutant strain impaired in the photosynthetic oxygen evolution process.

    PubMed Central

    Bartsevich, V V; Pakrasi, H B

    1995-01-01

    During photosynthesis, the photosystem II (PSII) pigment-protein complex catalyzes oxygen evolution, a reaction in which a four-manganese ensemble plays a crucial role. Using a newly developed selection scheme, we have isolated BP13, a random photosynthesis-deficient mutant strain of the cyanobacterium, Synechocystis 6803. This mutant grew slowly under photoautotrophic conditions, and had a low oxygen evolution activity. Biochemical analysis revealed that the lesion in this mutant strain had specifically affected the Mn ensemble in PSII. Interestingly, incubation of BP13 cells with micromolar levels of added Mn induced rapid recovery of oxygen evolution activity. The mutant could be complemented with a fragment of wild-type chromosomal DNA containing three closely linked genes, mntA, mntB and mntC. These gene products showed significant sequence similarities with polypeptide components of bacterial permeases that are members of the 'ABC (ATP binding cassette) superfamily' of transporter proteins. We determined that in the BP13 strain, a single nucleotide change had resulted in the replacement of an alanine by an aspartic acid residue in MntA, a soluble protein containing ATP binding motifs. These results suggest that the mntCAB gene cluster encodes polypeptide components of a Mn transporter, the first such protein complex identified in any organism. PMID:7743991

  15. Pharmacotherapy in pregnancy; effect of ABC and SLC transporters on drug transport across the placenta and fetal drug exposure.

    PubMed

    Staud, Frantisek; Cerveny, Lukas; Ceckova, Martina

    2012-11-01

    Pharmacotherapy during pregnancy is often inevitable for medical treatment of the mother, the fetus or both. The knowledge of drug transport across placenta is, therefore, an important topic to bear in mind when deciding treatment in pregnant women. Several drug transporters of the ABC and SLC families have been discovered in the placenta, such as P-glycoprotein, breast cancer resistance protein, or organic anion/cation transporters. It is thus evident that the passage of drugs across the placenta can no longer be predicted simply on the basis of their physical-chemical properties. Functional expression of placental drug transporters in the trophoblast and the possibility of drug-drug interactions must be considered to optimize pharmacotherapy during pregnancy. In this review we summarize current knowledge on the expression and function of ABC and SLC transporters in the trophoblast. Furthermore, we put this data into context with medical conditions that require maternal and/or fetal treatment during pregnancy, such as gestational diabetes, HIV infection, fetal arrhythmias and epilepsy. Proper understanding of the role of placental transporters should be of great interest not only to clinicians but also to pharmaceutical industry for future drug design and development to control the degree of fetal exposure.

  16. The ABC transporter Tba of Amycolatopsis balhimycina is required for efficient export of the glycopeptide antibiotic balhimycin.

    PubMed

    Menges, R; Muth, G; Wohlleben, W; Stegmann, E

    2007-11-01

    All known gene clusters for glycopeptide antibiotic biosynthesis contain a conserved gene supposed to encode an ABC-transporter. In the balhimycin-producer Amycolatopsis balhimycina this gene (tba) is localised between the prephenate dehydrogenase gene pdh and the peptide synthetase gene bpsA. Inactivation of tba in A. balhimycina by gene replacement did not interfere with growth and did not affect balhimycin resistance. However, in the supernatant of the tba mutant RM43 less balhimycin was accumulated compared to the wild type; and the intra-cellular balhimycin concentration was ten times higher in the tba mutant RM43 than in the wild type. These data suggest that the ABC transporter encoded in the balhimycin biosynthesis gene cluster is not involved in resistance but is required for the efficient export of the antibiotic. To elucidate the activity of Tba it was heterologously expressed in Escherichia coli with an N-terminal His-tag and purified by nickel chromatography. A photometric assay revealed that His(6)-Tba solubilised in dodecylmaltoside possesses ATPase activity, characteristic for ABC-transporters.

  17. The mitochondrial transporter ABC-me (ABCB10), a downstream target of GATA-1, is essential for erythropoiesis in vivo.

    PubMed

    Hyde, B B; Liesa, M; Elorza, A A; Qiu, W; Haigh, S E; Richey, L; Mikkola, H K; Schlaeger, T M; Shirihai, O S

    2012-07-01

    The mitochondrial transporter ATP binding cassette mitochondrial erythroid (ABC-me/ABCB10) is highly induced during erythroid differentiation by GATA-1 and its overexpression increases hemoglobin production rates in vitro. However, the role of ABC-me in erythropoiesis in vivo is unknown. Here we report for the first time that erythrocyte development in mice requires ABC-me. ABC-me-/- mice die at day 12.5 of gestation, showing nearly complete eradication of primitive erythropoiesis and lack of hemoglobinized cells at day 10.5. ABC-me-/- erythroid cells fail to differentiate because they exhibit a marked increase in apoptosis, both in vivo and ex vivo. Erythroid precursors are particularly sensitive to oxidative stress and ABC-me in the heart and its yeast ortholog multidrug resistance-like 1 have been shown to protect against oxidative stress. Thus, we hypothesized that increased apoptosis in ABC-me-/- erythroid precursors was caused by oxidative stress. Within this context, ABC-me deletion causes an increase in mitochondrial superoxide production and protein carbonylation in erythroid precursors. Furthermore, treatment of ABC-me-/- erythroid progenitors with the mitochondrial antioxidant MnTBAP (superoxide dismutase 2 mimetic) supports survival, ex vivo differentiation and increased hemoglobin production. Altogether, our findings demonstrate that ABC-me is essential for erythropoiesis in vivo.

  18. The mitochondrial transporter ABC-me (ABCB10), a downstream target of GATA-1, is essential for erythropoiesis in vivo

    PubMed Central

    Hyde, B B; Liesa, M; Elorza, A A; Qiu, W; Haigh, S E; Richey, L; Mikkola, H K; Schlaeger, T M; Shirihai, O S

    2012-01-01

    The mitochondrial transporter ATP binding cassette mitochondrial erythroid (ABC-me/ABCB10) is highly induced during erythroid differentiation by GATA-1 and its overexpression increases hemoglobin production rates in vitro. However, the role of ABC-me in erythropoiesis in vivo is unknown. Here we report for the first time that erythrocyte development in mice requires ABC-me. ABC-me−/− mice die at day 12.5 of gestation, showing nearly complete eradication of primitive erythropoiesis and lack of hemoglobinized cells at day 10.5. ABC-me−/− erythroid cells fail to differentiate because they exhibit a marked increase in apoptosis, both in vivo and ex vivo. Erythroid precursors are particularly sensitive to oxidative stress and ABC-me in the heart and its yeast ortholog multidrug resistance-like 1 have been shown to protect against oxidative stress. Thus, we hypothesized that increased apoptosis in ABC-me−/− erythroid precursors was caused by oxidative stress. Within this context, ABC-me deletion causes an increase in mitochondrial superoxide production and protein carbonylation in erythroid precursors. Furthermore, treatment of ABC-me−/− erythroid progenitors with the mitochondrial antioxidant MnTBAP (superoxide dismutase 2 mimetic) supports survival, ex vivo differentiation and increased hemoglobin production. Altogether, our findings demonstrate that ABC-me is essential for erythropoiesis in vivo. PMID:22240895

  19. Novel understanding of ABC transporters ABCB1/MDR/P-glycoprotein, ABCC2/MRP2, and ABCG2/BCRP in colorectal pathophysiology

    PubMed Central

    Andersen, Vibeke; Svenningsen, Katrine; Knudsen, Lina Almind; Hansen, Axel Kornerup; Holmskov, Uffe; Stensballe, Allan; Vogel, Ulla

    2015-01-01

    AIM: To evaluate ATP-binding cassette (ABC) transporters in colonic pathophysiology as they had recently been related to colorectal cancer (CRC) development. METHODS: Literature search was conducted on PubMed using combinations of the following terms: ABC transporters, ATP binding cassette transporter proteins, inflammatory bowel disease, ulcerative, colitis, Crohns disease, colorectal cancer, colitis, intestinal inflammation, intestinal carcinogenesis, ABCB1/P-glycoprotein (P-gp/CD243/MDR1), ABCC2/multidrug resistance protein 2 (MRP2) and ABCG2/breast cancer resistance protein (BCRP), Abcb1/Mdr1a, abcc2/Mrp2, abcg2/Bcrp, knock-out mice, tight junction, membrane lipid function. RESULTS: Recently, human studies reported that changes in the levels of ABC transporters were early events in the adenoma-carcinoma sequence leading to CRC. A link between ABCB1, high fat diet and gut microbes in relation to colitis was suggested by the animal studies. The finding that colitis was preceded by altered gut bacterial composition suggests that deletion of Abcb1 leads to fundamental changes of host-microbiota interaction. Also, high fat diet increases the frequency and severity of colitis in specific pathogen-free Abcb1 KO mice. The Abcb1 KO mice might thus serve as a model in which diet/environmental factors and microbes may be controlled and investigated in relation to intestinal inflammation. Potential molecular mechanisms include defective transport of inflammatory mediators and/or phospholipid translocation from one side to the other of the cell membrane lipid bilayer by ABC transporters affecting inflammatory response and/or function of tight junctions, phagocytosis and vesicle trafficking. Also, diet and microbes give rise to molecules which are potential substrates for the ABC transporters and which may additionally affect ABC transporter function through nuclear receptors and transcriptional regulation. Another critical role of ABCB1 was suggested by the finding that

  20. Mechanistic determinants of the directionality and energetics of active export by a heterodimeric ABC transporter

    DOE PAGES

    Grossmann, Nina; Vakkasoglu, Ahmet S.; Hulpke, Sabine; ...

    2014-11-07

    The ATP-binding cassette (ABC) transporter associated with antigen processing (TAP) participates in immune surveillance by moving proteasomal products into the endoplasmic reticulum (ER) lumen for major histocompatibility complex class I loading and cell surface presentation to cytotoxic T cells. Here we delineate the mechanistic basis for antigen translocation. Notably, TAP works as a molecular diode, translocating peptide substrates against the gradient in a strict unidirectional way. We reveal the importance of the D-loop at the dimer interface of the two nucleotide-binding domains (NBDs) in coupling substrate translocation with ATP hydrolysis and defining transport vectoriality. Substitution of the converved aspartate, whichmore » coordinates the ATP-binding site, decreases NBD dimerization affinity and turns the unidirectional primary active pump into a passive bidirectional nucleotide-gated facilitator. Thus, ATP hydrolysis is not required for translocation per se, but is essential for both active and unidirectional transport. As a result, our data provide detailed mechanistic insight into how heterodimeric ABC exporters operate.« less

  1. Iowa ABC connections : [tech transfer summary].

    DOT National Transportation Integrated Search

    2015-06-01

    The Iowa Department of Transportation (DOT) and other organizations have : been developing accelerated bridge construction (ABC) concepts, details, and : processes, and Iowa has come to be viewed as a national leader in the area of : ABC. However, th...

  2. Effect of β-elemene on the kinetics of intracellular transport of d-luciferin potassium salt (ABC substrate) in doxorubicin-resistant breast cancer cells and the associated molecular mechanism.

    PubMed

    Tang, Chao-Yuan; Zhu, Li-Xin; Yu, Jian-Dong; Chen, Zhi; Gu, Man-Cang; Mu, Chao-Feng; Liu, Qi; Xiong, Yang

    2018-07-30

    In order to explore the mechanism of the reversing multidrug resistance (MDR) phenotypes by β-elemene (β-ELE) in doxorubicin (DOX)-resistant breast cancer cells (MCF-7/DOX), both the functionality and quantity of the ABC transporters in MCF-7/DOX were studied. Bioluminescence imaging (BLI) was used to study the efflux of d-luciferin potassium salt, the substrate of ATP-binding cassette transporters (ABC transporters), in MCF-7/DOX cells treated by β-ELE. At the same time three major ABC transport proteins and genes-related MDR, P-glycoprotein (P-gp, ABCB1) and multidrug resistance-associated protein 1 (MRP, ABCC1) as well as breast cancer resistance protein (BCRP, ABCG2) were analyzed by q-PCR and Western blot. To investigate the efflux functionality of ABC transporters, MCF-7/DOX Fluc cell line with stably-overexpressed luciferase was established. BLI was then used to real-time monitor the efflux kinetics of d-luciferin potassium salt before and after MCF-7/DOX Fluc cells being treated with β-ELE or not. The results showed that the efflux of d-luciferin potassium salt from MCF-7/DOX Fluc was lessened when pretreated with β-ELE, which means that β-ELE may dampen the functionality of ABC transporters, thus decrease the efflux of d-fluorescein potassium or other chemotherapies which also serve as the substrates of ABC transporters. As the effect of β-ELE on the expression of ABC transporters, the results of q-PCR and Western blot showed that gene and protein expression of ABC transporters such as P-gp, MRP, and BCRP were down-regulated after the treatment of β-ELE. To verify the efficacy of β-ELE on reversing MDR, MCF-7/DOX cells were treated with the combination of DOX and β-ELE. MTT assay showed that β-ELE increased the inhibitory effect of DOX on the proliferation of MCF-7/DOX, and the IC 50 of the combination group was much lower than that of the single DOX or β-ELE treatment. In all, β-ELE may reverse MDR through the substrates of ABC transporters

  3. Drug resistance-associated changes in sphingolipids and ABC transporters occur in different regions of membrane domains.

    PubMed

    Hinrichs, John W J; Klappe, Karin; van Riezen, Manon; Kok, Jan W

    2005-11-01

    We have recently shown that two ATP binding cassette (ABC) transporters are enriched in Lubrol-resistant noncaveolar membrane domains in multidrug-resistant human cancer cells [Hinrichs, J. W. J., K. Klappe, I. Hummel, and J. W. Kok. 2004. ATP-binding cassette transporters are enriched in non-caveolar detergent-insoluble glycosphingolipid-enriched membrane domains (DIGs) in human multidrug-resistant cancer cells. J. Biol. Chem. 279: 5734-5738]. Here, we show that aminophospholipids are relatively enriched in Lubrol-resistant membrane domains compared with Triton X-100-resistant membrane domains, whereas sphingolipids are relatively enriched in the latter. Moreover, Lubrol-resistant membrane domains contain more protein and lipid mass. Based on these results, we postulate a model for detergent-insoluble glycosphingolipid-enriched membrane domains consisting of a Lubrol-insoluble/Triton X-100-insoluble region and a Lubrol-insoluble/Triton X-100-soluble region. The latter region contains most of the ABC transporters as well as lipids known to be necessary for their efflux activity. Compared with drug-sensitive cells, the detergent-insoluble glycosphingolipid-enriched membrane domains (DIGs) in drug-resistant cells differ specifically in sphingolipid content and not in protein, phospholipid, or cholesterol content. In drug-resistant cells, sphingolipids with specific fatty acids (especially C24:1) are enriched in these membrane domains. Together, these data show that multidrug resistance-associated changes in both sphingolipids and ABC transporters occur in DIGs, but in different regions of these domains.

  4. Salinomycin overcomes ABC transporter-mediated multidrug and apoptosis resistance in human leukemia stem cell-like KG-1a cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fuchs, Dominik; Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg; Daniel, Volker

    2010-04-16

    Leukemia stem cells are known to exhibit multidrug resistance by expression of ATP-binding cassette (ABC) transporters which constitute transmembrane proteins capable of exporting a wide variety of chemotherapeutic drugs from the cytosol. We show here that human promyeloblastic leukemia KG-1a cells exposed to the histone deacetylase inhibitor phenylbutyrate resemble many characteristics of leukemia stem cells, including expression of functional ABC transporters such as P-glycoprotein, BCRP and MRP8. Consequently, KG-1a cells display resistance to the induction of apoptosis by various chemotherapeutic drugs. Resistance to apoptosis induction by chemotherapeutic drugs can be reversed by cyclosporine A, which effectively inhibits the activity ofmore » P-glycoprotein and BCRP, thus demonstrating ABC transporter-mediated drug resistance in KG-1a cells. However, KG-1a are highly sensitive to apoptosis induction by salinomycin, a polyether ionophore antibiotic that has recently been shown to kill human breast cancer stem cell-like cells and to induce apoptosis in human cancer cells displaying multiple mechanisms of drug and apoptosis resistance. Whereas KG-1a cells can be adapted to proliferate in the presence of apoptosis-inducing concentrations of bortezomib and doxorubicin, salinomycin does not permit long-term adaptation of the cells to apoptosis-inducing concentrations. Thus, salinomycin should be regarded as a novel and effective agent for the elimination of leukemia stem cells and other tumor cells exhibiting ABC transporter-mediated multidrug resistance.« less

  5. Genome-Wide Identification and Expression Profiling of ATP-Binding Cassette (ABC) Transporter Gene Family in Pineapple (Ananas comosus (L.) Merr.) Reveal the Role of AcABCG38 in Pollen Development

    PubMed Central

    Chen, Piaojuan; Li, Yi; Zhao, Lihua; Hou, Zhimin; Yan, Maokai; Hu, Bingyan; Liu, Yanhui; Azam, Syed Muhammad; Zhang, Ziyan; Rahman, Zia ur; Liu, Liping; Qin, Yuan

    2017-01-01

    Pineapple (Ananas comosus L.) cultivation commonly relies on asexual reproduction which is easily impeded by many factors in agriculture production. Sexual reproduction might be a novel approach to improve the pineapple planting. However, genes controlling pineapple sexual reproduction are still remain elusive. In different organisms a conserved superfamily proteins known as ATP binding cassette (ABC) participate in various biological processes. Whereas, till today the ABC gene family has not been identified in pineapple. Here 100 ABC genes were identified in the pineapple genome and grouped into eight subfamilies (5 ABCAs, 20 ABCBs, 16 ABCCs, 2 ABCDs, one ABCEs, 5 ABCFs, 42 ABCGs and 9 ABCIs). Gene expression profiling revealed the dynamic expression pattern of ABC gene family in various tissues and different developmental stages. AcABCA5, AcABCB6, AcABCC4, AcABCC7, AcABCC9, AcABCG26, AcABCG38 and AcABCG42 exhibited preferential expression in ovule and stamen. Over-expression of AcABCG38 in the Arabidopsis double mutant abcg1-2abcg16-2 partially restored its pollen abortion defects, indicating that AcABCG38 plays important roles in pollen development. Our study on ABC gene family in pineapple provides useful information for developing sexual pineapple plantation which could be utilized to improve pineapple agricultural production. PMID:29312399

  6. Genome-Wide Identification and Expression Profiling of ATP-Binding Cassette (ABC) Transporter Gene Family in Pineapple (Ananas comosus (L.) Merr.) Reveal the Role of AcABCG38 in Pollen Development.

    PubMed

    Chen, Piaojuan; Li, Yi; Zhao, Lihua; Hou, Zhimin; Yan, Maokai; Hu, Bingyan; Liu, Yanhui; Azam, Syed Muhammad; Zhang, Ziyan; Rahman, Zia Ur; Liu, Liping; Qin, Yuan

    2017-01-01

    Pineapple ( Ananas comosus L .) cultivation commonly relies on asexual reproduction which is easily impeded by many factors in agriculture production. Sexual reproduction might be a novel approach to improve the pineapple planting. However, genes controlling pineapple sexual reproduction are still remain elusive. In different organisms a conserved superfamily proteins known as ATP binding cassette (ABC) participate in various biological processes. Whereas, till today the ABC gene family has not been identified in pineapple. Here 100 ABC genes were identified in the pineapple genome and grouped into eight subfamilies (5 ABCAs , 20 ABCB s, 16 ABCCs , 2 ABCDs , one ABCEs , 5 ABCFs , 42 ABCGs and 9 ABCIs ). Gene expression profiling revealed the dynamic expression pattern of ABC gene family in various tissues and different developmental stages. AcABCA5, AcABCB6, AcABCC4 , AcABCC7 , AcABCC9 , AcABCG26 , AcABCG38 and AcABCG42 exhibited preferential expression in ovule and stamen. Over-expression of AcABCG38 in the Arabidopsis double mutant abcg1-2abcg16-2 partially restored its pollen abortion defects, indicating that AcABCG38 plays important roles in pollen development. Our study on ABC gene family in pineapple provides useful information for developing sexual pineapple plantation which could be utilized to improve pineapple agricultural production.

  7. Osmoregulated ABC-transport system of Lactococcus lactis senses water stress via changes in the physical state of the membrane.

    PubMed

    van der Heide, T; Poolman, B

    2000-06-20

    An osmoregulated ABC transporter (OpuA) with novel structural features has been identified that responds to water stress. This glycine betaine transport system consists of an ATP-binding/hydrolyzing subunit (OpuAA) and a protein (OpuABC) that contains both the translocator and the substrate-binding domain. The components of OpuA have been overexpressed, purified, and functionally incorporated into liposomes with an ATP-regenerating system in the vesicle lumen. A transmembrane osmotic gradient (outside hyperosmotic relative to the inside) of both ionic and nonionic compounds was able to osmotically activate OpuA in the proteoliposomal system. Hypoosmotic medium conditions inhibited the basal activity of the system. The data show that OpuAA and OpuABC are sufficient for osmoregulated transport, indicating that OpuA can act both as osmosensor and osmoregulator. Strikingly, OpuA could also be activated by low concentrations of cationic and anionic amphipaths, which interact with the membrane. This result indicates that activation by a transmembrane osmotic gradient is mediated by changes in membrane properties/protein-lipid interactions.

  8. Osmoregulated ABC-transport system of Lactococcus lactis senses water stress via changes in the physical state of the membrane

    PubMed Central

    van der Heide, Tiemen; Poolman, Bert

    2000-01-01

    An osmoregulated ABC transporter (OpuA) with novel structural features has been identified that responds to water stress. This glycine betaine transport system consists of an ATP-binding/hydrolyzing subunit (OpuAA) and a protein (OpuABC) that contains both the translocator and the substrate-binding domain. The components of OpuA have been overexpressed, purified, and functionally incorporated into liposomes with an ATP-regenerating system in the vesicle lumen. A transmembrane osmotic gradient (outside hyperosmotic relative to the inside) of both ionic and nonionic compounds was able to osmotically activate OpuA in the proteoliposomal system. Hypoosmotic medium conditions inhibited the basal activity of the system. The data show that OpuAA and OpuABC are sufficient for osmoregulated transport, indicating that OpuA can act both as osmosensor and osmoregulator. Strikingly, OpuA could also be activated by low concentrations of cationic and anionic amphipaths, which interact with the membrane. This result indicates that activation by a transmembrane osmotic gradient is mediated by changes in membrane properties/protein–lipid interactions. PMID:10860977

  9. The ABC transporter ABCG29 is involved in H2O2 tolerance and biocontrol traits in the fungus Clonostachys rosea.

    PubMed

    Dubey, Mukesh; Jensen, Dan Funck; Karlsson, Magnus

    2016-04-01

    For successful biocontrol interactions, biological control organisms must tolerate toxic metabolites produced by themselves or plant pathogens during mycoparasitic/antagonistic interactions, by host plant during colonization of the plant, and xenobiotics present in the environment. ATP-binding cassette (ABC) transporters can play a significant role in tolerance of toxic compounds by mediating active transport across the cellular membrane. This paper reports on functional characterization of an ABC transporter ABCG29 in the biocontrol fungus Clonostachys rosea strain IK726. Gene expression analysis showed induced expression of abcG29 during exposure to the Fusarium spp. mycotoxin zearalenone (ZEA) and the fungicides Cantus, Chipco Green and Apron. Expression of abcG29 in C. rosea was significantly higher during C. rosea-C. rosea (Cr-Cr) interaction or in exposure to C. rosea culture filtrate for 2 h, compared to interaction with Fusarium graminearum or 2 h exposure to F. graminearum culture filtrate. In contrast with gene expression data, ΔabcG29 strains did not display reduced tolerance towards ZEA, fungicides or chemical agents known for inducing oxidative, cell wall or osmotic stress, compared to C. rosea WT. The exception was a significant reduction in tolerance to H2O2 (10 mM) in ΔabcG29 strains when conidia were used as an inoculum. The antagonistic ability of ΔabcG29 strains towards F. graminearum, Fusarium oxysporum or Botrytis cinerea in dual plate assays were not different compared with WT. However, in biocontrol assays ΔabcG29 strains displayed reduced ability to protect Arabidopsis thaliana leaves from B. cinerea, and barley seedling from F. graminearum as measured by an A. thaliana detached leaf assay and a barley foot rot disease assay, respectively. These data show that the ABCG29 is dispensable for ZEA and fungicides tolerance, and antagonism but not H2O2 tolerance and biocontrol effects in C. rosea.

  10. Implications of Mycobacterium Major Facilitator Superfamily for Novel Measures against Tuberculosis.

    PubMed

    Wang, Rui; Zhang, Zhen; Xie, Longxiang; Xie, Jianping

    2015-01-01

    Major facilitator superfamily (MFS) is an important secondary membrane transport protein superfamily conserved from prokaryotes to eukaryotes. The MFS proteins are widespread among bacteria and are responsible for the transfer of substrates. Pathogenic Mycobacterium MFS transporters, their distribution, function, phylogeny, and predicted crystal structures were studied to better understand the function of MFS and to discover specific inhibitors of MFS for better tuberculosis control.

  11. Iowa ABC connections.

    DOT National Transportation Integrated Search

    2015-06-01

    For several years the Iowa Department of Transportation (DOT), Iowa State University, the Federal Highway Administration, : and several Iowa counties have been working to develop accelerated bridge construction (ABC) concepts, details, and processes....

  12. Eukaryotic major facilitator superfamily transporter modeling based on the prokaryotic GlpT crystal structure.

    PubMed

    Lemieux, M Joanne

    2007-01-01

    The major facilitator superfamily (MFS) of transporters represents the largest family of secondary active transporters and has a diverse range of substrates. With structural information for four MFS transporters, we can see a strong structural commonality suggesting, as predicted, a common architecture for MFS transporters. The rate for crystal structure determination of MFS transporters is slow, making modeling of both prokaryotic and eukaryotic transporters more enticing. In this review, models of eukaryotic transporters Glut1, G6PT, OCT1, OCT2 and Pho84, based on the crystal structures of the prokaryotic GlpT, based on the crystal structure of LacY are discussed. The techniques used to generate the different models are compared. In addition, the validity of these models and the strategy of using prokaryotic crystal structures to model eukaryotic proteins are discussed. For comparison, E. coli GlpT was modeled based on the E. coli LacY structure and compared to the crystal structure of GlpT demonstrating that experimental evidence is essential for accurate modeling of membrane proteins.

  13. Barley has two peroxisomal ABC transporters with multiple functions in β-oxidation

    PubMed Central

    Mendiondo, Guillermina M.; Medhurst, Anne; van Roermund, Carlo W.; Zhang, Xuebin; Devonshire, Jean; Scholefield, Duncan; Fernández, José; Axcell, Barry; Ramsay, Luke; Waterham, Hans R.; Waugh, Robbie; Theodoulou, Frederica L.; Holdsworth, Michael J.

    2014-01-01

    In oilseed plants, peroxisomal β-oxidation functions not only in lipid catabolism but also in jasmonate biosynthesis and metabolism of pro-auxins. Subfamily D ATP-binding cassette (ABC) transporters mediate import of β-oxidation substrates into the peroxisome, and the Arabidopsis ABCD protein, COMATOSE (CTS), is essential for this function. Here, the roles of peroxisomal ABCD transporters were investigated in barley, where the main storage compound is starch. Barley has two CTS homologues, designated HvABCD1 and HvABCD2, which are widely expressed and present in embryo and aleurone tissues during germination. Suppression of both genes in barley RNA interference (RNAi) lines indicated roles in metabolism of 2,4-dichlorophenoxybutyrate (2,4-DB) and indole butyric acid (IBA), jasmonate biosynthesis, and determination of grain size. Transformation of the Arabidopsis cts-1 null mutant with HvABCD1 and HvABCD2 confirmed these findings. HvABCD2 partially or completely complemented all tested phenotypes of cts-1. In contrast, HvABCD1 failed to complement the germination and establishment phenotypes of cts-1 but increased the sensitivity of hypocotyls to 100 μM IBA and partially complemented the seed size phenotype. HvABCD1 also partially complemented the yeast pxa1/pxa2Δ mutant for fatty acid β-oxidation. It is concluded that the core biochemical functions of peroxisomal ABC transporters are largely conserved between oilseeds and cereals but that their physiological roles and importance may differ. PMID:24913629

  14. Multiple ABC glucoside transporters mediate sugar-stimulated growth in the heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120.

    PubMed

    Nieves-Morión, Mercedes; Flores, Enrique

    2018-02-01

    Cyanobacteria are generally capable of photoautotrophic growth and are widely distributed on Earth. The model filamentous, heterocyst-forming strain Anabaena sp. PCC 7120 has long been considered as a strict photoautotroph but is now known to be able to assimilate fructose. We have previously described two components of ABC glucoside uptake transporters from Anabaena that are involved in uptake of the sucrose analog esculin: GlsC [a nucleotide-binding domain subunit (NBD)] and GlsP [a transmembrane component (TMD)]. Here, we created Anabaena mutants of genes encoding three further ABC transporter components needed for esculin uptake: GlsD (NBD), GlsQ (TMD) and GlsR (periplasmic substrate-binding protein). Phototrophic growth of Anabaena was significantly stimulated by sucrose, fructose and glucose. Whereas the glsC and glsD mutants were drastically hampered in sucrose-stimulated growth, the different gls mutants were generally impaired in sugar-dependent growth. Our results suggest the participation of Gls and other ABC transporters encoded in the Anabaena genome in sugar-stimulated growth. Additionally, Gls transporter components influence the function of septal junctions in the Anabaena filament. We suggest that mixotrophic growth is important in cyanobacterial physiology and may be relevant for the wide success of these organisms in diverse environments. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  15. ABC transporters and cytochromes P450 in the human central nervous system: influence on brain pharmacokinetics and contribution to neurodegenerative disorders.

    PubMed

    Dutheil, Fabien; Jacob, Aude; Dauchy, Sandrine; Beaune, Philippe; Scherrmann, Jean-Michel; Declèves, Xavier; Loriot, Marie-Anne

    2010-10-01

    The identification of xenobiotic metabolizing enzymes (i.e., CYP) and transporters (i.e., ABC transporters) (XMET) in the human brain, including the BBB, raises the question whether these transporters and enzymes have specific functions in brain physiology, neuropharmacology and toxicology. Relevant literature was identified using PubMed search articles published up to March 2010. Search terms included 'ABC transporters and P450 or CYP', 'drug metabolism, effect and toxicity' and 'neurodegenerative disease (Alzheimer and Parkinson diseases)' restricted to the field of 'brain or human brain'. This review aims to provide a better understanding of XMET functions in the human brain and show their pharmacological importance for improving drug delivery and efficacy and also for managing their side effects. Finally, the impact of brain XMET activity during neurodegenerative processes is discussed, giving an opportunity to identify new markers of human brain diseases. During the last 2 decades, much evidence concerning the specific distribution patterns of XMET, their induction by xenobiotics and endobiotics and their genetic variations have made cerebral ABC transporters and CYP enzymes key elements in the way individual patients respond to centrally acting drugs.

  16. Phosphorylation is required for the pathogen defense function of the Arabidopsis PEN3 ABC transporter

    USDA-ARS?s Scientific Manuscript database

    The Arabidopsis PEN3 ABC transporter accumulates at sites of pathogen detection, where it is involved in defense against multiple pathogens. Perception of PAMPs by pattern recognition receptors initiates recruitment of PEN3 and also leads to PEN3 phosphorylation at multiple amino acid residues. Whet...

  17. Inhibition of the Human ABC Efflux Transporters P-gp and ...

    EPA Pesticide Factsheets

    High body burdens of polybrominated diphenyl ethers (PBDEs) in infants and young children have led to increased concern over their potential impact on human development. PBDE exposure can alter the expression of genes involved in thyroid homeostasis, including those of ATP-binding cassette (ABC) transporters, which mediate cellular xenobiotic efflux. However, little information exists on how PBDEs interact with ABC transporters such as P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). The purpose of this study was to evaluate the interactions of 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) and its hydroxylated metabolite 6-OH-BDE-47 with P-gp and BCRP, using human MDR1- and BCRP-expressing membrane vesicles and stably transfected NIH-3T3-MDR1 and MDCK-BCRP cells. In P-gp membranes, BDE-47 did not affect P-gp activity; however, 6-OH-BDE-47 inhibited P-gp activity at low µM concentrations (IC50 = 11.7 µM). In BCRP membranes, BDE-47 inhibited BCRP activity; however, 6-OH-BDE-47 was a stronger inhibitor [IC50 = 45.9 µM (BDE-47) vs. IC50 = 9.4 µM (6-OH-BDE-47)]. Intracellular concentrations of known P-gp and BCRP substrates [(3H)-paclitaxel and (3H)-prazosin, respectively] were significantly higher (indicating less efflux) in NIH-3T3-MDR1 and MDCK-BCRP cells in the presence of 6-OH-BDE-47, but not BDE-47. Collectively, our results indicate that the BDE-47 metabolite 6-OH-BDE-47 is an inhibitor of both P-gp and BCRP efflux activity.

  18. NtPDR3, an iron-deficiency inducible ABC transporter in Nicotiana tabacum.

    PubMed

    Ducos, Eric; Fraysse, Staffan; Boutry, Marc

    2005-12-19

    In plants, the ABC transporter PDR (pleiotropic drug resistance) subfamily is composed of approximately 15 genes, few of which have been analyzed. We have identified NtPDR3, a Nicotiana tabacum PDR gene belonging to a cluster for which no functional data was previously available. NtPDR3 was found to be induced in suspension cells treated with methyl jasmonate, salicylic acid, 1-naphthalene acetic acid, or cembrene, a macrocyclic diterpene. In agreement with the identification of a putative iron deficiency element in the NtPDR3 transcription promoter region, we found that iron deficiency in the culture medium induced NtPDR3 expression, thus suggesting a new function of the PDR transporter family.

  19. Effects of selected OATP and/or ABC transporter inhibitors on the brain and whole-body distribution of glyburide.

    PubMed

    Tournier, Nicolas; Saba, Wadad; Cisternino, Salvatore; Peyronneau, Marie-Anne; Damont, Annelaure; Goutal, Sébastien; Dubois, Albertine; Dollé, Frédéric; Scherrmann, Jean-Michel; Valette, Héric; Kuhnast, Bertrand; Bottlaender, Michel

    2013-10-01

    Glyburide (glibenclamide, GLB) is a widely prescribed antidiabetic with potential beneficial effects in central nervous system injury and diseases. In vitro studies show that GLB is a substrate of organic anion transporting polypeptide (OATP) and ATP-binding cassette (ABC) transporter families, which may influence GLB distribution and pharmacokinetics in vivo. In the present study, we used [(11)C]GLB positron emission tomography (PET) imaging to non-invasively observe the distribution of GLB at a non-saturating tracer dose in baboons. The role of OATP and P-glycoprotein (P-gp) in [(11)C]GLB whole-body distribution, plasma kinetics, and metabolism was assessed using the OATP inhibitor rifampicin and the dual OATP/P-gp inhibitor cyclosporine. Finally, we used in situ brain perfusion in mice to pinpoint the effect of ABC transporters on GLB transport at the blood-brain barrier (BBB). PET revealed the critical role of OATP on liver [(11)C]GLB uptake and its subsequent impact on [(11)C]GLB metabolism and plasma clearance. OATP-mediated uptake also occurred in the myocardium and kidney parenchyma but not the brain. The inhibition of P-gp in addition to OATP did not further influence [(11)C]GLB tissue and plasma kinetics. At the BBB, the inhibition of both P-gp and breast cancer resistance protein (BCRP) was necessary to demonstrate the role of ABC transporters in limiting GLB brain uptake. This study demonstrates that GLB distribution, metabolism, and elimination are greatly dependent on OATP activity, the first step in GLB hepatic clearance. Conversely, P-gp, BCRP, and probably multidrug resistance protein 4 work in synergy to limit GLB brain uptake.

  20. NFκBP65 transcription factor modulates resistance to doxorubicin through ABC transporters in breast cancer.

    PubMed

    Velaei, Kobra; Samadi, Nasser; Soltani, Sina; Barazvan, Balal; Soleimani Rad, Jafar

    2017-07-01

    Shedding light on chemoresistance biology of breast cancer could contribute to enhance the clinical outcome. Intrinsic or acquired resistance to chemotherapy is a major problem in breast cancer treatment. The NFκB pathway by siRNAP65 and JSH-23 as a translocational inhibitor of NFκBP65 in the doxorubicin-resistant MCF-7 (MCF-7/Dox) and MCF-7 cells was blocked. Then, the ABC transporter expression and function were assessed by real-time qRT-PCR and flow cytometry, respectively. Induction of apoptosis was evaluated after inhibition of the NFΚB pathway as well. Our study underlined the upregulation of NFκBP65 and anti-apoptotic Bcl-2 and downregulation of pro-apoptotic Bax in the MCF-7/Dox cells compared with control MCF-7 cells. Here, we showed that interplay between nuclear factor kappa B P65 (NFkBP65) as a transcriptional regulator and ABC transporters in the MCF-7/Dox cancer cells. We found that inhibition of the elevated expression of NFκBP65 in the resistant breast cancer, whether translocational inhibition or silencing by siRNA, decreased the expression and function of MDR1 and MRP1 efflux pumps. Furthermore, the blockade of NFκBP65 promoted apoptosis via modulating Bcl-2 and BAX expression. After inhibition of the NFκBP65 signaling pathway, elevated baseline expression of survival Bcl-2 gene in the resistant breast cells significantly decreased. Suppression of the NFκB pathway has a profound dual impact on promoting the intrinsic apoptotic pathway and reducing ABC transporter function and expression, which are some of the chemoresistance features. It was speculated that the NFκB pathway directly acts on doxorubicin-induced MDR1 and MRP1 expression in MCF-7/Dox cells.

  1. Two Major Facilitator Superfamily Sugar Transporters from Trichoderma reesei and Their Roles in Induction of Cellulase Biosynthesis*

    PubMed Central

    Zhang, Weixin; Kou, Yanbo; Xu, Jintao; Cao, Yanli; Zhao, Guolei; Shao, Jing; Wang, Hai; Wang, Zhixing; Bao, Xiaoming; Chen, Guanjun; Liu, Weifeng

    2013-01-01

    Proper perception of the extracellular insoluble cellulose is key to initiating the rapid synthesis of cellulases by cellulolytic Trichoderma reesei. Uptake of soluble oligosaccharides derived from cellulose hydrolysis represents a potential point of control in the induced cascade. In this study, we identified a major facilitator superfamily sugar transporter Stp1 capable of transporting cellobiose by reconstructing a cellobiose assimilation system in Saccharomyces cerevisiae. The absence of Stp1 in T. reesei resulted in differential cellulolytic response to Avicel versus cellobiose. Transcriptional profiling revealed a different expression profile in the Δstp1 strain from that of wild-type strain in response to Avicel and demonstrated that Stp1 somehow repressed induction of the bulk of major cellulase and hemicellulose genes. Two other putative major facilitator superfamily sugar transporters were, however, up-regulated in the profiling. Deletion of one of them identified Crt1 that was required for growth and enzymatic activity on cellulose or lactose, but was not required for growth or hemicellulase activity on xylan. The essential role of Crt1 in cellulase induction did not seem to rely on its transporting activity because the overall uptake of cellobiose or sophorose by T. reesei was not compromised in the absence of Crt1. Phylogenetic analysis revealed that orthologs of Crt1 exist in the genomes of many filamentous ascomycete fungi capable of degrading cellulose. These data thus shed new light on the mechanism by which T. reesei senses and transmits the cellulose signal and offers potential strategies for strain improvement. PMID:24085297

  2. Characterization of SiaA, a streptococcal heme-binding protein associated with a heme ABC transport system.

    PubMed

    Sook, Brian R; Block, Darci R; Sumithran, Suganya; Montañez, Griselle E; Rodgers, Kenton R; Dawson, John H; Eichenbaum, Zehava; Dixon, Dabney W

    2008-02-26

    Many pathogenic bacteria require heme and obtain it from their environment. Heme transverses the cytoplasmic membrane via an ATP binding cassette (ABC) pathway. Although a number of heme ABC transport systems have been described in pathogenic bacteria, there is as yet little biophysical characterization of the proteins in these systems. The sia (hts) gene cluster encodes a heme ABC transporter in the Gram positive Streptococcus pyogenes. The lipoprotein-anchored heme binding protein (HBP) of this transporter is SiaA (HtsA). In the current study, resonance Raman (rR), magnetic circular dichroism (MCD), and nuclear magnetic resonance (NMR) spectroscopies were used to determine the coordination state and spin state of both the ferric and ferrous forms of this protein. Identifiers from these techniques suggest that the heme is six-coordinate and low-spin in both oxidation states of the protein, with methionine and histidine as axial ligands. SiaA has a pKa of 9.7 +/- 0.1, attributed to deprotonation of the axial histidine. Guanidinium titration studies show that the ferric state is less stable than the ferrous state, with DeltaG(H2O) values for the oxidized and reduced proteins of 7.3 +/- 0.8 and 16.0 +/- 3.6 kcal mol-1, respectively. The reductive and oxidative midpoint potentials determined via spectroelectrochemistry are 83 +/- 3 and 64 +/- 3 mV, respectively; the irreversibility of heme reduction suggests that redox cycling of the heme is coupled to a kinetically sluggish change in structure or conformation. The biophysical characterization described herein will significantly advance our understanding of structure-function relationships in HBP.

  3. ABC transporters P-gp and Bcrp do not limit the brain uptake of the novel antipsychotic and anticonvulsant drug cannabidiol in mice

    PubMed Central

    Brzozowska, Natalia; Li, Kong M.; Wang, Xiao Suo; Booth, Jessica; Stuart, Jordyn; McGregor, Iain S.

    2016-01-01

    Cannabidiol (CBD) is currently being investigated as a novel therapeutic for the treatment of CNS disorders like schizophrenia and epilepsy. ABC transporters such as P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp) mediate pharmacoresistance in these disorders. P-gp and Bcrp are expressed at the blood brain barrier (BBB) and reduce the brain uptake of substrate drugs including various antipsychotics and anticonvulsants. It is therefore important to assess whether CBD is prone to treatment resistance mediated by P-gp and Bcrp. Moreover, it has become common practice in the drug development of CNS agents to screen against ABC transporters to help isolate lead compounds with optimal pharmacokinetic properties. The current study aimed to assess whether P-gp and Bcrp impacts the brain transport of CBD by comparing CBD tissue concentrations in wild-type (WT) mice versus mice devoid of ABC transporter genes. P-gp knockout (Abcb1a/b−∕−), Bcrp knockout (Abcg2−∕−), combined P-gp/Bcrp knockout (Abcb1a/b−∕−Abcg2−∕−) and WT mice were injected with CBD, before brain and plasma samples were collected at various time-points. CBD results were compared with the positive control risperidone and 9-hydroxy risperidone, antipsychotic drugs that are established ABC transporter substrates. Brain and plasma concentrations of CBD were not greater in P-gp, Bcrp or P-gp/Bcrp knockout mice than WT mice. In comparison, the brain/plasma concentration ratios of risperidone and 9-hydroxy risperidone were profoundly higher in P-gp knockout mice than WT mice. These results suggest that CBD is not a substrate of P-gp or Bcrp and may be free from the complication of reduced brain uptake by these transporters. Such findings provide favorable evidence for the therapeutic development of CBD in the treatment of various CNS disorders. PMID:27257556

  4. ABC transporters P-gp and Bcrp do not limit the brain uptake of the novel antipsychotic and anticonvulsant drug cannabidiol in mice.

    PubMed

    Brzozowska, Natalia; Li, Kong M; Wang, Xiao Suo; Booth, Jessica; Stuart, Jordyn; McGregor, Iain S; Arnold, Jonathon C

    2016-01-01

    Cannabidiol (CBD) is currently being investigated as a novel therapeutic for the treatment of CNS disorders like schizophrenia and epilepsy. ABC transporters such as P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp) mediate pharmacoresistance in these disorders. P-gp and Bcrp are expressed at the blood brain barrier (BBB) and reduce the brain uptake of substrate drugs including various antipsychotics and anticonvulsants. It is therefore important to assess whether CBD is prone to treatment resistance mediated by P-gp and Bcrp. Moreover, it has become common practice in the drug development of CNS agents to screen against ABC transporters to help isolate lead compounds with optimal pharmacokinetic properties. The current study aimed to assess whether P-gp and Bcrp impacts the brain transport of CBD by comparing CBD tissue concentrations in wild-type (WT) mice versus mice devoid of ABC transporter genes. P-gp knockout (Abcb1a/b (-∕-)), Bcrp knockout (Abcg2 (-∕-)), combined P-gp/Bcrp knockout (Abcb1a/b (-∕-) Abcg2 (-∕-)) and WT mice were injected with CBD, before brain and plasma samples were collected at various time-points. CBD results were compared with the positive control risperidone and 9-hydroxy risperidone, antipsychotic drugs that are established ABC transporter substrates. Brain and plasma concentrations of CBD were not greater in P-gp, Bcrp or P-gp/Bcrp knockout mice than WT mice. In comparison, the brain/plasma concentration ratios of risperidone and 9-hydroxy risperidone were profoundly higher in P-gp knockout mice than WT mice. These results suggest that CBD is not a substrate of P-gp or Bcrp and may be free from the complication of reduced brain uptake by these transporters. Such findings provide favorable evidence for the therapeutic development of CBD in the treatment of various CNS disorders.

  5. A new ABC half-transporter in Leishmania major is involved in resistance to antimony.

    PubMed

    Manzano, J I; García-Hernández, R; Castanys, S; Gamarro, F

    2013-08-01

    The characterization of ABCI4, a new intracellular ATP-binding cassette (ABC) half-transporter in Leishmania major, is described. We show that ABCI4 is involved in heavy metal export, thereby conferring resistance to Pentostam, to Sb(III), and to As(III) and Cd(II). Parasites overexpressing ABCI4 showed a lower mitochondrial toxic effect of antimony by decreasing reactive oxygen species production and maintained higher values of both the mitochondrial electrochemical potential and total ATP levels with respect to controls. The ABCI4 half-transporter forms homodimers as determined by a coimmunoprecipitation assay. A combination of subcellular localization studies under a confocal microscope and a surface biotinylation assay using parasites expressing green fluorescent protein- and FLAG-tagged ABCI4 suggests that the transporter presents a dual localization in both mitochondria and the plasma membrane. Parasites overexpressing ABCI4 present an increased replication in mouse peritoneal macrophages. We have determined that porphyrins are substrates for ABCI4. Consequently, the overexpression of ABCI4 confers resistance to some toxic porphyrins, such as zinc-protoporphyrin, due to the lower accumulation resulting from a significant efflux, as determined using the fluorescent zinc-mesoporphyrin, a validated heme analog. In addition, ABCI4 has a significant ability to efflux thiol after Sb(III) incubation, thus meaning that ABCI4 could be considered to be a potential thiol-X-pump that is able to recognize metal-conjugated thiols. In summary, we have shown that this new ABC transporter is involved in drug sensitivity to antimony and other compounds by efflux as conjugated thiol complexes.

  6. Pharmacogenomics of the human ABC transporter ABCG2: from functional evaluation to drug molecular design

    NASA Astrophysics Data System (ADS)

    Ishikawa, Toshihisa; Tamura, Ai; Saito, Hikaru; Wakabayashi, Kanako; Nakagawa, Hiroshi

    2005-10-01

    In the post-genome-sequencing era, emerging genomic technologies are shifting the paradigm for drug discovery and development. Nevertheless, drug discovery and development still remain high-risk and high-stakes ventures with long and costly timelines. Indeed, the attrition of drug candidates in preclinical and development stages is a major problem in drug design. For at least 30% of the candidates, this attrition is due to poor pharmacokinetics and toxicity. Thus, pharmaceutical companies have begun to seriously re-evaluate their current strategies of drug discovery and development. In that light, we propose that a transport mechanism-based design might help to create new, pharmacokinetically advantageous drugs, and as such should be considered an important component of drug design strategy. Performing enzyme- and/or cell-based drug transporter, interaction tests may greatly facilitate drug development and allow the prediction of drug-drug interactions. We recently developed methods for high-speed functional screening and quantitative structure-activity relationship analysis to study the substrate specificity of ABC transporters and to evaluate the effect of genetic polymorphisms on their function. These methods would provide a practical tool to screen synthetic and natural compounds, and these data can be applied to the molecular design of new drugs. In this review article, we present an overview on the genetic polymorphisms of human ABC transporter ABCG2 and new camptothecin analogues that can circumvent AGCG2-associated multidrug resistance of cancer.

  7. A Neisseria meningitidis fbpABC mutant is incapable of using nonheme iron for growth.

    PubMed

    Khun, H H; Kirby, S D; Lee, B C

    1998-05-01

    The neisserial fbpABC locus has been proposed to act as an iron-specific ABC transporter system. To confirm this assigned function, we constructed an fbpABC mutant in Neisseria meningitidis by insertional inactivation of fbpABC with a selectable antibiotic marker. The mutant was unable to use iron supplied from human transferrin, human lactoferrin, or iron chelates. However, the use of iron from heme and human hemoglobin was unimpaired. These results support the obligatory participation of fbpABC in neisserial periplasmic iron transport and do not indicate a role for this genetic locus in the heme iron pathway.

  8. A Neisseria meningitidis fbpABC Mutant Is Incapable of Using Nonheme Iron for Growth

    PubMed Central

    Khun, Heng H.; Kirby, Shane D.; Lee, B. Craig

    1998-01-01

    The neisserial fbpABC locus has been proposed to act as an iron-specific ABC transporter system. To confirm this assigned function, we constructed an fbpABC mutant in Neisseria meningitidis by insertional inactivation of fbpABC with a selectable antibiotic marker. The mutant was unable to use iron supplied from human transferrin, human lactoferrin, or iron chelates. However, the use of iron from heme and human hemoglobin was unimpaired. These results support the obligatory participation of fbpABC in neisserial periplasmic iron transport and do not indicate a role for this genetic locus in the heme iron pathway. PMID:9573125

  9. 75 FR 49549 - ABC & D Recycling, Inc.-Lease and Operation Exemption-a Line of Railroad in Ware, MA

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-13

    ... DEPARTMENT OF TRANSPORTATION Surface Transportation Board [Docket No. FD 35397] ABC & D Recycling, Inc.--Lease and Operation Exemption--a Line of Railroad in Ware, MA ABC & D Recycling, Inc. (ABC & D..., ABC & D Recycling, Inc.--Lease and Operation Exemption--a Line of Railroad in Ware, Massachusetts (STB...

  10. Independent activity of the homologous small regulatory RNAs AbcR1 and AbcR2 in the legume symbiont Sinorhizobium meliloti.

    PubMed

    Torres-Quesada, Omar; Millán, Vicenta; Nisa-Martínez, Rafael; Bardou, Florian; Crespi, Martín; Toro, Nicolás; Jiménez-Zurdo, José I

    2013-01-01

    The legume symbiont Sinorhizobium meliloti expresses a plethora of small noncoding RNAs (sRNAs) whose function is mostly unknown. Here, we have functionally characterized two tandemly encoded S. meliloti Rm1021 sRNAs that are similar in sequence and structure. Homologous sRNAs (designated AbcR1 and AbcR2) have been shown to regulate several ABC transporters in the related α-proteobacteria Agrobacterium tumefaciens and Brucella abortus. In Rm1021, AbcR1 and AbcR2 exhibit divergent unlinked regulation and are stabilized by the RNA chaperone Hfq. AbcR1 is transcribed in actively dividing bacteria, either in culture, rhizosphere or within the invasion zone of mature alfalfa nodules. Conversely, AbcR2 expression is induced upon entry into stationary phase and under abiotic stress. Only deletion of AbcR1 resulted into a discrete growth delay in rich medium, but both are dispensable for symbiosis. Periplasmic proteome profiling revealed down-regulation of the branched-chain amino acid binding protein LivK by AbcR1, but not by AbcR2. A double-plasmid reporter assay confirmed the predicted specific targeting of the 5'-untranslated region of the livK mRNA by AbcR1 in vivo. Our findings provide evidences of independent regulatory functions of these sRNAs, probably to fine-tune nutrient uptake in free-living and undifferentiated symbiotic rhizobia.

  11. Independent Activity of the Homologous Small Regulatory RNAs AbcR1 and AbcR2 in the Legume Symbiont Sinorhizobium meliloti

    PubMed Central

    Torres-Quesada, Omar; Millán, Vicenta; Nisa-Martínez, Rafael; Bardou, Florian; Crespi, Martín; Toro, Nicolás; Jiménez-Zurdo, José I.

    2013-01-01

    The legume symbiont Sinorhizobium meliloti expresses a plethora of small noncoding RNAs (sRNAs) whose function is mostly unknown. Here, we have functionally characterized two tandemly encoded S. meliloti Rm1021 sRNAs that are similar in sequence and structure. Homologous sRNAs (designated AbcR1 and AbcR2) have been shown to regulate several ABC transporters in the related α-proteobacteria Agrobacterium tumefaciens and Brucella abortus. In Rm1021, AbcR1 and AbcR2 exhibit divergent unlinked regulation and are stabilized by the RNA chaperone Hfq. AbcR1 is transcribed in actively dividing bacteria, either in culture, rhizosphere or within the invasion zone of mature alfalfa nodules. Conversely, AbcR2 expression is induced upon entry into stationary phase and under abiotic stress. Only deletion of AbcR1 resulted into a discrete growth delay in rich medium, but both are dispensable for symbiosis. Periplasmic proteome profiling revealed down-regulation of the branched-chain amino acid binding protein LivK by AbcR1, but not by AbcR2. A double-plasmid reporter assay confirmed the predicted specific targeting of the 5′-untranslated region of the livK mRNA by AbcR1 in vivo. Our findings provide evidences of independent regulatory functions of these sRNAs, probably to fine-tune nutrient uptake in free-living and undifferentiated symbiotic rhizobia. PMID:23869210

  12. Convergent Loss of ABC Transporter Genes From Clostridioides difficile Genomes Is Associated With Impaired Tyrosine Uptake and p-Cresol Production.

    PubMed

    Steglich, Matthias; Hofmann, Julia D; Helmecke, Julia; Sikorski, Johannes; Spröer, Cathrin; Riedel, Thomas; Bunk, Boyke; Overmann, Jörg; Neumann-Schaal, Meina; Nübel, Ulrich

    2018-01-01

    We report the frequent, convergent loss of two genes encoding the substrate-binding protein and the ATP-binding protein of an ATP-binding cassette (ABC) transporter from the genomes of unrelated Clostridioides difficile strains. This specific genomic deletion was strongly associated with the reduced uptake of tyrosine and phenylalanine and production of derived Stickland fermentation products, including p -cresol, suggesting that the affected ABC transporter had been responsible for the import of aromatic amino acids. In contrast, the transporter gene loss did not measurably affect bacterial growth or production of enterotoxins. Phylogenomic analysis of publically available genome sequences indicated that this transporter gene deletion had occurred multiple times in diverse clonal lineages of C. difficile , with a particularly high prevalence in ribotype 027 isolates, where 48 of 195 genomes (25%) were affected. The transporter gene deletion likely was facilitated by the repetitive structure of its genomic location. While at least some of the observed transporter gene deletions are likely to have occurred during the natural life cycle of C. difficile , we also provide evidence for the emergence of this mutation during long-term laboratory cultivation of reference strain R20291.

  13. Convergent Loss of ABC Transporter Genes From Clostridioides difficile Genomes Is Associated With Impaired Tyrosine Uptake and p-Cresol Production

    PubMed Central

    Steglich, Matthias; Hofmann, Julia D.; Helmecke, Julia; Sikorski, Johannes; Spröer, Cathrin; Riedel, Thomas; Bunk, Boyke; Overmann, Jörg; Neumann-Schaal, Meina; Nübel, Ulrich

    2018-01-01

    We report the frequent, convergent loss of two genes encoding the substrate-binding protein and the ATP-binding protein of an ATP-binding cassette (ABC) transporter from the genomes of unrelated Clostridioides difficile strains. This specific genomic deletion was strongly associated with the reduced uptake of tyrosine and phenylalanine and production of derived Stickland fermentation products, including p-cresol, suggesting that the affected ABC transporter had been responsible for the import of aromatic amino acids. In contrast, the transporter gene loss did not measurably affect bacterial growth or production of enterotoxins. Phylogenomic analysis of publically available genome sequences indicated that this transporter gene deletion had occurred multiple times in diverse clonal lineages of C. difficile, with a particularly high prevalence in ribotype 027 isolates, where 48 of 195 genomes (25%) were affected. The transporter gene deletion likely was facilitated by the repetitive structure of its genomic location. While at least some of the observed transporter gene deletions are likely to have occurred during the natural life cycle of C. difficile, we also provide evidence for the emergence of this mutation during long-term laboratory cultivation of reference strain R20291. PMID:29867812

  14. Investigating the dynamic nature of the ABC transporters: ABCB1 and MsbA as examples for the potential synergies of MD theory and EPR applications.

    PubMed

    Stockner, Thomas; Mullen, Anna; MacMillan, Fraser

    2015-10-01

    ABC transporters are primary active transporters found in all kingdoms of life. Human multidrug resistance transporter ABCB1, or P-glycoprotein, has an extremely broad substrate spectrum and confers resistance against chemotherapy drug treatment in cancer cells. The bacterial ABC transporter MsbA is a lipid A flippase and a homolog to the human ABCB1 transporter, with which it partially shares its substrate spectrum. Crystal structures of MsbA and ABCB1 have been solved in multiple conformations, providing a glimpse into the possible conformational changes the transporter could be going through during the transport cycle. Crystal structures are inherently static, while a dynamic picture of the transporter in motion is needed for a complete understanding of transporter function. Molecular dynamics (MD) simulations and electron paramagnetic resonance (EPR) spectroscopy can provide structural information on ABC transporters, but the strength of these two methods lies in the potential to characterise the dynamic regime of these transporters. Information from the two methods is quite complementary. MD simulations provide an all atom dynamic picture of the time evolution of the molecular system, though with a narrow time window. EPR spectroscopy can probe structural, environmental and dynamic properties of the transporter in several time regimes, but only through the attachment sites of an exogenous spin label. In this review the synergistic effects that can be achieved by combining the two methods are highlighted, and a brief methodological background is also presented. © 2015 Authors; published by Portland Press Limited.

  15. Polymorphisms in ABC Transporter Genes and Concentrations of Mercury in Newborns – Evidence from Two Mediterranean Birth Cohorts

    PubMed Central

    Llop, Sabrina; Engström, Karin; Ballester, Ferran; Franforte, Elisa; Alhamdow, Ayman; Pisa, Federica; Tratnik, Janja Snoj; Mazej, Datja; Murcia, Mario; Rebagliato, Marisa; Bustamante, Mariona; Sunyer, Jordi; Sofianou-Katsoulis, Αikaterini; Prasouli, Alexia; Antonopoulou, Eleni; Antoniadou, Ioanna; Nakou, Sheena; Barbone, Fabio; Horvat, Milena; Broberg, Karin

    2014-01-01

    Background The genetic background may influence methylmercury (MeHg) metabolism and neurotoxicity. ATP binding cassette (ABC) transporters actively transport various xenobiotics across biological membranes. Objective To investigate the role of ABC polymorphisms as modifiers of prenatal exposure to MeHg. Methods The study population consisted of participants (n = 1651) in two birth cohorts, one in Italy and Greece (PHIME) and the other in Spain (INMA). Women were recruited during pregnancy in Italy and Spain, and during the perinatal period in Greece. Total mercury concentrations were measured in cord blood samples by atomic absorption spectrometry. Maternal fish intake during pregnancy was determined from questionnaires. Polymorphisms (n = 5) in the ABC genes ABCA1, ABCB1, ABCC1 and ABCC2 were analysed in both cohorts. Results ABCB1 rs2032582, ABCC1 rs11075290, and ABCC2 rs2273697 modified the associations between maternal fish intake and cord blood mercury concentrations. The overall interaction coefficient between rs2032582 and log2-transformed fish intake was negative for carriers of GT (β = −0.29, 95%CI −0.47, −0.12) and TT (β = −0.49, 95%CI −0.71, −0.26) versus GG, meaning that for a doubling in fish intake of the mothers, children with the rs2032582 GG genotype accumulated 35% more mercury than children with TT. For rs11075290, the interaction coefficient was negative for carriers of TC (β = −0.12, 95%CI −0.33, 0.09), and TT (β = −0.28, 95%CI −0.51, −0.06) versus CC. For rs2273697, the interaction coefficient was positive when combining GA+AA (β = 0.16, 95%CI 0.01, 0.32) versus GG. Conclusion The ABC transporters appear to play a role in accumulation of MeHg during early development. PMID:24831289

  16. The yeast plasma membrane ATP binding cassette (ABC) transporter Aus1: purification, characterization, and the effect of lipids on its activity.

    PubMed

    Marek, Magdalena; Milles, Sigrid; Schreiber, Gabriele; Daleke, David L; Dittmar, Gunnar; Herrmann, Andreas; Müller, Peter; Pomorski, Thomas Günther

    2011-06-17

    The ATP binding cassette (ABC) transporter Aus1 is expressed under anaerobic growth conditions at the plasma membrane of the yeast Saccharomyces cerevisiae and is required for sterol uptake. These observations suggest that Aus1 promotes the translocation of sterols across membranes, but the precise transport mechanism has yet to be identified. In this study, an extraction and purification procedure was developed to characterize the Aus1 transporter. The detergent-solubilized protein was able to bind and hydrolyze ATP. Mutagenesis of the conserved lysine to methionine in the Walker A motif abolished ATP hydrolysis. Likewise, ATP hydrolysis was inhibited by classical inhibitors of ABC transporters. Upon reconstitution into proteoliposomes, the ATPase activity of Aus1 was specifically stimulated by phosphatidylserine (PS) in a stereoselective manner. We also found that Aus1-dependent sterol uptake, but not Aus1 expression and trafficking to the plasma membrane, was affected by changes in cellular PS levels. These results suggest a direct interaction between Aus1 and PS that is critical for the activity of the transporter.

  17. The phytoestrogen genistein enhances multidrug resistance in breast cancer cell lines by translational regulation of ABC transporters.

    PubMed

    Rigalli, Juan Pablo; Tocchetti, Guillermo Nicolás; Arana, Maite Rocío; Villanueva, Silvina Stella Maris; Catania, Viviana Alicia; Theile, Dirk; Ruiz, María Laura; Weiss, Johanna

    2016-06-28

    Breast cancer is the most frequent malignancy in women. Multidrug resistance due to overexpression of ABC drug transporters is a common cause of chemotherapy failure and disease recurrence. Genistein (GNT) is a phytoestrogen present in soybeans and hormone supplements. We investigated the effect of GNT on the expression and function of ABC transporters in MCF-7 and MDA-MB-231 breast cancer cell lines. Results demonstrated an induction at the protein level of ABCC1 and ABCG2 and of ABCC1 in MCF-7 and MDA-MB-231, respectively. MCF-7 cells showed a concomitant increase in doxorubicin and mitoxantrone efflux and resistance, dependent on ABCG2 activity. ABCC1 induction by GNT in MDA-MB-231 cells modified neither drug efflux nor chemoresistance due to simultaneous acute inhibition of the transporter activity by GNT. All inductions took place at the translational level, as no increment in mRNA was observed and protein increase was prevented by cycloheximide. miR-181a, already demonstrated to inhibit ABCG2 translation, was down-regulated by GNT, explaining translational induction. Effects were independent of classical estrogen receptors. Results suggest potential nutrient-drug interactions that could threaten chemotherapy efficacy, especially in ABCG2-expressing tumors treated with substrates of this transporter. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  18. Multidrug efflux transporter, AcrB--the pumping mechanism.

    PubMed

    Murakami, Satoshi

    2008-08-01

    Resistance nodulation cell division (RND) transporters are one of the main causes of the bacterial multidrug resistance. They pump a wide range of antibiotics out of the cell by proton motive force. AcrB is the major RND transporter in Escherichia coli. Recently, the crystal structures of AcrB have been determined by different space groups. All these structures are consistent with asymmetric trimer. Each monomer has different conformation corresponding to one of the three functional states of the transport cycle. Transporting hydrophobic drug was bound in the periplasmic domain on one of the three monomers. The transport pathway with alternating access mechanism is located at the hydrophilic domain protruded into the periplasmic space while this mechanism of other transporter families like ATP binding cassette (ABC) and major facilitator superfamily (MFS) transporter is located in the membrane-embedded region. For the RND, protonation might also take place asymmetrically at the functionally important charged residues in the transmembrane (TM) region. The structures indicate that drugs are transported by a three-step functional rotation in which substrates undergo ordered binding change.

  19. A Major Facilitator Superfamily Transporter Plays a Dual Role in Polar Auxin Transport and Drought Stress Tolerance in Arabidopsis[W

    PubMed Central

    Remy, Estelle; Cabrito, Tânia R.; Baster, Pawel; Batista, Rita A.; Teixeira, Miguel C.; Friml, Jiri; Sá-Correia, Isabel; Duque, Paula

    2013-01-01

    Many key aspects of plant development are regulated by the polarized transport of the phytohormone auxin. Cellular auxin efflux, the rate-limiting step in this process, has been shown to rely on the coordinated action of PIN-formed (PIN) and B-type ATP binding cassette (ABCB) carriers. Here, we report that polar auxin transport in the Arabidopsis thaliana root also requires the action of a Major Facilitator Superfamily (MFS) transporter, Zinc-Induced Facilitator-Like 1 (ZIFL1). Sequencing, promoter-reporter, and fluorescent protein fusion experiments indicate that the full-length ZIFL1.1 protein and a truncated splice isoform, ZIFL1.3, localize to the tonoplast of root cells and the plasma membrane of leaf stomatal guard cells, respectively. Using reverse genetics, we show that the ZIFL1.1 transporter regulates various root auxin-related processes, while the ZIFL1.3 isoform mediates drought tolerance by regulating stomatal closure. Auxin transport and immunolocalization assays demonstrate that ZIFL1.1 indirectly modulates cellular auxin efflux during shootward auxin transport at the root tip, likely by regulating plasma membrane PIN2 abundance. Finally, heterologous expression in yeast revealed that ZIFL1.1 and ZIFL1.3 share H+-coupled K+ transport activity. Thus, by determining the subcellular and tissue distribution of two isoforms, alternative splicing dictates a dual function for the ZIFL1 transporter. We propose that this MFS carrier regulates stomatal movements and polar auxin transport by modulating potassium and proton fluxes in Arabidopsis cells. PMID:23524662

  20. Phosphorylation is required for the pathogen defense function of the Arabidopsis PEN3 ABC transporter.

    PubMed

    Underwood, William; Somerville, Shauna C

    2017-10-03

    The Arabidopsis PEN3 ABC transporter accumulates at sites of pathogen detection, where it is involved in defense against a number of pathogens. Perception of PAMPs by pattern recognition receptors initiates recruitment of PEN3 and also leads to PEN3 phosphorylation at multiple amino acid residues. Whether PAMP-induced phosphorylation of PEN3 is important for its defense function or focal recruitment has not been addressed. In this study, we evaluated the role of PEN3 phosphorylation in modulating the localization and defense function of the transporter. We report that PEN3 phosphorylation is critical for its function in defense, but dispensable for recruitment to powdery mildew penetration sites. These results indicate that PAMP-induced phosphorylation is likely to regulate the transport activity of PEN3.

  1. Classification of a Haemophilus influenzae ABC Transporter HI1470/71 through Its Cognate Molybdate Periplasmic Binding Protein, MolA

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tirado-Lee, Leidamarie; Lee, Allen; Rees, Douglas C.

    2014-10-02

    molA (HI1472) from H. influenzae encodes a periplasmic binding protein (PBP) that delivers substrate to the ABC transporter MolB{sub 2}C{sub 2} (formerly HI1470/71). The structures of MolA with molybdate and tungstate in the binding pocket were solved to 1.6 and 1.7 {angstrom} resolution, respectively. The MolA-binding protein binds molybdate and tungstate, but not other oxyanions such as sulfate and phosphate, making it the first class III molybdate-binding protein structurally solved. The {approx}100 {mu}M binding affinity for tungstate and molybdate is significantly lower than observed for the class II ModA molybdate-binding proteins that have nanomolar to low micromolar affinity for molybdate.more » The presence of two molybdate loci in H. influenzae suggests multiple transport systems for one substrate, with molABC constituting a low-affinity molybdate locus.« less

  2. Vibrio cholerae NspS, a homologue of ABC-type periplasmic solute binding proteins, facilitates transduction of polyamine signals independent of their transport

    PubMed Central

    Cockerell, Steven R.; Rutkovsky, Alex C.; Zayner, Josiah P.; Cooper, Rebecca E.; Porter, Lindsay R.; Pendergraft, Sam S.; Parker, Zach M.; McGinnis, Marcus W.

    2014-01-01

    The polyamines norspermidine and spermidine are among the environmental signals that regulate Vibrio cholerae biofilm formation. The effects of these polyamines are mediated by NspS, a member of the bacterial periplasmic solute binding protein superfamily. Almost all members of this superfamily characterized to date are components of ATP-binding cassette-type transporters involved in nutrient uptake. Consequently, in the current annotation of the V. cholerae genome, NspS has been assigned a function in transport. The objective of this study was to further characterize NspS and investigate its potential role in transport. Our results support a role for NspS in signal transduction in response to norspermidine and spermidine, but not their transport. In addition, we provide evidence that these polyamine signals are processed by c-di-GMP signalling networks in the cell. Furthermore, we present comparative genomics analyses which reveal the presence of NspS-like proteins in a variety of bacteria, suggesting that periplasmic ligand binding proteins may be widely utilized for sensory transduction. PMID:24530989

  3. The TULIP superfamily of eukaryotic lipid-binding proteins as a mediator of lipid sensing and transport.

    PubMed

    Alva, Vikram; Lupas, Andrei N

    2016-08-01

    The tubular lipid-binding (TULIP) superfamily has emerged in recent years as a major mediator of lipid sensing and transport in eukaryotes. It currently encompasses three protein families, SMP-like, BPI-like, and Takeout-like, which share a common fold. This fold consists of a long helix wrapped in a highly curved anti-parallel β-sheet, enclosing a central, lipophilic cavity. The SMP-like proteins, which include subunits of the ERMES complex and the extended synaptotagmins (E-Syts), appear to be mainly located at membrane contacts sites (MCSs) between organelles, mediating inter-organelle lipid exchange. The BPI-like proteins, which include the bactericidal/permeability-increasing protein (BPI), the LPS (lipopolysaccharide)-binding protein (LBP), the cholesteryl ester transfer protein (CETP), and the phospholipid transfer protein (PLTP), are either involved in innate immunity against bacteria through their ability to sense lipopolysaccharides, as is the case for BPI and LBP, or in lipid exchange between lipoprotein particles, as is the case for CETP and PLTP. The Takeout-like proteins, which are comprised of insect juvenile hormone-binding proteins and arthropod allergens, transport, where known, lipid hormones to target tissues during insect development. In all cases, the activity of these proteins is underpinned by their ability to bind large, hydrophobic ligands in their central cavity and segregate them away from the aqueous environment. Furthermore, where they are involved in lipid exchange, recent structural studies have highlighted their ability to establish lipophilic, tubular channels, either between organelles in the case of SMP domains or between lipoprotein particles in the case of CETP. Here, we review the current knowledge on the structure, versatile functions, and evolution of the TULIP superfamily. We propose a deep evolutionary split in this superfamily, predating the Last Eukaryotic Common Ancestor, between the SMP-like proteins, which act on

  4. Human and Rat ABC Transporter Efflux of Bisphenol A and Bisphenol A Glucuronide: Interspecies Comparison and Implications for Pharmacokinetic Assessment

    EPA Science Inventory

    Significant interspecies differences exist between human and rodent with respect to absorption, distribution, and excretion of bisphenol A (BPA) and its primary metabolite, BPA-glucuronide (BPA-G). ATP-Binding Cassette (ABC) transporter enzymes play important roles in these physi...

  5. Biochemical characterization of an ABC transporter LptBFGC complex required for the outer membrane sorting of lipopolysaccharides.

    PubMed

    Narita, Shin-ichiro; Tokuda, Hajime

    2009-07-07

    Seven Lpt proteins (A through G) are thought to be involved in lipopolysaccharide transport from the inner to outer membrane of Escherichia coli. LptB belongs to the ATP-binding cassette transporter superfamily. Although the lptB gene lacks neighboring genes encoding membrane subunits, bioinformatic analyses recently indicated that two distantly located consecutive genes, lptF and lptG, could encode membrane subunits. To examine this possibility, LptB was expressed with LptF and LptG. We report here that both LptF and LptG formed a complex with LptB. Furthermore, an inner membrane protein, LptC, which had been implicated in lipopolysaccharide transport, was also included in this complex.

  6. An ATP-driven efflux pump is a novel pathogenicity factor in rice blast disease.

    PubMed Central

    Urban, M; Bhargava, T; Hamer, J E

    1999-01-01

    Cells tolerate exposure to cytotoxic compounds through the action of ATP-driven efflux pumps belonging to the ATP-binding cassette (ABC) superfamily of membrane transporters. Phytopathogenic fungi encounter toxic environments during plant invasion as a result of the plant defense response. Here we demonstrate the requirement for an ABC transporter during host infection by the fungal plant pathogen Magnaporthe grisea. The ABC1 gene was identified in an insertional mutagenesis screen for pathogenicity mutants. The ABC1 insertional mutant and a gene-replacement mutant arrest growth and die shortly after penetrating either rice or barley epidermal cells. The ABC1-encoded protein is similar to yeast ABC transporters implicated in multidrug resistance, and ABC1 gene transcripts are inducible by toxic drugs and a rice phytoalexin. However, abc1 mutants are not hypersensitive to antifungal compounds. The non-pathogenic, insertional mutation in ABC1 occurs in the promoter region and dramatically reduces transcript induction by metabolic poisons. These data strongly suggest that M.grisea requires the up-regulation of specific ABC transporters for pathogenesis; most likely to protect itself against plant defense mechanisms. PMID:9927411

  7. 75 FR 11991 - ABC & D Recycling, Inc.-Lease and Operation Exemption-a Line of Railroad in Ware, MA

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-03-12

    ... DEPARTMENT OF TRANSPORTATION Surface Transportation Board [STB Finance Docket No. 35356] ABC & D Recycling, Inc.--Lease and Operation Exemption--a Line of Railroad in Ware, MA ABC & D Recycling, Inc. (ABC & D), a noncarrier, has filed a verified notice of exemption under 49 CFR 1150.31 to lease from O...

  8. The Role of the Photoreceptor ABC Transporter ABCA4 in Lipid Transport and Stargardt Macular Degeneration

    PubMed Central

    Molday, Robert S.; Zhong, Ming; Quazi, Faraz

    2009-01-01

    ABCA4 is a member of the ABCA subfamily of ATP binding cassette (ABC) transporters that is expressed in rod and cone photoreceptors of the vertebrate retina. ABCA4, also known as the Rim protein and ABCR, is a large 2273 amino acid glycoprotein organized as two tandem halves, each containing a single membrane spanning segment followed sequentially by a large exocytoplasmic domain, a multispanning membrane domain and a nucleotide binding domain. Over 500 mutations in the gene encoding ABCA4 are associated with a spectrum of related autosomal recessive retinal degenerative diseases including Stargardt macular degeneration, cone-rod dystrophy and a subset of retinitis pigmentosa. Biochemical studies on the purified ABCA4 together with analysis of abca4 knockout mice and patients with Stargardt disease have implicated ABCA4 as a retinylidene-phosphatidylethanolamine transporter that facilitates the removal of potentially reactive retinal derivatives from photoreceptors following photoexcitation. Knowledge of the genetic and molecular basis for ABCA4 related retinal degenerative diseases is being used to develop rationale therapeutic treatments for this set of disorders. PMID:19230850

  9. Identification of Residues in the Lipopolysaccharide ABC Transporter That Coordinate ATPase Activity with Extractor Function.

    PubMed

    Simpson, Brent W; Owens, Tristan W; Orabella, Matthew J; Davis, Rebecca M; May, Janine M; Trauger, Sunia A; Kahne, Daniel; Ruiz, Natividad

    2016-10-18

    The surface of most Gram-negative bacteria is covered with lipopolysaccharide (LPS), creating a permeability barrier against toxic molecules, including many antimicrobials. To assemble LPS on their surface, Gram-negative bacteria must extract newly synthesized LPS from the inner membrane, transport it across the aqueous periplasm, and translocate it across the outer membrane. The LptA to -G proteins assemble into a transenvelope complex that transports LPS from the inner membrane to the cell surface. The Lpt system powers LPS transport from the inner membrane by using a poorly characterized ATP-binding cassette system composed of the ATPase LptB and the transmembrane domains LptFG. Here, we characterize a cluster of residues in the groove region of LptB that is important for controlling LPS transport. We also provide the first functional characterization of LptFG and identify their coupling helices that interact with the LptB groove. Substitutions at conserved residues in these coupling helices compromise both the assembly and function of the LptB 2 FG complex. Defects in LPS transport conferred by alterations in the LptFG coupling helices can be rescued by changing a residue in LptB that is adjacent to functionally important residues in the groove region. This suppression is achieved by increasing the ATPase activity of the LptB 2 FG complex. Taken together, these data identify a specific binding site in LptB for the coupling helices of LptFG that is responsible for coupling of ATP hydrolysis by LptB with LptFG function to achieve LPS extraction. Lipopolysaccharide (LPS) is synthesized at the cytoplasmic membrane of Gram-negative bacteria and transported across several compartments to the cell surface, where it forms a barrier that protects these organisms from antibiotics. The LptB 2 FG proteins form an ATP-binding cassette (ABC) transporter that uses energy from ATP hydrolysis in the cytoplasm to facilitate extraction of LPS from the outer face of the

  10. Inactivation of the Ecs ABC transporter of Staphylococcus aureus attenuates virulence by altering composition and function of bacterial wall.

    PubMed

    Jonsson, Ing-Marie; Juuti, Jarmo T; François, Patrice; AlMajidi, Rana; Pietiäinen, Milla; Girard, Myriam; Lindholm, Catharina; Saller, Manfred J; Driessen, Arnold J M; Kuusela, Pentti; Bokarewa, Maria; Schrenzel, Jacques; Kontinen, Vesa P

    2010-12-02

    Ecs is an ATP-binding cassette (ABC) transporter present in aerobic and facultative anaerobic gram-positive Firmicutes. Inactivation of Bacillus subtilis Ecs causes pleiotropic changes in the bacterial phenotype including inhibition of intramembrane proteolysis. The molecule(s) transported by Ecs is (are) still unknown. In this study we mutated the ecsAB operon in two Staphylococcus aureus strains, Newman and LS-1. Phenotypic and functional characterization of these Ecs deficient mutants revealed a defect in growth, increased autolysis and lysostaphin sensitivity, altered composition of cell wall proteins including the precursor form of staphylokinase and an altered bacterial surface texture. DNA microarray analysis indicated that the Ecs deficiency changed expression of the virulence factor regulator protein Rot accompanied by differential expression of membrane transport proteins, particularly ABC transporters and phosphate-specific transport systems, protein A, adhesins and capsular polysaccharide biosynthesis proteins. Virulence of the ecs mutants was studied in a mouse model of hematogenous S. aureus infection. Mice inoculated with the ecs mutant strains developed markedly milder infections than those inoculated with the wild-type strains and had consequently lower mortality, less weight loss, milder arthritis and decreased persistence of staphylococci in the kidneys. The ecs mutants had higher susceptibility to ribosomal antibiotics and plant alkaloids chelerythrine and sanguinarine. Our results show that Ecs is essential for staphylococcal virulence and antimicrobial resistance probably since the transport function of Ecs is essential for the normal structure and function of the cell wall. Thus targeting Ecs may be a new approach in combating staphylococcal infection.

  11. Transporter-mediated natural product-drug interactions for the treatment of cardiovascular diseases.

    PubMed

    Zha, Weibin

    2018-04-01

    The growing use of natural products in cardiovascular (CV) patients has been greatly raising the concerns about potential natural product-CV drug interactions. Some of these may lead to unexpected cardiovascular adverse effects and it is, therefore, essential to identify or predict potential natural product-CV drug interactions, and to understand the underlying mechanisms. Drug transporters are important determinants for the pharmacokinetics of drugs and alterations of drug transport has been recognized as one of the major causes of natural product-drug interactions. In last two decades, many CV drugs (e.g., angiotensin II receptor blockers, beta-blockers and statins) have been identified to be substrates and inhibitors of the solute carrier (SLC) transporters and the ATP-binding cassette (ABC) transporters, which are two major transporter superfamilies. Meanwhile, in vitro and in vivo studies indicate that a growing number of natural products showed cardioprotective effects (e.g., gingko biloba, danshen and their active ingredients) are also substrates and inhibitors of drug transporters. Thus, to understand transporter-mediated natural product-CV drug interactions is important and some transporter-mediated interactions have already shown to have clinical relevance. In this review, we review the current knowledge on the role of ABC and SLC transporters in CV therapy, as well as transporter modulation by natural products used in CV diseases and their induced natural product-CV drug interactions through alterations of drug transport. We hope our review will aid in a comprehensive summary of transporter-mediated natural product-CV drug interactions and help public and physicians understand these type of interactions. Copyright © 2017. Published by Elsevier B.V.

  12. An alternative physiological role for the EmhABC efflux pump in Pseudomonas fluorescens cLP6a

    PubMed Central

    2011-01-01

    Background Efflux pumps belonging to the resistance-nodulation-division (RND) superfamily in bacteria are involved in antibiotic resistance and solvent tolerance but have an unknown physiological role. EmhABC, a RND-type efflux pump in Pseudomonas fluorescens strain cLP6a, extrudes hydrophobic antibiotics, dyes and polycyclic aromatic hydrocarbons including phenanthrene. The effects of physico-chemical factors such as temperature or antibiotics on the activity and expression of EmhABC were determined in order to deduce its physiological role(s) in strain cLP6a in comparison to the emhB disruptant strain, cLP6a-1. Results Efflux assays conducted with 14C-phenanthrene showed that EmhABC activity is affected by incubation temperature. Increased phenanthrene efflux was measured in cLP6a cells grown at 10°C and decreased efflux was observed at 35°C compared with cells grown at the optimum temperature of 28°C. Membrane fatty acids in cLP6a cells were substantially altered by changes in growth temperature and in the presence of tetracycline. Changed membrane fatty acids and increased membrane permeability were associated with ~30-fold increased expression of emhABC in cLP6a cells grown at 35°C, and with increased extracellular free fatty acids. Growth of P. fluorescens cLP6a at supra-optimal temperature was enhanced by the presence of EmhABC compared to strain cLP6a-1. Conclusions Combined, these observations suggest that the EmhABC efflux pump may be involved in the management of membrane stress effects such as those due to unfavourable incubation temperatures. Efflux of fatty acids replaced as a result of membrane damage or phospholipid turnover may be the primary physiological role of the EmhABC efflux pump in P. fluorescens cLP6a. PMID:22085438

  13. Regulation of the ATPase activity of ABCE1 from Pyrococcus abyssi by Fe-S cluster status and Mg²⁺: implication for ribosomal function.

    PubMed

    Sims, Lynn M; Igarashi, Robert Y

    2012-08-15

    Ribosomal function is dependent on multiple proteins. The ABCE1 ATPase, a unique ABC superfamily member that bears two Fe₄S₄ clusters, is crucial for ribosomal biogenesis and recycling. Here, the ATPase activity of the Pyrococcus abyssi ABCE1 (PabABCE1) was studied using both apo- (without reconstituted Fe-S clusters) and holo- (with full complement of Fe-S clusters reconstituted post-purification) forms, and is shown to be jointly regulated by the status of Fe-S clusters and Mg²⁺. Typically ATPases require Mg²⁺, as is true for PabABCE1, but Mg²⁺ also acts as a negative allosteric effector that modulates ATP affinity of PabABCE1. Physiological [Mg²⁺] inhibits the PabABCE1 ATPase (K(i) of ∼1 μM) for both apo- and holo-PabABCE1. Comparative kinetic analysis of Mg²⁺ inhibition shows differences in degree of allosteric regulation between the apo- and holo-PabABCE1 where the apparent ATP K(m) of apo-PabABCE1 increases >30-fold from ∼30 μM to over 1 mM with M²⁺. This effect would significantly convert the ATPase activity of PabABCE1 from being independent of cellular energy charge (φ) to being dependent on φ with cellular [Mg²⁺]. These findings uncover intricate overlapping effects by both [Mg²⁺] and the status of Fe-S clusters that regulate ABCE1's ATPase activity with implications to ribosomal function. Copyright © 2012 Elsevier Inc. All rights reserved.

  14. The Pseudomonas aeruginosa PA14 ABC Transporter NppA1A2BCD Is Required for Uptake of Peptidyl Nucleoside Antibiotics.

    PubMed

    Pletzer, Daniel; Braun, Yvonne; Dubiley, Svetlana; Lafon, Corinne; Köhler, Thilo; Page, Malcolm G P; Mourez, Michael; Severinov, Konstantin; Weingart, Helge

    2015-07-01

    Analysis of the genome sequence of Pseudomonas aeruginosa PA14 revealed the presence of an operon encoding an ABC-type transporter (NppA1A2BCD) showing homology to the Yej transporter of Escherichia coli. The Yej transporter is involved in the uptake of the peptide-nucleotide antibiotic microcin C, a translation inhibitor that targets the enzyme aspartyl-tRNA synthetase. Furthermore, it was recently shown that the Opp transporter from P. aeruginosa PAO1, which is identical to Npp, is required for uptake of the uridyl peptide antibiotic pacidamycin, which targets the enzyme translocase I (MraY), which is involved in peptidoglycan synthesis. We used several approaches to further explore the substrate specificity of the Npp transporter. Assays of growth in defined minimal medium containing peptides of various lengths and amino acid compositions as sole nitrogen sources, as well as Biolog Phenotype MicroArrays, showed that the Npp transporter is not required for di-, tri-, and oligopeptide uptake. Overexpression of the npp operon increased susceptibility not just to pacidamycin but also to nickel chloride and the peptidyl nucleoside antibiotic blasticidin S. Furthermore, heterologous expression of the npp operon in a yej-deficient mutant of E. coli resulted in increased susceptibility to albomycin, a naturally occurring sideromycin with a peptidyl nucleoside antibiotic. Additionally, heterologous expression showed that microcin C is recognized by the P. aeruginosa Npp system. Overall, these results suggest that the NppA1A2BCD transporter is involved in the uptake of peptidyl nucleoside antibiotics by P. aeruginosa PA14. One of the world's most serious health problems is the rise of antibiotic-resistant bacteria. There is a desperate need to find novel antibiotic therapeutics that either act on new biological targets or are able to bypass known resistance mechanisms. Bacterial ABC transporters play an important role in nutrient uptake from the environment. These uptake

  15. ROLE OF ATP BINDING CASSETTE SUB-FAMILY MEMBER 2 (ABCG2) IN MOUSE EMBRYONIC STEM CELL DEVELOPMENT.

    EPA Science Inventory

    ATP binding cassette sub-family member 2 (ABCG2), is a member of the ABC transporter superfamily and a principal xenobiotic transporter. ABCG2 is also highly expressed in certain stem cell populations where it is thought to be related to stem cell plasticity, although the role o...

  16. Regulation of Expression of abcA and Its Response to Environmental Conditions

    PubMed Central

    Villet, Regis A.; Truong-Bolduc, Que Chi; Wang, Yin; Estabrooks, Zoe; Medeiros, Heidi

    2014-01-01

    The ATP-dependent transporter gene abcA in Staphylococcus aureus confers resistance to hydrophobic β-lactams. In strain ISP794, abcA is regulated by the transcriptional regulators MgrA and NorG and shares a 420-nucleotide intercistronic region with the divergently transcribed pbp4 gene, which encodes the transpeptidase Pbp4. Exposure of exponentially growing cells to iron-limited media, oxidative stress, and acidic pH (5.5) for 0.5 to 2 h had no effect on abcA expression. In contrast, nutrient limitation produced a significant increase in abcA transcripts. We identified three additional regulators (SarA, SarZ, and Rot) that bind to the overlapping promoter region of abcA and pbp4 in strain MW2 and investigated their role in the regulation of abcA expression. Expression of abcA is decreased by 10.0-fold in vivo in a subcutaneous abscess model. In vitro, abcA expression depends on rot and sarZ regulators. Moenomycin A exposure of strain MW2 produced an increase in abcA transcripts. Relative to MW2, the MIC of moenomycin was decreased 8-fold for MW2ΔabcA and increased 10-fold for the MW2 abcA overexpresser, suggesting that moenomycin is a substrate of AbcA. PMID:24509312

  17. Characterization of a major facilitator superfamily transporter in Shiraia bambusicola.

    PubMed

    Deng, Huaxiang; Gao, Ruijie; Liao, Xiangru; Cai, Yujie

    2017-09-01

    Reactive oxygen species (ROS) generated by photo-activated hypocrellin from Shiraia bambusicola are detrimental to cellular macromolecules. However, S. bambusicola can still maintain excellent morphology during continuous hypocrellin production, indicating an extraordinary autoresistance system that protects against the harmful ROS. In this study, a major facilitator superfamily transporter (MFS) was isolated from S. bambusicola and deleted using the clustered regularly interspaced short palindromic repeat sequences (CRISPR)/Cas9 system. The ΔMFS mutant abolished hypocrellin production and was slightly sensitive to 40-μM hypocrellin, while the ΔMFS compliment strain restored hypocrellin production and resistance. Hypocrellin treatment also enhanced the relative expression of MFS in wild-type S. bambusicola. Subsequent pathogenicity assays showed that MFS deletion reduced damage to bamboo leaves. By contrast, restoration of hypocrellin production in the MFS compliment strain generated similar necrotic lesions on bamboo leaves to those observed with the wild-type strain. These results revealed that the identified MFS is involved in efflux of hypocrellin from cells, which reduces the hypocrellin toxicity. Furthermore, hypocrellin contributed to the virulence of S. bambusicola on bamboo leaves. These findings could help to reduce plant loss by disrupting hypocrellin biosynthesis in S. bambusicola, or overexpressing the associated resistance gene in transgenic plants. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  18. Isolation and functional analysis of Thmfs1, the first major facilitator superfamily transporter from the biocontrol fungus Trichoderma harzianum.

    PubMed

    Liu, Mu; Liu, Jun; Wang, Wei Min

    2012-10-01

    A novel major facilitator superfamily (MFS) transporter gene, Thmfs1, was isolated from Trichoderma harzianum (T. harzianum). A Thmfs1 over-expressing mutant displayed enhanced antifungal activity and fungicide tolerance, while the Thmfs1 disruption mutant showed the opposite trend. Trichodermin production in Thmfs1 disruption group (185 mg l(-1)) was decreased by less than 17 % compared to the parental strain, suggesting that Thmfs1 is not mainly responsible for trichodermin secretion. Real-time PCR showed that Thmfs1 transcript level could be induced by a certain range of trichodermin concentrations, while expression of Tri5, encoding a trichodiene synthase, was strongly inhibited under these conditions. To our knowledge, Thmfs1 is the first MFS transporter gene identified in T. harzianum.

  19. Role of nuclear factor-erythroid 2-related factor 2 (Nrf2) in the transcriptional regulation of brain ABC transporters during acute acetaminophen (APAP) intoxication in mice.

    PubMed

    Ghanem, Carolina I; Rudraiah, Swetha; Bataille, Amy M; Vigo, María B; Goedken, Michael J; Manautou, José E

    2015-04-01

    Changes in expression of liver ABC transporters have been described during acute APAP intoxication. However, the effect of APAP on brain ABC transporters is poorly understood. The aim of this study was to evaluate the effect of APAP on brain ABC transporters expression and the role of the oxidative stress sensor Nrf2. Male C57BL/6J mice were administered APAP (400mg/kg) for analysis of brain mRNA and protein expression of Mrp1-6, Bcrp and P-gp. The results show induction of P-gp, Mrp2 and Mrp4 proteins, with no changes in Bcrp, Mrp1 or Mrp5-6. The protein values were accompanied by corresponding changes in mRNA levels. Additionally, brain Nrf2 nuclear translocation and expression of two Nrf2 target genes, quinone oxidoreductase 1 (Nqo1) and Hemoxygenase 1 (Ho-1), was evaluated at 6, 12 and 24h after APAP treatment. Nrf2 nuclear content increased by 58% at 12h after APAP along with significant increments in mRNA and protein expression of Nqo1 and Ho-1. Furthermore, APAP treated Nrf2 knockout mice did not increase mRNA or protein expression of Mrp2 and Mrp4 as observed in wildtypes. In contrast, P-gp induction by APAP was observed in both genotypes. In conclusion, acute APAP intoxication induces protein expression of brain P-gp, Mrp2 and Mrp4. This study also suggests that brain changes in Mrp2 and Mrp4 expression may be due to in situ Nrf2 activation by APAP, while P-gp induction is independent of Nrf2 function. The functional consequences of these changes in brain ABC transporters by APAP deserve further attention. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Emission of volatile organic compounds from petunia flowers is facilitated by an ABC transporter.

    PubMed

    Adebesin, Funmilayo; Widhalm, Joshua R; Boachon, Benoît; Lefèvre, François; Pierman, Baptiste; Lynch, Joseph H; Alam, Iftekhar; Junqueira, Bruna; Benke, Ryan; Ray, Shaunak; Porter, Justin A; Yanagisawa, Makoto; Wetzstein, Hazel Y; Morgan, John A; Boutry, Marc; Schuurink, Robert C; Dudareva, Natalia

    2017-06-30

    Plants synthesize a diversity of volatile molecules that are important for reproduction and defense, serve as practical products for humans, and influence atmospheric chemistry and climate. Despite progress in deciphering plant volatile biosynthesis, their release from the cell has been poorly understood. The default assumption has been that volatiles passively diffuse out of cells. By characterization of a Petunia hybrida adenosine triphosphate-binding cassette (ABC) transporter, PhABCG1, we demonstrate that passage of volatiles across the plasma membrane relies on active transport. PhABCG1 down-regulation by RNA interference results in decreased emission of volatiles, which accumulate to toxic levels in the plasma membrane. This study provides direct proof of a biologically mediated mechanism of volatile emission. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  1. TNF superfamily: costimulation and clinical applications

    PubMed Central

    Vinay, Dass S; Kwon, Byoung S

    2009-01-01

    The molecules concerned with costimulation belong either to the immunoglobulin (Ig) or tumor necrosis factor (TNF) superfamilies. The tumor necrosis superfamily comprises molecules capable of providing both costimulation and cell death. In this review we briefly summarize certain TNF superfamily receptor-ligand pairs that are endowed with costimulatory properties and their importance in health and disease. PMID:19230849

  2. Maltose Uptake by the Novel ABC Transport System MusEFGK2I Causes Increased Expression of ptsG in Corynebacterium glutamicum

    PubMed Central

    Henrich, Alexander; Kuhlmann, Nora; Eck, Alexander W.; Krämer, Reinhard

    2013-01-01

    The Gram-positive Corynebacterium glutamicum efficiently metabolizes maltose by a pathway involving maltodextrin and glucose formation by 4-α-glucanotransferase, glucose phosphorylation by glucose kinases, and maltodextrin degradation via maltodextrin phosphorylase and α-phosphoglucomutase. However, maltose uptake in C. glutamicum has not been investigated. Interestingly, the presence of maltose in the medium causes increased expression of ptsG in C. glutamicum by an unknown mechanism, although the ptsG-encoded glucose-specific EII permease of the phosphotransferase system itself is not required for maltose utilization. We identified the maltose uptake system as an ABC transporter encoded by musK (cg2708; ATPase subunit), musE (cg2705; substrate binding protein), musF (cg2704; permease), and musG (cg2703; permease) by combination of data obtained from characterization of maltose uptake and reanalyses of transcriptome data. Deletion of the mus gene cluster in C. glutamicum Δmus abolished maltose uptake and utilization. Northern blotting and reverse transcription-PCR experiments revealed that musK and musE are transcribed monocistronically, whereas musF and musG are part of an operon together with cg2701 (musI), which encodes a membrane protein of unknown function with no homologies to characterized proteins. Characterization of growth and [14C]maltose uptake in the musI insertion strain C. glutamicum IMcg2701 showed that musI encodes a novel essential component of the maltose ABC transporter of C. glutamicum. Finally, ptsG expression during cultivation on different carbon sources was analyzed in the maltose uptake-deficient strain C. glutamicum Δmus. Indeed, maltose uptake by the novel ABC transport system MusEFGK2I is required for the positive effect of maltose on ptsG expression in C. glutamicum. PMID:23543710

  3. The multidrug ABC transporter BmrC/BmrD of Bacillus subtilis is regulated via a ribosome-mediated transcriptional attenuation mechanism

    PubMed Central

    Reilman, Ewoud; Mars, Ruben A. T.; van Dijl, Jan Maarten; Denham, Emma L.

    2014-01-01

    Expression of particular drug transporters in response to antibiotic pressure is a critical element in the development of bacterial multidrug resistance, and represents a serious concern for human health. To obtain a better understanding of underlying regulatory mechanisms, we have dissected the transcriptional activation of the ATP-binding cassette (ABC) transporter BmrC/BmrD of the Gram-positive model bacterium Bacillus subtilis. By using promoter-GFP fusions and live cell array technology, we demonstrate a temporally controlled transcriptional activation of the bmrCD genes in response to antibiotics that target protein synthesis. Intriguingly, bmrCD expression only occurs during the late-exponential and stationary growth stages, irrespective of the timing of the antibiotic challenge. We show that this is due to tight transcriptional control by the transition state regulator AbrB. Moreover, our results show that the bmrCD genes are co-transcribed with bmrB (yheJ), a small open reading frame immediately upstream of bmrC that harbors three alternative stem-loop structures. These stem-loops are apparently crucial for antibiotic-induced bmrCD transcription. Importantly, the antibiotic-induced bmrCD expression requires translation of bmrB, which implies that BmrB serves as a regulatory leader peptide. Altogether, we demonstrate for the first time that a ribosome-mediated transcriptional attenuation mechanism can control the expression of a multidrug ABC transporter. PMID:25217586

  4. Separating the roles of acropetal and basipetal auxin transport on gravitropism with mutations in two Arabidopsis multidrug resistance-like ABC transporter genes.

    PubMed

    Lewis, Daniel R; Miller, Nathan D; Splitt, Bessie L; Wu, Guosheng; Spalding, Edgar P

    2007-06-01

    Two Arabidopsis thaliana ABC transporter genes linked to auxin transport by various previous results were studied in a reverse-genetic fashion. Mutations in Multidrug Resistance-Like1 (MDR1) reduced acropetal auxin transport in roots by 80% without affecting basipetal transport. Conversely, mutations in MDR4 blocked 50% of basipetal transport without affecting acropetal transport. Developmental and auxin distribution phenotypes associated with these altered auxin flows were studied with a high-resolution morphometric system and confocal microscopy, respectively. Vertically grown mdr1 roots produced positive and negative curvatures threefold greater than the wild type, possibly due to abnormal auxin distribution observed in the elongation zone. However, upon 90 degrees reorientation, mdr1 gravitropism was inseparable from the wild type. Thus, acropetal auxin transport maintains straight growth but contributes surprisingly little to gravitropism. Conversely, vertically maintained mdr4 roots grew as straight as the wild type, but their gravitropism was enhanced. Upon reorientation, curvature in this mutant developed faster, was distributed more basally, and produced a greater total angle than the wild type. An amplified auxin asymmetry may explain the mdr4 hypertropism. Double mutant analysis indicated that the two auxin transport streams are more independent than interdependent. The hypothesis that flavanols regulate MDR-dependent auxin transport was supported by the epistatic relationship of mdr4 to the tt4 phenylpropanoid pathway mutation.

  5. Structural basis for allosteric cross-talk between the asymmetric nucleotide binding sites of a heterodimeric ABC exporter.

    PubMed

    Hohl, Michael; Hürlimann, Lea M; Böhm, Simon; Schöppe, Jendrik; Grütter, Markus G; Bordignon, Enrica; Seeger, Markus A

    2014-07-29

    ATP binding cassette (ABC) transporters mediate vital transport processes in every living cell. ATP hydrolysis, which fuels transport, displays positive cooperativity in numerous ABC transporters. In particular, heterodimeric ABC exporters exhibit pronounced allosteric coupling between a catalytically impaired degenerate site, where nucleotides bind tightly, and a consensus site, at which ATP is hydrolyzed in every transport cycle. Whereas the functional phenomenon of cooperativity is well described, its structural basis remains poorly understood. Here, we present the apo structure of the heterodimeric ABC exporter TM287/288 and compare it to the previously solved structure with adenosine 5'-(β,γ-imido)triphosphate (AMP-PNP) bound at the degenerate site. In contrast to other ABC exporter structures, the nucleotide binding domains (NBDs) of TM287/288 remain in molecular contact even in the absence of nucleotides, and the arrangement of the transmembrane domains (TMDs) is not influenced by AMP-PNP binding, a notion confirmed by double electron-electron resonance (DEER) measurements. Nucleotide binding at the degenerate site results in structural rearrangements, which are transmitted to the consensus site via two D-loops located at the NBD interface. These loops owe their name from a highly conserved aspartate and are directly connected to the catalytically important Walker B motif. The D-loop at the degenerate site ties the NBDs together even in the absence of nucleotides and substitution of its aspartate by alanine is well-tolerated. By contrast, the D-loop of the consensus site is flexible and the aspartate to alanine mutation and conformational restriction by cross-linking strongly reduces ATP hydrolysis and substrate transport.

  6. Characterization of acquired paclitaxel resistance of breast cancer cells and involvement of ABC transporters

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Němcová-Fürstová, Vlasta, E-mail: vlasta.furstova@

    Development of taxane resistance has become clinically very important issue. The molecular mechanisms underlying the resistance are still unclear. To address this issue, we established paclitaxel-resistant sublines of the SK-BR-3 and MCF-7 breast cancer cell lines that are capable of long-term proliferation in 100 nM and 300 nM paclitaxel, respectively. Application of these concentrations leads to cell death in the original counterpart cells. Both sublines are cross-resistant to doxorubicin, indicating the presence of the MDR phenotype. Interestingly, resistance in both paclitaxel-resistant sublines is circumvented by the second-generation taxane SB-T-1216. Moreover, we demonstrated that it was not possible to establish sublinesmore » of SK-BR-3 and MCF-7 cells resistant to this taxane. It means that at least the tested breast cancer cells are unable to develop resistance to some taxanes. Employing mRNA expression profiling of all known human ABC transporters and subsequent Western blot analysis of the expression of selected transporters, we demonstrated that only the ABCB1/PgP and ABCC3/MRP3 proteins were up-regulated in both paclitaxel-resistant sublines. We found up-regulation of ABCG2/BCRP and ABCC4 proteins only in paclitaxel-resistant SK-BR-3 cells. In paclitaxel-resistant MCF-7 cells, ABCB4/MDR3 and ABCC2/MRP2 proteins were up-regulated. Silencing of ABCB1 expression using specific siRNA increased significantly, but did not completely restore full sensitivity to both paclitaxel and doxorubicin. Thus we showed a key, but not exclusive, role for ABCB1 in mechanisms of paclitaxel resistance. It suggests the involvement of multiple mechanisms in paclitaxel resistance in tested breast cancer cells. - Highlights: • Expression of all ABC transporters in paclitaxel-resistant sublines of SK-BR-3 and MCF-7 cells was analyzed. • SK-BR-3 and MCF-7 cells are unable to develop resistance to some taxanes. • Some taxanes are able to overcome developed resistance to

  7. ATP-binding cassette (ABC) transporters in caprine preantral follicles: gene and protein expression.

    PubMed

    Guerreiro, Denise Damasceno; de Lima, Laritza Ferreira; Mbemya, Gildas Tetaping; Maside, Carolina Mielgo; Miranda, André Marrocos; Tavares, Kaio César Simiano; Alves, Benner Geraldo; Faustino, Luciana Rocha; Smitz, Johan; de Figueiredo, José Ricardo; Rodrigues, Ana Paula Ribeiro

    2018-06-01

    The multidrug resistance proteins ABCB1, ABCC2 and ABCG2 are an energy-dependent efflux pump that functions in systemic detoxification processes. Physiologically expressed in a variety of tissues, most abundantly in the liver and intestinal epithelia, placenta, blood-brain barrier and various stem cells, until now, these pumps were not identified in goat ovarian tissue. Therefore, the aim of this study is to analyze ABCB1, ABCC2, and ABCG2 mRNA and protein expression in goat preantral follicles. Fragments (3 × 3 × 1 mm) from five pairs of ovary (n = 10) obtained from five goat were collected and immediately submitted to qPCR, Western blot, and immunofluorescence assay for mRNA detection and identification and localization of the ABC transporters, respectively. mRNA for ABCB1, ABCC2, and ABCG2 and the presence of their proteins were observed on ovarian tissue samples. Positive marks were observed for the three transport proteins in all follicular categories studied. However, the marks were primarily localized in the oocyte of primordial, transition and primary follicle categories. In conclusion, goat ovarian tissue expresses mRNA for the ABCB1, ABCC2 and ABCG2 transporters and the expression of these proteins in the preantral follicles is a follicle-dependent stage.

  8. The multidrug ABC transporter BmrC/BmrD of Bacillus subtilis is regulated via a ribosome-mediated transcriptional attenuation mechanism.

    PubMed

    Reilman, Ewoud; Mars, Ruben A T; van Dijl, Jan Maarten; Denham, Emma L

    2014-10-01

    Expression of particular drug transporters in response to antibiotic pressure is a critical element in the development of bacterial multidrug resistance, and represents a serious concern for human health. To obtain a better understanding of underlying regulatory mechanisms, we have dissected the transcriptional activation of the ATP-binding cassette (ABC) transporter BmrC/BmrD of the Gram-positive model bacterium Bacillus subtilis. By using promoter-GFP fusions and live cell array technology, we demonstrate a temporally controlled transcriptional activation of the bmrCD genes in response to antibiotics that target protein synthesis. Intriguingly, bmrCD expression only occurs during the late-exponential and stationary growth stages, irrespective of the timing of the antibiotic challenge. We show that this is due to tight transcriptional control by the transition state regulator AbrB. Moreover, our results show that the bmrCD genes are co-transcribed with bmrB (yheJ), a small open reading frame immediately upstream of bmrC that harbors three alternative stem-loop structures. These stem-loops are apparently crucial for antibiotic-induced bmrCD transcription. Importantly, the antibiotic-induced bmrCD expression requires translation of bmrB, which implies that BmrB serves as a regulatory leader peptide. Altogether, we demonstrate for the first time that a ribosome-mediated transcriptional attenuation mechanism can control the expression of a multidrug ABC transporter. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Application of fluorescent dye substrates for functional characterization of ABC multidrug transporters at a single cell level.

    PubMed

    Nerada, Zsuzsanna; Hegyi, Zoltán; Szepesi, Áron; Tóth, Szilárd; Hegedüs, Csilla; Várady, György; Matula, Zsolt; Homolya, László; Sarkadi, Balázs; Telbisz, Ágnes

    2016-09-01

    ABC multidrug transporters are key players in cancer multidrug resistance and in determining the ADME-Tox properties of drugs and xenobiotics. The most sensitive and specific detection of these transporters is based on functional assays. Assessment of the transporter-dependent reduction of cellular uptake of the fluorescent dyes, such as Hoechst 33342 (Ho) and more recently DyeCycle Violet (DCV), have been widely advocated for the characterization of both ABCB1 and ABCG2 multidrug transporters. Detailed comparison of these supravital DNA-binding dyes revealed that DCV is less toxic to ABCG2- and ABCB1-expressing cells than Ho. ATPase measurements imply that DCV and Ho are similarly handled by ABCB1, whereas ABCG2 seems to transport DVC more effectively. In addition, we have developed an image-based high content microscopy screening method for simultaneous in situ measurement of the cellular activity and expression of the ABCG2 multidrug transporter. We demonstrated the applicability of this method for identifying ABCG2-positive cells in heterogeneous cell population by a single dye uptake measurement. These results may promote multidrug transporter studies at a single cell level and allow the quantitative detection of clinically important drug-resistant sub-populations. © 2016 International Society for Advancement of Cytometry. © 2016 International Society for Advancement of Cytometry.

  10. Environmental Conditions Influence Induction of Key ABC-Transporter Genes Affecting Glyphosate Resistance Mechanism in Conyza canadensis.

    PubMed

    Tani, Eleni; Chachalis, Demosthenis; Travlos, Ilias S; Bilalis, Dimitrios

    2016-04-20

    Conyza canadensis has been reported to be the most frequent weed species that evolved resistance to glyphosate in various parts of the world. The objective of the present study was to investigate the effect of environmental conditions (temperature and light) on the expression levels of the EPSPS gene and two major ABC-transporter genes (M10 and M11) on glyphosate susceptible (GS) and glyphosate resistant (GR) horseweed populations, collected from several regions across Greece. Real-time PCR was conducted to determine the expression level of the aforementioned genes when glyphosate was applied at normal (1×; 533 g·a.e.·ha(-1)) and high rates (4×, 8×), measured at an early one day after treatment (DAT) and a later stage (four DAT) of expression. Plants were exposed to light or dark conditions, at three temperature regimes (8, 25, 35 °C). GR plants were made sensitive when exposed to 8 °C with light; those sensitized plants behaved biochemically (shikimate accumulation) and molecularly (expression of EPSPS and ABC-genes) like the GS plants. Results from the current study show the direct link between the environmental conditions and the induction level of the above key genes that likely affect the efficiency of the proposed mechanism of glyphosate resistance.

  11. Environmental Conditions Influence Induction of Key ABC-Transporter Genes Affecting Glyphosate Resistance Mechanism in Conyza canadensis

    PubMed Central

    Tani, Eleni; Chachalis, Demosthenis; Travlos, Ilias S.; Bilalis, Dimitrios

    2016-01-01

    Conyza canadensis has been reported to be the most frequent weed species that evolved resistance to glyphosate in various parts of the world. The objective of the present study was to investigate the effect of environmental conditions (temperature and light) on the expression levels of the EPSPS gene and two major ABC-transporter genes (M10 and M11) on glyphosate susceptible (GS) and glyphosate resistant (GR) horseweed populations, collected from several regions across Greece. Real-time PCR was conducted to determine the expression level of the aforementioned genes when glyphosate was applied at normal (1×; 533 g·a.e.·ha−1) and high rates (4×, 8×), measured at an early one day after treatment (DAT) and a later stage (four DAT) of expression. Plants were exposed to light or dark conditions, at three temperature regimes (8, 25, 35 °C). GR plants were made sensitive when exposed to 8 °C with light; those sensitized plants behaved biochemically (shikimate accumulation) and molecularly (expression of EPSPS and ABC-genes) like the GS plants. Results from the current study show the direct link between the environmental conditions and the induction level of the above key genes that likely affect the efficiency of the proposed mechanism of glyphosate resistance. PMID:27104532

  12. ALS3 encodes a phloem-localized ABC transporter-like protein that is required for aluminum tolerance in Arabidopsis.

    PubMed

    Larsen, Paul B; Geisler, Matt J B; Jones, Carol A; Williams, Kelly M; Cancel, Jesse D

    2005-02-01

    Aluminum (Al) toxicity in acid soils is a worldwide agricultural problem that severely limits crop productivity through inhibition of root growth. Previously, Arabidopsis mutants with increased Al sensitivity were isolated in order to identify genes important for Al tolerance in plants. One mutant, als3, exhibited extreme root growth inhibition in the presence of Al, suggesting that this mutation negatively impacts a gene required for Al tolerance. Map-based cloning of the als3-1 mutation resulted in the isolation of a novel gene that encodes a previously undescribed ABC transporter-like protein, which is highly homologous to a putative bacterial metal resistance protein, ybbM. Northern analysis for ALS3 expression revealed that it is found in all organs examined, which is consistent with the global nature of Al sensitivity displayed by als3, and that expression increases in roots following Al treatment. Based on GUS fusion and in situ hybridization analyses, ALS3 is primarily expressed in leaf hydathodes and the phloem throughout the plant, along with the root cortex following Al treatment. Immunolocalization indicates that ALS3 predominantly accumulates in the plasma membrane of cells that express ALS3. From our results, it appears that ALS3 encodes an ABC transporter-like protein that is required for Al resistance/tolerance and may function to redistribute accumulated Al away from sensitive tissues in order to protect the growing root from the toxic effects of Al.

  13. The feoABC Locus of Yersinia pestis Likely Has Two Promoters Causing Unique Iron Regulation

    PubMed Central

    O'Connor, Lauren; Fetherston, Jacqueline D.; Perry, Robert D.

    2017-01-01

    The FeoABC ferrous transporter is a wide-spread bacterial system. While the feoABC locus is regulated by a number of factors in the bacteria studied, we have previously found that regulation of feoABC in Yersinia pestis appears to be unique. None of the non-iron responsive transcriptional regulators that control expression of feoABC in other bacteria do so in Y. pestis. Another unique factor is the iron and Fur regulation of the Y. pestis feoABC locus occurs during microaerobic but not aerobic growth. Here we show that this unique iron-regulation is not due to a unique aspect of the Y. pestis Fur protein but to DNA sequences that regulate transcription. We have used truncations, alterations, and deletions of the feoA::lacZ reporter to assess the mechanism behind the failure of iron to repress transcription under aerobic conditions. These studies plus EMSAs and DNA sequence analysis have led to our proposal that the feoABC locus has two promoters: an upstream P1 promoter whose expression is relatively iron-independent but repressed under microaerobic conditions and the known downstream Fur-regulated P2 promoter. In addition, we have identified two regions that bind Y. pestis protein(s), although we have not identified these protein(s) or their function. Finally we used iron uptake assays to demonstrate that both FeoABC and YfeABCD transport ferrous iron in an energy-dependent manner and also use ferric iron as a substrate for uptake. PMID:28785546

  14. Requirement of ABC transporter inhibition and Hoechst 33342 dye deprivation for the assessment of side population-defined C6 glioma stem cell metabolism using fluorescent probes.

    PubMed

    Murota, Yoshitaka; Tabu, Kouichi; Taga, Tetsuya

    2016-11-04

    Elucidating the precise properties of cancer stem cells (CSCs) is indispensable for the development of effective therapies against tumors, because CSCs are key drivers of tumor development, metastasis and relapse. We previously reported that the Hoechst 33342 dye-low staining side population (SP) method can enrich for CSCs in the C6 glioma cell line, and that the positively stained main population (MP) cells are non-CSCs. Presence of cancer stem-like SP cells is reported in various types of cancer. Although altered cellular energy metabolism is a hallmark of cancer, very little has been studied on the applicability of fluorescent probes for the understanding of CSC energy metabolism. The metabolic status of C6 SP and MP cells are evaluated by CellROX, MitoTracker Green (MTG) and JC-1 for cellular oxidative stress, mitochondrial amount, and mitochondrial membrane potential, respectively. SP cells were found to exhibit significantly lower fluorescent intensities of CellROX and MTG than MP cells. However, inhibition of ATP binding cassette (ABC) transporters by verapamil enhanced the intensities of these probes in SP cells to the levels similar to those in MP cells, indicating that SP cells expel the probes outside of the cells through ABC transporters. Next, SP cells were stained with JC-1 dye which exhibits membrane potential dependent accumulation in mitochondrial matrix, followed by formation of aggregates. The mitochondrial membrane potential indicated by the aggregates of JC-1 was 5.0-fold lower in SP cells than MP cells. Inhibition of ABC transporters enhanced the fluorescent intensities of the JC-1 aggregates in both SP and MP cells, the former of which was still 2.2-fold lower than the latter. This higher JC-1 signal in MP cells was further found to be due to the Hoechst 33342 dye existing in MP cells. When SP and MP cells were recultured to deprive the intracellular Hoechst 33342 dye and then stained with JC-1 in the presence of verapamil, the intensities of

  15. [Biological characteristics of an enteroinvasive Escherichia coli strain with tatABC deletion].

    PubMed

    Gong, Zhaolong; Ye, Changyun; Liu, Xiaobing; Zhang, Min; Zhuo, Qin

    2013-05-04

    To study the relationship between twin-arginine translocation system (Tat) system with the biological characteristics of enteroinvasive Escherichia coli (EIEC). Through homologous recombination, we constructed EIEC's tatABC gene deletion strain and complementary strain, and explored their impact on bacterial form, substrate transport function as well as on HeLa cells and guinea pig's corneal invasion force. The tatABC gene deletion strain had apparent changes in bacterial form, loss of substrate transporter function, and significant weakened bacterial invasion force (the number of the deletion strain invading into HeLa cells was decreased significantly, and the ability of its corneal lesion capacity of the guinea pig was significantly weakened), while the complementary strain was similar to the wild strain in the above respects. EIEC's Tat protein transport system is closely related with the biological characteristics of EIEC.

  16. Overexpression of the ABC transporter AvtAB increases avermectin production in Streptomyces avermitilis.

    PubMed

    Qiu, Jingfan; Zhuo, Ying; Zhu, Dongqing; Zhou, Xiufen; Zhang, Lixin; Bai, Linquan; Deng, Zixin

    2011-10-01

    Avermectins are 16-membered macrocyclic polyketides with potent antiparasitic activities, produced by Streptomyces avermitilis. Upstream of the avermectin biosynthetic gene cluster, there is the avtAB operon encoding the ABC transporter AvtAB, which is highly homologous to the mammalian multidrug efflux pump P-glycoprotein (Pgp). Inactivation of avtAB had no effect, but increasing the concentration of avtAB mRNA 30-500-fold, using a multi-copy plasmid in S. avermitilis, enhanced avermectin production about two-fold both in the wild-type and in a high-yield producer strain on agar plates. In liquid industrial fermentation medium, the overall productivity of avermectin B1a in the engineered high-yield producer was improved for about 50%, from 3.3 to 4.8 g/l. In liquid YMG medium, moreover, the ratio of intracellular to extracellular accumulation of avermectin B1a was dropped from 6:1 to 4.5:1 in response to multiple copies of avtAB. Additionally, the overexpression of avtAB did not cause any increased expression of the avermectin biosynthetic genes through RT-PCR analysis. We propose that the AvtAB transporter exports avermectin, and thus reduces the feedback inhibition on avermectin production inside the cell. This strategy may be useful for enhancing the production of other antibiotics.

  17. Trichothecene resistance in wheat: Development of molecular markers for PDR-type ABC transporter genes.

    PubMed

    Mitterbauer, R; Heinrich, M; Rauscher, R; Lemmens, M; Bürstmayr, H; Adam, G

    2003-03-01

    Infection withFusarium graminearum andF. culmorum not only causes severe yield and quality losses, the most relevant concern is the contamination of cereal foods and feeds with trichothecenes (e.g. deoxynivalenol, DON). The ability to synthesize trichothecenes has been shown to be a virulence factor ofF. graminearum on wheat and, on the other hand, toxin resistance is most likely an important component of field resistance. Our hypothesis is that pleiotropic drug resistance mediated by PDR-type ABC transporter proteins (acting as membrane located drug efflux pumps) is a relevant mechanism of DON resistance not only in yeast but also in wheat. Goal of this project is the development of molecular markers for this gene family for use in marker-assisted plant breeding programs. The technical difficulties caused by the large size of the PDR-family are discussed.

  18. Accelerated bridge construction (ABC) decision making and economic modeling tool.

    DOT National Transportation Integrated Search

    2011-12-01

    In this FHWA-sponsored pool funded study, a set of decision making tools, based on the Analytic Hierarchy Process (AHP) was developed. This tool set is prepared for transportation specialists and decision-makers to determine if ABC is more effective ...

  19. Alkyl-Lysophospholipid Resistance in Multidrug-Resistant Leishmania tropica and Chemosensitization by a Novel P-Glycoprotein-Like Transporter Modulator

    PubMed Central

    Pérez-Victoria, José M.; Pérez-Victoria, F. Javier; Parodi-Talice, Adriana; Jiménez, Ignacio A.; Ravelo, Angel G.; Castanys, Santiago; Gamarro, Francisco

    2001-01-01

    Drug resistance has emerged as a major impediment in the treatment of leishmaniasis. Alkyl-lysophospholipids (ALP), originally developed as anticancer drugs, are considered to be the most promising antileishmanial agents. In order to anticipate probable clinical failure in the near future, we have investigated possible mechanisms of resistance to these drugs in Leishmania spp. The results presented here support the involvement of a member of the ATP-binding cassette (ABC) superfamily, the Leishmania P-glycoprotein-like transporter, in the resistance to ALP. (i) First, a multidrug resistance (MDR) Leishmania tropica line overexpressing a P-glycoprotein-like transporter displays significant cross-resistance to the ALP miltefosine and edelfosine, with resistant indices of 9.2- and 7.1-fold, respectively. (ii) Reduced expression of P-glycoprotein in the MDR line correlates with a significant decrease in ALP resistance. (iii) The ALP were able to modulate the P-glycoprotein-mediated resistance to daunomycin in the MDR line. (iv) We have found a new inhibitor of this transporter, the sesquiterpene C-3, that completely sensitizes MDR parasites to ALP. (v) Finally, the MDR line exhibits a lower accumulation than the wild-type line of bodipy-C5-PC, a fluorescent analogue of phosphatidylcholine that has a structure resembling that of edelfosine. Also, C-3 significantly increases the accumulation of the fluorescent analogue to levels similar to those of wild-type parasites. The involvement of the Leishmania P-glycoprotein-like transporter in resistance to drugs used in the treatment of leishmaniasis also supports the importance of developing new specific inhibitors of this ABC transporter. PMID:11502516

  20. ATP-binding cassette transporters in reproduction: a new frontier

    PubMed Central

    Bloise, E.; Ortiga-Carvalho, T.M.; Reis, F.M.; Lye, S.J.; Gibb, W.; Matthews, S.G.

    2016-01-01

    BACKGROUND The transmembrane ATP-binding cassette (ABC) transporters actively efflux an array of clinically relevant compounds across biological barriers, and modulate biodistribution of many physiological and pharmacological factors. To date, over 48 ABC transporters have been identified and shown to be directly and indirectly involved in peri-implantation events and fetal/placental development. They efflux cholesterol, steroid hormones, vitamins, cytokines, chemokines, prostaglandins, diverse xenobiotics and environmental toxins, playing a critical role in regulating drug disposition, immunological responses and lipid trafficking, as well as preventing fetal accumulation of drugs and environmental toxins. METHODS This review examines ABC transporters as important mediators of placental barrier functions and key reproductive processes. Expression, localization and function of all identified ABC transporters were systematically reviewed using PubMed and Google Scholar websites to identify relevant studies examining ABC transporters in reproductive tissues in physiological and pathophysiological states. Only reports written in English were incorporated with no restriction on year of publication. While a major focus has been placed on the human, extensive evidence from animal studies is utilized to describe current understanding of the regulation and function of ABC transporters relevant to human reproduction. RESULTS ABC transporters are modulators of steroidogenesis, fertilization, implantation, nutrient transport and immunological responses, and function as ‘gatekeepers’ at various barrier sites (i.e. blood-testes barrier and placenta) against potentially harmful xenobiotic factors, including drugs and environmental toxins. These roles appear to be species dependent and change as a function of gestation and development. The best-described ABC transporters in reproductive tissues (primarily in the placenta) are the multidrug transporters p-glycoprotein and

  1. Marine Natural Products with P-Glycoprotein Inhibitor Properties

    PubMed Central

    Lopez, Dioxelis; Martinez-Luis, Sergio

    2014-01-01

    P-glycoprotein (P-gp) is a protein belonging to the ATP-binding cassette (ABC) transporters superfamily that has clinical relevance due to its role in drug metabolism and multi-drug resistance (MDR) in several human pathogens and diseases. P-gp is a major cause of drug resistance in cancer, parasitic diseases, epilepsy and other disorders. This review article aims to summarize the research findings on the marine natural products with P-glycoprotein inhibitor properties. Natural compounds that modulate P-gp offer great possibilities for semi-synthetic modification to create new drugs and are valuable research tools to understand the function of complex ABC transporters. PMID:24451193

  2. The SUPERFAMILY database in 2004: additions and improvements.

    PubMed

    Madera, Martin; Vogel, Christine; Kummerfeld, Sarah K; Chothia, Cyrus; Gough, Julian

    2004-01-01

    The SUPERFAMILY database provides structural assignments to protein sequences and a framework for analysis of the results. At the core of the database is a library of profile Hidden Markov Models that represent all proteins of known structure. The library is based on the SCOP classification of proteins: each model corresponds to a SCOP domain and aims to represent an entire superfamily. We have applied the library to predicted proteins from all completely sequenced genomes (currently 154), the Swiss-Prot and TrEMBL databases and other sequence collections. Close to 60% of all proteins have at least one match, and one half of all residues are covered by assignments. All models and full results are available for download and online browsing at http://supfam.org. Users can study the distribution of their superfamily of interest across all completely sequenced genomes, investigate with which other superfamilies it combines and retrieve proteins in which it occurs. Alternatively, concentrating on a particular genome as a whole, it is possible first, to find out its superfamily composition, and secondly, to compare it with that of other genomes to detect superfamilies that are over- or under-represented. In addition, the webserver provides the following standard services: sequence search; keyword search for genomes, superfamilies and sequence identifiers; and multiple alignment of genomic, PDB and custom sequences.

  3. Constitutive mRNA expression and protein activity levels of nine ABC efflux transporters in seven permanent cell lines derived from different tissues of rainbow trout (Oncorhynchus mykiss).

    PubMed

    Fischer, Stephan; Loncar, Jovica; Zaja, Roko; Schnell, Sabine; Schirmer, Kristin; Smital, Tvrtko; Luckenbach, Till

    2011-01-25

    Permanent fish cell lines have become common model systems for determining ecotoxicological effects of pollutants. For these cell lines little is known on the cellular active transport mechanisms that control the amount of a compound entering the cell, such as the MXR (multixenobiotic resistance) system mediated by ATP binding cassette (ABC) transport proteins. Therefore, for toxic evaluation of chemicals with those cells information on MXR is important. We here present data on constitutive mRNA expression and protein activity levels of a series of ABC efflux transporters in seven permanent cell lines derived from liver (RTL-W1; R1) and liver hepatoma (RTH-149), gill (RTgill-W1), gonad (RTG-2), gut (RTgutGC) and brain (RTbrain) of rainbow trout (Oncorhynchus mykiss). In addition to known transporters abcb1 (designated here abcb1a), abcb11, abcc1-3, abcc5 and abcg2, we quantified expression levels of a newly identified abcb1 isoform (abcb1b) and abcc4, previously unknown in trout. Quantitative real time PCR (qPCR) indicated that mRNA of the examined ABC transporters was constitutively expressed in all cell lines. Transporter mRNA expression patterns were similar in all cell lines, with expression levels of abcc transporters being 80 to over 1000 fold higher than for abcg2, abcb1a/b and abcb11 (abcc1-5>abcg2>abcb1a/b, 11). Transporter activity in the cell lines was determined by measuring uptake of transporter type specific fluorescent substrates in the presence of activity inhibitors. The combination of the ABCB1 and ABCC transporter substrate calcein-AM with inhibitors cyclosporine A, PSC833 and MK571 resulted in a concentration-dependent fluorescence increase of up to 3-fold, whereas reversin 205 caused a slight, but not concentration-dependent fluorescence increase. Accumulation of the dyes Hoechst 33342 and 2',7'-dichlorodihydrofluorescein diacetate was basically unchanged in the presence of Ko134 and taurocholate, respectively, indicating low Abcg2 and Abcb11

  4. ABC transporter Cdr1p harbors charged residues in the intracellular loop and nucleotide-binding domain critical for protein trafficking and drug resistance.

    PubMed

    Shah, Abdul Haseeb; Banerjee, Atanu; Rawal, Manpreet Kaur; Saxena, Ajay Kumar; Mondal, Alok Kumar; Prasad, Rajendra

    2015-08-01

    The ABC transporter Cdr1 protein of Candida albicans, which plays a major role in antifungal resistance, has two transmembrane domains (TMDs) and two nucleotide-binding domains (NBDs). The 12 transmembrane helices of TMDs that are interconnected by extracellular and intracellular loops (ICLs) mainly harbor substrate recognition sites where drugs bind while cytoplasmic NBDs hydrolyze ATP which powers drug efflux. The coupling of ATP hydrolysis to drug transport requires proper communication between NBDs and TMDs typically accomplished by ICLs. This study examines the role of cytoplasmic ICLs of Cdr1p by rationally predicting the critical residues on the basis of their interatomic distances. Among nine pairs that fall within a proximity of <4 Å, an ion pair between K577 of ICL1 and E315 of NBD1 was found to be critical. The substitution, swapping and changing of the length or charge of K577 or E315 by directed mutagenesis led to a misfolded, non-rescuable protein entrapped in intracellular structures. Furthermore, the equipositional ionic pair-forming residues from ICL3 and NBD2 (R1260 and E1014) did not impact protein trafficking. These results point to a new role for ICL/NBD interacting residues in PDR ABC transporters in protein folding and trafficking. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. Evolution and Diversity of the Ras Superfamily of Small GTPases in Prokaryotes

    PubMed Central

    Wuichet, Kristin; Søgaard-Andersen, Lotte

    2015-01-01

    The Ras superfamily of small GTPases are single domain nucleotide-dependent molecular switches that act as highly tuned regulators of complex signal transduction pathways. Originally identified in eukaryotes for their roles in fundamental cellular processes including proliferation, motility, polarity, nuclear transport, and vesicle transport, recent studies have revealed that single domain GTPases also control complex functions such as cell polarity, motility, predation, development and antibiotic resistance in bacteria. Here, we used a computational genomics approach to understand the abundance, diversity, and evolution of small GTPases in prokaryotes. We collected 520 small GTPase sequences present in 17% of 1,611 prokaryotic genomes analyzed that cover diverse lineages. We identified two discrete families of small GTPases in prokaryotes that show evidence of three distinct catalytic mechanisms. The MglA family includes MglA homologs, which are typically associated with the MglB GTPase activating protein, whereas members of the Rup (Ras superfamily GTPase of unknown function in prokaryotes) family are not predicted to interact with MglB homologs. System classification and genome context analyses support the involvement of small GTPases in diverse prokaryotic signal transduction pathways including two component systems, laying the foundation for future experimental characterization of these proteins. Phylogenetic analysis of prokaryotic and eukaryotic GTPases supports that the last universal common ancestor contained ancestral MglA and Rup family members. We propose that the MglA family was lost from the ancestral eukaryote and that the Ras superfamily members in extant eukaryotes are the result of vertical and horizontal gene transfer events of ancestral Rup GTPases. PMID:25480683

  6. A novel major facilitator superfamily transporter in Penicillium digitatum (PdMFS2) is required for prochloraz resistance, conidiation and full virulence.

    PubMed

    Wu, Zhi; Wang, Shengqiang; Yuan, Yongze; Zhang, Tingfu; Liu, Jing; Liu, Deli

    2016-08-01

    To clone a novel major facilitator superfamily (MFS, a large protein family with diverse physiological functions in all kingdoms) transporter gene, Pdmfs2, and characterize its function in Penicillium digitatum. A novel MFS transporter gene, Pdmfs2, was isolated from P. digitatum. The full-length DNA of Pdmfs2 had a 1590 bp ORF encoding a full-size MFS transporter with 529 amino acids. In a prochloraz-resistant strain (PdHS-F6), Pdmfs2 transcript level was up-regulated compared with the prochloraz-sensitive strain (PdHS-E3) and could be induced by 7 μg prochloraz/ml. The deletion of Pdmfs2 (ΔPdmfs2) in PdHS-F6 led to increased susceptibility to prochloraz and lower EC50 value (the concentration of prochloraz producing 50 % growth inhibition) compared with the PdHS-F6 or complementation strain (COPdmfs2). The ΔPdmfs2 strain was defective in conidia yield and virulence towards citrus fruits, while the complementation of Pdmfs2 could restore the phenotypic features to a large extent. Pdmfs2 is the second MFS transporter gene in P. digitatum and is required for prochloraz resistance, conidiation and full virulence.

  7. Secondary Metabolites from Plants Inhibiting ABC Transporters and Reversing Resistance of Cancer Cells and Microbes to Cytotoxic and Antimicrobial Agents

    PubMed Central

    Wink, Michael; Ashour, Mohamed L.; El-Readi, Mahmoud Zaki

    2012-01-01

    Fungal, bacterial, and cancer cells can develop resistance against antifungal, antibacterial, or anticancer agents. Mechanisms of resistance are complex and often multifactorial. Mechanisms include: (1) Activation of ATP-binding cassette (ABC) transporters, such as P-gp, which pump out lipophilic compounds that have entered a cell, (2) Activation of cytochrome p450 oxidases which can oxidize lipophilic agents to make them more hydrophilic and accessible for conjugation reaction with glucuronic acid, sulfate, or amino acids, and (3) Activation of glutathione transferase, which can conjugate xenobiotics. This review summarizes the evidence that secondary metabolites (SM) of plants, such as alkaloids, phenolics, and terpenoids can interfere with ABC transporters in cancer cells, parasites, bacteria, and fungi. Among the active natural products several lipophilic terpenoids [monoterpenes, diterpenes, triterpenes (including saponins), steroids (including cardiac glycosides), and tetraterpenes] but also some alkaloids (isoquinoline, protoberberine, quinoline, indole, monoterpene indole, and steroidal alkaloids) function probably as competitive inhibitors of P-gp, multiple resistance-associated protein 1, and Breast cancer resistance protein in cancer cells, or efflux pumps in bacteria (NorA) and fungi. More polar phenolics (phenolic acids, flavonoids, catechins, chalcones, xanthones, stilbenes, anthocyanins, tannins, anthraquinones, and naphthoquinones) directly inhibit proteins forming several hydrogen and ionic bonds and thus disturbing the 3D structure of the transporters. The natural products may be interesting in medicine or agriculture as they can enhance the activity of active chemotherapeutics or pesticides or even reverse multidrug resistance, at least partially, of adapted and resistant cells. If these SM are applied in combination with a cytotoxic or antimicrobial agent, they may reverse resistance in a synergistic fashion. PMID:22536197

  8. ABC transporter content diversity in Streptococcus pneumoniae impacts competence regulation and bacteriocin production.

    PubMed

    Wang, Charles Y; Patel, Nisha; Wholey, Wei-Yun; Dawid, Suzanne

    2018-06-19

    The opportunistic pathogen Streptococcus pneumoniae (pneumococcus) uses natural genetic competence to increase its adaptability through horizontal gene transfer. One method of acquiring DNA is through predation of neighboring strains with antimicrobial peptides called "bacteriocins." Competence and production of the major family of pneumococcal bacteriocins, pneumocins, are regulated by the quorum-sensing systems com and blp , respectively. In the classical paradigm, the ABC transporters ComAB and BlpAB each secretes its own system's signaling pheromone and in the case of BlpAB also secretes the pneumocins. While ComAB is found in all pneumococci, only 25% of strains encode an intact version of BlpAB [BlpAB(+)] while the rest do not [BlpAB(-)]. Contrary to the classical paradigm, it was previously shown that BlpAB(-) strains can activate blp through ComAB-mediated secretion of the blp pheromone during brief periods of competence. To better understand the full extent of com - blp crosstalk, we examined the contribution of each transporter to competence development and pneumocin secretion. We found that BlpAB(+) strains have a greater capacity for competence activation through BlpAB-mediated secretion of the com pheromone. Similarly, we show that ComAB and BlpAB are promiscuous and both can secrete pneumocins. Consequently, differences in pneumocin secretion between BlpAB(+) and BlpAB(-) strains derive from the regulation and kinetics of transporter expression rather than substrate specificity. We speculate that BlpAB(-) strains (opportunists) use pneumocins mainly in a narrowly tailored role for DNA acquisition and defense during competence while BlpAB(+) strains (aggressors) expand their use for the general inhibition of rival strains. Copyright © 2018 the Author(s). Published by PNAS.

  9. Interaction of BDE-47 and its Hydroxylated Metabolite 6-OH-BDE-47 with the Human ABC Efflux Transporters P-gp and BCRP: Considerations for Human Exposure and Risk Assessment

    EPA Science Inventory

    ATP binding cassette (ABC) transporters, including P-glycoprotein (P-gp; also known as MDR1, ABCB1) and breast cancer resistance protein (BCRP; also known as ABCG2), are membrane-bound proteins that mediate the cellular efflux of xenobiotics as an important defense against chemic...

  10. The ABC and AUSSAT.

    ERIC Educational Resources Information Center

    McGarritty, Ian

    1985-01-01

    Discusses the Australian Broadcasting Corporation's (ABC) utilization of the AUSSAT telecommunications satellite to extend its television and radio transmission range to reach remote Australian audiences; the satellite's program gathering and interchange capabilities; and ABC's generation of other benefits to offset cost of satellite services.…

  11. Characterization and regulation of the resistance-nodulation-cell division-type multidrug efflux pumps MdtABC and MdtUVW from the fire blight pathogen Erwinia amylovora.

    PubMed

    Pletzer, Daniel; Weingart, Helge

    2014-07-11

    The Gram-negative bacterium Erwinia amylovora is the causal agent of the devastating disease fire blight in rosaceous plants such as apple, pear, quince, raspberry, and cotoneaster. In order to survive and multiply in a host, microbes must be able to circumvent the toxic effects of antimicrobial plant compounds, such as flavonoids and tannins. E. amylovora uses multidrug efflux transporters that recognize and actively export toxic compounds out of the cells. Here, two heterotrimeric resistance-nodulation-cell division (RND)-type multidrug efflux pumps, MdtABC and MdtUVW, from E. amylovora were identified. These RND systems are unusual in that they contain two different RND proteins forming a functional pump. To find the substrate specificities of the two efflux systems, we overexpressed the transporters in a hypersensitive mutant lacking the major RND pump AcrB. Both transporters mediated resistance to several flavonoids, fusidic acid and novobiocin. Additionally, MdtABC mediated resistance towards josamycin, bile salts and silver nitrate, and MdtUVW towards clotrimazole. The ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock was reduced. Quantitative RT-PCR analyses revealed that the expression of the transporter genes was induced during infection of apple rootstock. The polyphenolic plant compound tannin, as well as the heavy metal salt tungstate was found to induce the expression of mdtABC. Finally, the expression of the mdtABC genes was shown to be regulated by BaeR, the response regulator of the two-component system BaeSR, a cell envelope stress response system that controls the adaptive responses to changes in the environment. The expression of MdtABC and MdtUVW is induced during growth of E. amylovora in planta. We identified the plant polyphenol tannin as inducer of mdtABC expression. The reduced ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock suggests that the efflux pumps are involved in

  12. Characterization and regulation of the Resistance-Nodulation-Cell Division-type multidrug efflux pumps MdtABC and MdtUVW from the fire blight pathogen Erwinia amylovora

    PubMed Central

    2014-01-01

    Background The Gram-negative bacterium Erwinia amylovora is the causal agent of the devastating disease fire blight in rosaceous plants such as apple, pear, quince, raspberry, and cotoneaster. In order to survive and multiply in a host, microbes must be able to circumvent the toxic effects of antimicrobial plant compounds, such as flavonoids and tannins. E. amylovora uses multidrug efflux transporters that recognize and actively export toxic compounds out of the cells. Here, two heterotrimeric resistance-nodulation-cell division (RND)-type multidrug efflux pumps, MdtABC and MdtUVW, from E. amylovora were identified. These RND systems are unusual in that they contain two different RND proteins forming a functional pump. Results To find the substrate specificities of the two efflux systems, we overexpressed the transporters in a hypersensitive mutant lacking the major RND pump AcrB. Both transporters mediated resistance to several flavonoids, fusidic acid and novobiocin. Additionally, MdtABC mediated resistance towards josamycin, bile salts and silver nitrate, and MdtUVW towards clotrimazole. The ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock was reduced. Quantitative RT-PCR analyses revealed that the expression of the transporter genes was induced during infection of apple rootstock. The polyphenolic plant compound tannin, as well as the heavy metal salt tungstate was found to induce the expression of mdtABC. Finally, the expression of the mdtABC genes was shown to be regulated by BaeR, the response regulator of the two-component system BaeSR, a cell envelope stress response system that controls the adaptive responses to changes in the environment. Conclusions The expression of MdtABC and MdtUVW is induced during growth of E. amylovora in planta. We identified the plant polyphenol tannin as inducer of mdtABC expression. The reduced ability of the mdtABC- and mdtUVW-deficient mutants to multiply in apple rootstock suggests that the

  13. Caenorhabditis elegans ABCRNAi transporters interact genetically with rde-2 and mut-7.

    PubMed

    Sundaram, Prema; Han, Wang; Cohen, Nancy; Echalier, Benjamin; Albin, John; Timmons, Lisa

    2008-02-01

    RNA interference (RNAi) mechanisms are conserved and consist of an interrelated network of activities that not only respond to exogenous dsRNA, but also perform endogenous functions required in the fine tuning of gene expression and in maintaining genome integrity. Not surprisingly, RNAi functions have widespread influences on cellular function and organismal development. Previously, we observed a reduced capacity to mount an RNAi response in nine Caenorhabditis elegans mutants that are defective in ABC transporter genes (ABC(RNAi) mutants). Here, we report an exhaustive study of mutants, collectively defective in 49 different ABC transporter genes, that allowed for the categorization of one additional transporter into the ABC(RNAi) gene class. Genetic complementation tests reveal functions for ABC(RNAi) transporters in the mut-7/rde-2 branch of the RNAi pathway. These second-site noncomplementation interactions suggest that ABC(RNAi) proteins and MUT-7/RDE-2 function together in parallel pathways and/or as multiprotein complexes. Like mut-7 and rde-2, some ABC(RNAi) mutants display transposon silencing defects. Finally, our analyses reveal a genetic interaction network of ABC(RNAi) gene function with respect to this part of the RNAi pathway. From our results, we speculate that the coordinated activities of ABC(RNAi) transporters, through their effects on endogenous RNAi-related mechanisms, ultimately affect chromosome function and integrity.

  14. Detergent Screening and Purification of the Human Liver ABC Transporters BSEP (ABCB11) and MDR3 (ABCB4) Expressed in the Yeast Pichia pastoris

    PubMed Central

    Stindt, Jan; Smits, Sander H. J.; Schmitt, Lutz

    2013-01-01

    The human liver ATP-binding cassette (ABC) transporters bile salt export pump (BSEP/ABCB11) and the multidrug resistance protein 3 (MDR3/ABCB4) fulfill the translocation of bile salts and phosphatidylcholine across the apical membrane of hepatocytes. In concert with ABCG5/G8, these two transporters are responsible for the formation of bile and mutations within these transporters can lead to severe hereditary diseases. In this study, we report the heterologous overexpression and purification of human BSEP and MDR3 as well as the expression of the corresponding C-terminal GFP-fusion proteins in the yeast Pichia pastoris. Confocal laser scanning microscopy revealed that BSEP-GFP and MDR3-GFP are localized in the plasma membrane of P. pastoris. Furthermore, we demonstrate the first purification of human BSEP and MDR3 yielding ∼1 mg and ∼6 mg per 100 g of wet cell weight, respectively. By screening over 100 detergents using a dot blot technique, we found that only zwitterionic, lipid-like detergents such as Fos-cholines or Cyclofos were able to extract both transporters in sufficient amounts for subsequent functional analysis. For MDR3, fluorescence-detection size exclusion chromatography (FSEC) screens revealed that increasing the acyl chain length of Fos-Cholines improved monodispersity. BSEP purified in n-dodecyl-β-D-maltoside or Cymal-5 after solubilization with Fos-choline 16 from P. pastoris membranes showed binding to ATP-agarose. Furthermore, detergent-solubilized and purified MDR3 showed a substrate-inducible ATPase activity upon addition of phosphatidylcholine lipids. These results form the basis for further biochemical analysis of human BSEP and MDR3 to elucidate the function of these clinically relevant ABC transporters. PMID:23593265

  15. The structure of the human ABC transporter ABCG2 reveals a novel mechanism for drug extrusion.

    PubMed

    Khunweeraphong, Narakorn; Stockner, Thomas; Kuchler, Karl

    2017-10-23

    The human ABC transporter ABCG2 (Breast Cancer Resistance Protein, BCRP) is implicated in anticancer resistance, in detoxification across barriers and linked to gout. Here, we generate a novel atomic model of ABCG2 using the crystal structure of ABCG5/G8. Extensive mutagenesis verifies the structure, disclosing hitherto unrecognized essential residues and domains in the homodimeric ABCG2 transporter. The elbow helix, the first intracellular loop (ICL1) and the nucleotide-binding domain (NBD) constitute pivotal elements of the architecture building the transmission interface that borders a central cavity which acts as a drug trap. The transmission interface is stabilized by salt-bridge interactions between the elbow helix and ICL1, as well as within ICL1, which is essential to control the conformational switch of ABCG2 to the outward-open drug-releasing conformation. Importantly, we propose that ICL1 operates like a molecular spring that holds the NBD dimer close to the membrane, thereby enabling efficient coupling of ATP hydrolysis during the catalytic cycle. These novel mechanistic data open new opportunities to therapeutically target ABCG2 in the context of related diseases.

  16. ABC Transporter Genes and Risk of Type 2 Diabetes

    PubMed Central

    Schou, Jesper; Tybjærg-Hansen, Anne; Møller, Holger J.; Nordestgaard, Børge G.; Frikke-Schmidt, Ruth

    2012-01-01

    OBJECTIVE Alterations of pancreatic β-cell cholesterol content may contribute to β-cell dysfunction. Two important determinants of intracellular cholesterol content are the ATP-binding cassette (ABC) transporters A1 (ABCA1) and -G1 (ABCG1). Whether genetic variation in ABCA1 and ABCG1 predicts risk of type 2 diabetes in the general population is unknown. RESEARCH DESIGN AND METHODS We tested whether genetic variation in the promoter and coding regions of ABCA1 and ABCG1 predicted risk of type 2 diabetes in the general population. Twenty-seven variants, identified by previous resequencing of both genes, were genotyped in the Copenhagen City Heart Study (CCHS) (n = 10,185). Two loss-of-function mutations (ABCA1 N1800H and ABCG1 g.-376C>T) (n = 322) and a common variant (ABCG1 g.-530A>G) were further genotyped in the Copenhagen General Population Study (CGPS) (n = 30,415). RESULTS Only one of the variants examined, ABCG1 g.-530A>G, predicted a decreased risk of type 2 diabetes in the CCHS (P for trend = 0.05). Furthermore, when validated in the CGPS or in the CCHS and CGPS combined (n = 40,600), neither the two loss-of-function mutations (ABCA1 N1800H, ABCG1 g.-376C>T) nor ABCG1 g.-530A>G were associated with type 2 diabetes (P values >0.57 and >0.30, respectively). CONCLUSIONS Genetic variations in ABCA1 and ABCG1 were not associated with increased risk of type 2 diabetes in the general population. These data were obtained in general population samples harboring the largest number of heterozygotes for loss-of-function mutations in ABCA1 and ABCG1. PMID:23139370

  17. Complete sequence of Enterococcus faecium pVEF3 and the detection of an omega-epsilon-zeta toxin-antitoxin module and an ABC transporter.

    PubMed

    Sletvold, H; Johnsen, P J; Hamre, I; Simonsen, G S; Sundsfjord, A; Nielsen, K M

    2008-07-01

    Glycopeptide resistant Enterococcus faecium (GREF) persists on Norwegian poultry farms despite the ban on the growth promoter avoparcin. The biological basis for long-term persistence of avoparcin resistance is not fully understood. This study presents the complete DNA sequence of the E. faecium R-plasmid pVEF3 and functional studies of some plasmid-encoded traits (a toxin-antitoxin (TA) system and an ABC transporter) that may be of importance for plasmid persistence. The pVEF3 (63.1 kbp), isolated from an E. faecium strain of poultry origin sampled in Norway in 1999, has 71 coding sequences including the vanA avoparcin/vancomycin resistance encoding gene cluster. pVEF3 encodes the TA system omega-epsilon-zeta, and plasmid stability tests and transcription analysis show that omega-epsilon-zeta is functional in Enterococcus faecalis OGIX, although with decreasing effect over time. The predicted ABC transporter was not found to confer reduced susceptibility to any of the 28 substances tested. The TA system identified in the pVEF-type plasmids may contribute to vanA plasmid persistence on Norwegian poultry farms. However, size and compositional heterogeneity among E. faecium vanA plasmids suggest that additional plasmid maintenance systems in combination with host specific factors and frequent horizontal gene transfer and rearrangement causes the observed plasmid composition and distribution patterns.

  18. In Vitro Reassembly of the Ribose ATP-binding Cassette Transporter Reveals a Distinct Set of Transport Complexes*

    PubMed Central

    Clifton, Matthew C.; Simon, Michael J.; Erramilli, Satchal K.; Zhang, Huide; Zaitseva, Jelena; Hermodson, Mark A.; Stauffacher, Cynthia V.

    2015-01-01

    Bacterial ATP-binding cassette (ABC) importers are primary active transporters that are critical for nutrient uptake. Based on structural and functional studies, ABC importers can be divided into two distinct classes, type I and type II. Type I importers follow a strict alternating access mechanism that is driven by the presence of the substrate. Type II importers accept substrates in a nucleotide-free state, with hydrolysis driving an inward facing conformation. The ribose transporter in Escherichia coli is a tripartite complex consisting of a cytoplasmic ATP-binding cassette protein, RbsA, with fused nucleotide binding domains; a transmembrane domain homodimer, RbsC2; and a periplasmic substrate binding protein, RbsB. To investigate the transport mechanism of the complex RbsABC2, we probed intersubunit interactions by varying the presence of the substrate ribose and the hydrolysis cofactors, ATP/ADP and Mg2+. We were able to purify a full complex, RbsABC2, in the presence of stable, transition state mimics (ATP, Mg2+, and VO4); a RbsAC complex in the presence of ADP and Mg2+; and a heretofore unobserved RbsBC complex in the absence of cofactors. The presence of excess ribose also destabilized complex formation between RbsB and RbsC. These observations suggest that RbsABC2 shares functional traits with both type I and type II importers, as well as possessing unique features, and employs a distinct mechanism relative to other ABC transporters. PMID:25533465

  19. Evolution of Enzyme Superfamilies: Comprehensive Exploration of Sequence-Function Relationships.

    PubMed

    Baier, F; Copp, J N; Tokuriki, N

    2016-11-22

    The sequence and functional diversity of enzyme superfamilies have expanded through billions of years of evolution from a common ancestor. Understanding how protein sequence and functional "space" have expanded, at both the evolutionary and molecular level, is central to biochemistry, molecular biology, and evolutionary biology. Integrative approaches that examine protein sequence, structure, and function have begun to provide comprehensive views of the functional diversity and evolutionary relationships within enzyme superfamilies. In this review, we outline the recent advances in our understanding of enzyme evolution and superfamily functional diversity. We describe the tools that have been used to comprehensively analyze sequence relationships and to characterize sequence and function relationships. We also highlight recent large-scale experimental approaches that systematically determine the activity profiles across enzyme superfamilies. We identify several intriguing insights from this recent body of work. First, promiscuous activities are prevalent among extant enzymes. Second, many divergent proteins retain "function connectivity" via enzyme promiscuity, which can be used to probe the evolutionary potential and history of enzyme superfamilies. Finally, we discuss open questions regarding the intricacies of enzyme divergence, as well as potential research directions that will deepen our understanding of enzyme superfamily evolution.

  20. SUPERFAMILY 1.75 including a domain-centric gene ontology method.

    PubMed

    de Lima Morais, David A; Fang, Hai; Rackham, Owen J L; Wilson, Derek; Pethica, Ralph; Chothia, Cyrus; Gough, Julian

    2011-01-01

    The SUPERFAMILY resource provides protein domain assignments at the structural classification of protein (SCOP) superfamily level for over 1400 completely sequenced genomes, over 120 metagenomes and other gene collections such as UniProt. All models and assignments are available to browse and download at http://supfam.org. A new hidden Markov model library based on SCOP 1.75 has been created and a previously ignored class of SCOP, coiled coils, is now included. Our scoring component now uses HMMER3, which is in orders of magnitude faster and produces superior results. A cloud-based pipeline was implemented and is publicly available at Amazon web services elastic computer cloud. The SUPERFAMILY reference tree of life has been improved allowing the user to highlight a chosen superfamily, family or domain architecture on the tree of life. The most significant advance in SUPERFAMILY is that now it contains a domain-based gene ontology (GO) at the superfamily and family levels. A new methodology was developed to ensure a high quality GO annotation. The new methodology is general purpose and has been used to produce domain-based phenotypic ontologies in addition to GO.

  1. Structure-Based Phylogenetic Analysis of the Lipocalin Superfamily.

    PubMed

    Lakshmi, Balasubramanian; Mishra, Madhulika; Srinivasan, Narayanaswamy; Archunan, Govindaraju

    2015-01-01

    Lipocalins constitute a superfamily of extracellular proteins that are found in all three kingdoms of life. Although very divergent in their sequences and functions, they show remarkable similarity in 3-D structures. Lipocalins bind and transport small hydrophobic molecules. Earlier sequence-based phylogenetic studies of lipocalins highlighted that they have a long evolutionary history. However the molecular and structural basis of their functional diversity is not completely understood. The main objective of the present study is to understand functional diversity of the lipocalins using a structure-based phylogenetic approach. The present study with 39 protein domains from the lipocalin superfamily suggests that the clusters of lipocalins obtained by structure-based phylogeny correspond well with the functional diversity. The detailed analysis on each of the clusters and sub-clusters reveals that the 39 lipocalin domains cluster based on their mode of ligand binding though the clustering was performed on the basis of gross domain structure. The outliers in the phylogenetic tree are often from single member families. Also structure-based phylogenetic approach has provided pointers to assign putative function for the domains of unknown function in lipocalin family. The approach employed in the present study can be used in the future for the functional identification of new lipocalin proteins and may be extended to other protein families where members show poor sequence similarity but high structural similarity.

  2. Structure and function of POTRA domains of Omp85/TPS superfamily.

    PubMed

    Simmerman, Richard F; Dave, Ashita M; Bruce, Barry D

    2014-01-01

    The Omp85/TPS (outer-membrane protein of 85 kDa/two-partner secretion) superfamily is a ubiquitous and major class of β-barrel proteins. This superfamily is restricted to the outer membranes of gram-negative bacteria, mitochondria, and chloroplasts. The common architecture, with an N-terminus consisting of repeats of soluble polypeptide-transport-associated (POTRA) domains and a C-terminal β-barrel pore is highly conserved. The structures of multiple POTRA domains and one full-length TPS protein have been solved, yet discovering roles of individual POTRA domains has been difficult. This review focuses on similarities and differences between POTRA structures, emphasizing POTRA domains in autotrophic organisms including plants and cyanobacteria. Unique roles, specific for certain POTRA domains, are examined in the context of POTRA location with respect to their attachment to the β-barrel pore, and their degree of biological dispensability. Finally, because many POTRA domains may have the ability to interact with thousands of partner proteins, possible modes of these interactions are also explored. © 2014 Elsevier Inc. All rights reserved.

  3. Structural diversity of domain superfamilies in the CATH database.

    PubMed

    Reeves, Gabrielle A; Dallman, Timothy J; Redfern, Oliver C; Akpor, Adrian; Orengo, Christine A

    2006-07-14

    The CATH database of domain structures has been used to explore the structural variation of homologous domains in 294 well populated domain structure superfamilies, each containing at least three sequence diverse relatives. Our analyses confirm some previously detected trends relating sequence divergence to structural variation but for a much larger dataset and in some superfamilies the new data reveal exceptional structural variation. Use of a new algorithm (2DSEC) to analyse variability in secondary structure compositions across a superfamily sheds new light on how structures evolve. 2DSEC detects inserted secondary structures that embellish the core of conserved secondary structures found throughout the superfamily. Analysis showed that for 56% of highly populated superfamilies (>9 sequence diverse relatives), there are twofold or more increases in the numbers of secondary structures in some relatives. In some families fivefold increases occur, sometimes modifying the fold of the domain. Manual inspection of secondary structure insertions or embellishments in 48 particularly variable superfamilies revealed that although these insertions were usually discontiguous in the sequence they were often co-located in 3D resulting in a larger structural motif that often modified the geometry of the active site or the surface conformation promoting diverse domain partnerships and protein interactions. These observations, supported by automatic analysis of all well populated CATH families, suggest that accretion of small secondary structure insertions may provide a simple mechanism for evolving new functions in diverse relatives. Some layered domain architectures (e.g. mainly-beta and alpha-beta sandwiches) that recur highly in the genomes more frequently exploit these types of embellishments to modify function. In these architectures, aggregation occurs most often at the edges, top or bottom of the beta-sheets. Information on structural variability across domain

  4. Basic Residues R260 and K357 Affect the Conformational Dynamics of the Major Facilitator Superfamily Multidrug Transporter LmrP

    PubMed Central

    Wang, Wei; van Veen, Hendrik W.

    2012-01-01

    Secondary-active multidrug transporters can confer resistance on cells to pharmaceuticals by mediating their extrusion away from intracellular targets via substrate/H+(Na+) antiport. While the interactions of catalytic carboxylates in these transporters with coupling ions and substrates (drugs) have been studied in some detail, the functional importance of basic residues has received much less attention. The only two basic residues R260 and K357 in transmembrane helices in the Major Facilitator Superfamily transporter LmrP from Lactococcus lactis are present on the outer surface of the protein, where they are exposed to the phospholipid head group region of the outer leaflet (R260) and inner leaflet (K357) of the cytoplasmic membrane. Although our observations on the proton-motive force dependence and kinetics of substrate transport, and substrate-dependent proton transport demonstrate that K357A and R260A mutants are affected in ethidium-proton and benzalkonium-proton antiport compared to wildtype LmrP, our findings suggest that R260 and K357 are not directly involved in the binding of substrates or the translocation of protons. Secondary-active multidrug transporters are thought to operate by a mechanism in which binding sites for substrates are alternately exposed to each face of the membrane. Disulfide crosslinking experiments were performed with a double cysteine mutant of LmrP that reports the substrate-stimulated transition from the outward-facing state to the inward-facing state with high substrate-binding affinity. In the experiments, the R260A and K357A mutations were found to influence the dynamics of these major protein conformations in the transport cycle, potentially by removing the interactions of R260 and K357 with phospholipids and/or other residues in LmrP. The R260A and K357A mutations therefore modify the maximum rate at which the transport cycle can operate and, as the transitions between conformational states are differently affected by

  5. A single-component multidrug transporter of the major facilitator superfamily is part of a network that protects E scherichia coli from bile salt stress

    PubMed Central

    Paul, Stephanie; Alegre, Kamela O; Holdsworth, Scarlett R; Rice, Matthew; Brown, James A; McVeigh, Paul; Kelly, Sharon M; Law, Christopher J

    2014-01-01

    Resistance to high concentrations of bile salts in the human intestinal tract is vital for the survival of enteric bacteria such as E scherichia coli. Although the tripartite AcrAB–TolC efflux system plays a significant role in this resistance, it is purported that other efflux pumps must also be involved. We provide evidence from a comprehensive suite of experiments performed at two different pH values (7.2 and 6.0) that reflect pH conditions that E . coli may encounter in human gut that MdtM, a single-component multidrug resistance transporter of the major facilitator superfamily, functions in bile salt resistance in E . coli by catalysing secondary active transport of bile salts out of the cell cytoplasm. Furthermore, assays performed on a chromosomal ΔacrB mutant transformed with multicopy plasmid encoding MdtM suggested a functional synergism between the single-component MdtM transporter and the tripartite AcrAB–TolC system that results in a multiplicative effect on resistance. Substrate binding experiments performed on purified MdtM demonstrated that the transporter binds to cholate and deoxycholate with micromolar affinity, and transport assays performed on inverted vesicles confirmed the capacity of MdtM to catalyse electrogenic bile salt/H+ antiport. PMID:24684269

  6. Two different groups of signal sequence in M-superfamily conotoxins.

    PubMed

    Wang, Qi; Jiang, Hui; Han, Yu-Hong; Yuan, Duo-Duo; Chi, Cheng-Wu

    2008-04-01

    M-superfamily conotoxins can be divided into four branches (M-1, M-2, M-3 and M-4) according to the number of amino acid residues in the third Cys loop. In general, it is widely accepted that the conotoxin signal peptides of each superfamily are strictly conserved. Recently, we cloned six cDNAs of novel M-superfamily conotoxins from Conus leopardus, Conus marmoreus and Conus quercinus, belonging to either M-1 or M-3 branch. These conotoxins, judging from the putative peptide sequences deducted from cDNAs, are rich in acidic residues and share highly conserved signal and pro-peptide region. However, they are quite different from the reported conotoxins of M-2 and M-4 branches even in their signal peptides, which in general are considered highly conserved for each superfamily of conotoxins. The signal sequences of M-1 and M-3 conotoxins composed of 24 residues start with MLKMGVVL-, while those of M-2 and M-4 conotoxins composed of 25 residues start with MMSKLGVL-. It is another example that different types of signal peptides can exist within a superfamily besides the I-conotoxin superfamily. In addition to the different disulfide connectivity of M-1 conotoxins from that of M-4 or M-2 conotoxins, the sequence alignment, preferential Cys codon usage and phylogenetic tree analysis suggest that M-1 and M-3 conotoxins have much closer relationship, being different from the conotoxins of other two branches (M-4 and M-2) of M-superfamily.

  7. An operon from Lactobacillus helveticus composed of a proline iminopeptidase gene (pepI) and two genes coding for putative members of the ABC transporter family of proteins.

    PubMed

    Varmanen, P; Rantanen, T; Palva, A

    1996-12-01

    A proline iminopeptidase gene (pepI) of an industrial Lactobacillus helveticus strain was cloned and found to be organized in an operon-like structure of three open reading frames (ORF1, ORF2 and ORF3). ORF1 was preceded by a typical prokaryotic promoter region, and a putative transcription terminator was found downstream of ORF3, identified as the pepI gene. Using primer-extension analyses, only one transcription start site, upstream of ORF1, was identifiable in the predicted operon. Although the size of mRNA could not be judged by Northern analysis either with ORF1-, ORF2- or pepI-specific probes, reverse transcription-PCR analyses further supported the operon structure of the three genes. ORF1, ORF2 and ORF3 had coding capacities for 50.7, 24.5 and 33.8 kDa proteins, respectively. The ORF3-encoded PepI protein showed 65% identity with the PepI proteins from Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus delbrueckii subsp. lactis. The ORF1-encoded protein had significant homology with several members of the ABC transporter family but, with two distinct putative ATP-binding sites, it would represent an unusual type among the bacterial ABC transporters. ORF2 encoded a putative integral membrane protein also characteristic of the ABC transporter family. The pepI gene was overexpressed in Escherichia coli. Purified PepI hydrolysed only di and tripeptides with proline in the first position. Optimum PepI activity was observed at pH 7.5 and 40 degrees C. A gel filtration analysis indicated that PepI is a dimer of M(r) 53,000. PepI was shown to be a metal-independent serine peptidase having thiol groups at or near the active site. Kinetic studies with proline-p-nitroanilide as substrate revealed Km and Vmax values of 0.8 mM and 350 mmol min-1 mg-1, respectively, and a very high turnover number of 135,000 s-1.

  8. Rice SPX-Major Facility Superfamily3, a Vacuolar Phosphate Efflux Transporter, Is Involved in Maintaining Phosphate Homeostasis in Rice1[OPEN

    PubMed Central

    Ying, Yinghui; Wang, Shoudong; Secco, David; Liu, Yu; Whelan, James; Tyerman, Stephen D.; Shou, Huixia

    2015-01-01

    To maintain a stable cytosol phosphate (Pi) concentration, plant cells store Pi in their vacuoles. When the Pi concentration in the cytosol decreases, Pi is exported from the vacuole into the cytosol. This export is mediated by Pi transporters on the tonoplast. In this study, we demonstrate that SYG1, PHO81, and XPR1 (SPX)-Major Facility Superfamily (MFS) proteins have a similar structure with yeast (Saccharomyces cerevisiae) low-affinity Pi transporters Phosphatase87 (PHO87), PHO90, and PHO91. OsSPX-MFS1, OsSPX-MFS2, and OsSPX-MFS3 all localized on the tonoplast of rice (Oryza sativa) protoplasts, even in the absence of the SPX domain. At high external Pi concentration, OsSPX-MFS3 could partially complement the yeast mutant strain EY917 under pH 5.5, which lacks all five Pi transporters present in yeast. In oocytes, OsSPX-MFS3 was shown to facilitate Pi influx or efflux depending on the external pH and Pi concentrations. In contrast to tonoplast localization in plants cells, OsSPX-MFS3 was localized to the plasma membrane when expressed in both yeast and oocytes. Overexpression of OsSPX-MFS3 results in decreased Pi concentration in the vacuole of rice tissues. We conclude that OsSPX-MFS3 is a low-affinity Pi transporter that mediates Pi efflux from the vacuole into cytosol and is coupled to proton movement. PMID:26424157

  9. Origin and evolution of TNF and TNF receptor superfamilies

    USDA-ARS?s Scientific Manuscript database

    The tumor necrosis factor superfamily (TNFSF) and the TNF receptor superfamily (TNFRSF) have an ancient evolutionary origin that can be traced back to single copy genes within Arthropods. In humans, 18 TNFSF and 29 TNFRSF genes have been identified. Evolutionary models account for the increase in g...

  10. ABC's of Construction. Final Report.

    ERIC Educational Resources Information Center

    Greater Baton Rouge Chamber of Commerce, LA.

    The ABC's of Construction project was a demonstration project designed to integrate basic skills training with an industry-developed vocational-craft training program. The program was located at the central training facility of the Pelican Chapter of Associated Builders and Contractors (ABC), an organization made up of nearly 300 member companies…

  11. Statistical Hypothesis Testing in Intraspecific Phylogeography: NCPA versus ABC

    PubMed Central

    Templeton, Alan R.

    2009-01-01

    Nested clade phylogeographic analysis (NCPA) and approximate Bayesian computation (ABC) have been used to test phylogeographic hypotheses. Multilocus NCPA tests null hypotheses, whereas ABC discriminates among a finite set of alternatives. The interpretive criteria of NCPA are explicit and allow complex models to be built from simple components. The interpretive criteria of ABC are ad hoc and require the specification of a complete phylogeographic model. The conclusions from ABC are often influenced by implicit assumptions arising from the many parameters needed to specify a complex model. These complex models confound many assumptions so that biological interpretations are difficult. Sampling error is accounted for in NCPA, but ABC ignores important sources of sampling error that creates pseudo-statistical power. NCPA generates the full sampling distribution of its statistics, but ABC only yields local probabilities, which in turn make it impossible to distinguish between a good fitting model, a non-informative model, and an over-determined model. Both NCPA and ABC use approximations, but convergences of the approximations used in NCPA are well defined whereas those in ABC are not. NCPA can analyze a large number of locations, but ABC cannot. Finally, the dimensionality of tested hypothesis is known in NCPA, but not for ABC. As a consequence, the “probabilities” generated by ABC are not true probabilities and are statistically non-interpretable. Accordingly, ABC should not be used for hypothesis testing, but simulation approaches are valuable when used in conjunction with NCPA or other methods that do not rely on highly parameterized models. PMID:19192182

  12. Overcoming Multidrug Resistance in Human Cancer Cells by Natural Compounds

    PubMed Central

    Nabekura, Tomohiro

    2010-01-01

    Multidrug resistance is a phenomenon whereby tumors become resistant to structurally unrelated anticancer drugs. P-glycoprotein belongs to the large ATP-binding cassette (ABC) transporter superfamily of membrane transport proteins. P-glycoprotein mediates resistance to various classes of anticancer drugs including vinblastine, daunorubicin, and paclitaxel, by actively extruding the drugs from the cells. The quest for inhibitors of anticancer drug efflux transporters has uncovered natural compounds, including (-)-epigallocatechin gallate, curcumin, capsaicin, and guggulsterone, as promising candidates. In this review, studies on the effects of natural compounds on P-glycoprotein and anticancer drug efflux transporters are summarized. PMID:22069634

  13. MetaSINEs: Broad Distribution of a Novel SINE Superfamily in Animals

    PubMed Central

    Nishihara, Hidenori; Plazzi, Federico; Passamonti, Marco; Okada, Norihiro

    2016-01-01

    SINEs (short interspersed elements) are transposable elements that typically originate independently in each taxonomic clade (order/family). However, some SINE families share a highly similar central sequence and are thus categorized as a SINE superfamily. Although only four SINE superfamilies (CORE-SINEs, V-SINEs, DeuSINEs, and Ceph-SINEs) have been reported so far, it is expected that new SINE superfamilies would be discovered by deep exploration of new SINEs in metazoan genomes. Here we describe 15 SINEs, among which 13 are novel, that have a similar 66-bp central region and therefore constitute a new SINE superfamily, MetaSINEs. MetaSINEs are distributed from fish to cnidarians, suggesting their common evolutionary origin at least 640 Ma. Because the 3′ tails of MetaSINEs are variable, these SINEs most likely survived by changing their partner long interspersed elements for retrotransposition during evolution. Furthermore, we examined the presence of members of other SINE superfamilies in bivalve genomes and characterized eight new SINEs belonging to the CORE-SINEs, V-SINEs, and DeuSINEs, in addition to the MetaSINEs. The broad distribution of bivalve SINEs suggests that at least three SINEs originated in the common ancestor of Bivalvia. Our comparative analysis of the central domains of the SINEs revealed that, in each superfamily, only a restricted region is shared among all of its members. Because the functions of the central domains of the SINE superfamilies remain unknown, such structural information of SINE superfamilies will be useful for future experimental and comparative analyses to reveal why they have been retained in metazoan genomes during evolution. PMID:26872770

  14. Do You Know Your ABC?

    ERIC Educational Resources Information Center

    Neale, Claire

    2013-01-01

    Within primary schools, the core subjects of literacy and numeracy are highly regarded, and rightly so, as children need to learn to read, write and be numerically literate. This means that all children learn their ABCs at an early age, But, what about the "other ABC"--"Airway, Breathing and Circulation?" Accidents and medical…

  15. Hydrogenase activity in the foodborne pathogen Campylobacter jejuni depends upon a novel ABC-type nickel transporter (NikZYXWV) and is SlyD-independent.

    PubMed

    Howlett, Robert M; Hughes, Bethan M; Hitchcock, Andrew; Kelly, David J

    2012-06-01

    Campylobacter jejuni is a human pathogen of worldwide significance. It is commensal in the gut of many birds and mammals, where hydrogen is a readily available electron donor. The bacterium possesses a single membrane-bound, periplasmic-facing NiFe uptake hydrogenase that depends on the acquisition of environmental nickel for activity. The periplasmic binding protein Cj1584 (NikZ) of the ATP binding cassette (ABC) transporter encoded by the cj1584c-cj1580c (nikZYXWV) operon in C. jejuni strain NCTC 11168 was found to be nickel-repressed and to bind free nickel ions with a submicromolar K(d) value, as measured by fluorescence spectroscopy. Unlike the Escherichia coli NikA protein, NikZ did not bind EDTA-chelated nickel and lacks key conserved residues implicated in metallophore interaction. A C. jejuni cj1584c null mutant strain showed an approximately 22-fold decrease in intracellular nickel content compared with the wild-type strain and a decreased rate of uptake of (63)NiCl(2). The inhibition of residual nickel uptake at higher nickel concentrations in this mutant by hexa-ammine cobalt (III) chloride or magnesium ions suggests that low-affinity uptake occurs partly through the CorA magnesium transporter. Hydrogenase activity was completely abolished in the cj1584c mutant after growth in unsupplemented media, but was fully restored after growth with 0.5 mM nickel chloride. Mutation of the putative metallochaperone gene slyD (cj0115) had no effect on either intracellular nickel accumulation or hydrogenase activity. Our data reveal a strict dependence of hydrogenase activity in C. jejuni on high-affinity nickel uptake through an ABC transporter that has distinct properties compared with the E. coli Nik system.

  16. Mutations in the Arabidopsis Peroxisomal ABC Transporter COMATOSE Allow Differentiation between Multiple Functions In Planta: Insights from an Allelic Series

    PubMed Central

    Dietrich, Daniela; Schmuths, Heike; Lousa, Carine De Marcos; Baldwin, Jocelyn M.; Baldwin, Stephen A.; Baker, Alison; Holdsworth, Michael J.

    2009-01-01

    COMATOSE (CTS), the Arabidopsis homologue of human Adrenoleukodystrophy protein (ALDP), is required for import of substrates for peroxisomal β-oxidation. A new allelic series and a homology model based on the bacterial ABC transporter, Sav1866, provide novel insights into structure-function relations of ABC subfamily D proteins. In contrast to ALDP, where the majority of mutations result in protein absence from the peroxisomal membrane, all CTS mutants produced stable protein. Mutation of conserved residues in the Walker A and B motifs in CTS nucleotide-binding domain (NBD) 1 resulted in a null phenotype but had little effect in NBD2, indicating that the NBDs are functionally distinct in vivo. Two alleles containing mutations in NBD1 outside the Walker motifs (E617K and C631Y) exhibited resistance to auxin precursors 2,4-dichlorophenoxybutyric acid (2,4-DB) and indole butyric acid (IBA) but were wild type in all other tests. The homology model predicted that the transmission interfaces are domain-swapped in CTS, and the differential effects of mutations in the conserved “EAA motif” of coupling helix 2 supported this prediction, consistent with distinct roles for each NBD. Our findings demonstrate that CTS functions can be separated by mutagenesis and the structural model provides a framework for interpretation of phenotypic data. PMID:19019987

  17. Comparative analysis of cation/proton antiporter superfamily in plants.

    PubMed

    Ye, Chu-Yu; Yang, Xiaohan; Xia, Xinli; Yin, Weilun

    2013-06-01

    The cation/proton antiporter superfamily is associated with the transport of monovalent cations across membranes. This superfamily was annotated in the Arabidopsis genome and some members were functionally characterized. In the present study, a systematic analysis of the cation/proton antiporter genes in diverse plant species was reported. We identified 240 cation/proton antiporters in alga, moss, and angiosperm. A phylogenetic tree was constructed showing these 240 members are separated into three families, i.e., Na(+)/H(+) exchangers, K(+) efflux antiporters, and cation/H(+) exchangers. Our analysis revealed that tandem and/or segmental duplications contribute to the expansion of cation/H(+) exchangers in the examined angiosperm species. Sliding window analysis of the nonsynonymous/synonymous substitution ratios showed some differences in the evolutionary fate of cation/proton antiporter paralogs. Furthermore, we identified over-represented motifs among these 240 proteins and found most motifs are family specific, demonstrating diverse evolution of the cation/proton antiporters among three families. In addition, we investigated the co-expressed genes of the cation/proton antiporters in Arabidopsis thaliana. The results showed some biological processes are enriched in the co-expressed genes, suggesting the cation/proton antiporters may be involved in these biological processes. Taken together, this study furthers our knowledge on cation/proton antiporters in plants. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Structure, function, and evolution of bacterial ATP-binding cassette systems

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Davidson, A.L.; Dassa, E.; Orelle, C.

    2010-07-27

    The ATP-binding cassette (ABC) systems constitute one of the largest superfamilies of paralogous sequences. All ABC systems share a highly conserved ATP-hydrolyzing domain or protein (the ABC; also referred to as a nucleotide-binding domain [NBD]) that is unequivocally characterized by three short sequence motifs (Fig. 1): these are the Walker A and Walker B motifs, indicative of the presence of a nucleotide-binding site, and the signature motif, unique to ABC proteins, located upstream of the Walker B motif (426). Other motifs diagnostic of ABC proteins are also indicated in Fig. 1. The biological significance of these motifs is discussed inmore » Structure, Function, and Dynamics of the ABC. ABC systems are widespread among living organisms and have been detected in all genera of the three kingdoms of life, with remarkable conservation in the primary sequence of the cassette and in the organization of the constitutive domains or subunits (203, 420). ABC systems couple the energy of ATP hydrolysis to an impressively large variety of essential biological phenomena, comprising not only transmembrane (TM) transport, for which they are best known, but also several non-transport-related processes, such as translation elongation (62) and DNA repair (174). Although ABC systems deserve much attention because they are involved in severe human inherited diseases (107), they were first discovered and characterized in detail in prokaryotes, as early as the 1970s (13, 148, 238, 468). The most extensively analyzed systems were the high-affinity histidine and maltose uptake systems of Salmonella enterica serovar Typhimurium and Escherichia coli. Over 2 decades ago, after the completion of the nucleotide sequences encoding these transporters in the respective laboratories of Giovanna Ames and Maurice Hofnung, Hiroshi Nikaido and colleagues noticed that the two systems displayed a global similarity in the nature of their components and, moreover, that the primary sequences of

  19. MetaSINEs: Broad Distribution of a Novel SINE Superfamily in Animals.

    PubMed

    Nishihara, Hidenori; Plazzi, Federico; Passamonti, Marco; Okada, Norihiro

    2016-02-12

    SINEs (short interspersed elements) are transposable elements that typically originate independently in each taxonomic clade (order/family). However, some SINE families share a highly similar central sequence and are thus categorized as a SINE superfamily. Although only four SINE superfamilies (CORE-SINEs, V-SINEs, DeuSINEs, and Ceph-SINEs) have been reported so far, it is expected that new SINE superfamilies would be discovered by deep exploration of new SINEs in metazoan genomes. Here we describe 15 SINEs, among which 13 are novel, that have a similar 66-bp central region and therefore constitute a new SINE superfamily, MetaSINEs. MetaSINEs are distributed from fish to cnidarians, suggesting their common evolutionary origin at least 640 Ma. Because the 3' tails of MetaSINEs are variable, these SINEs most likely survived by changing their partner long interspersed elements for retrotransposition during evolution. Furthermore, we examined the presence of members of other SINE superfamilies in bivalve genomes and characterized eight new SINEs belonging to the CORE-SINEs, V-SINEs, and DeuSINEs, in addition to the MetaSINEs. The broad distribution of bivalve SINEs suggests that at least three SINEs originated in the common ancestor of Bivalvia. Our comparative analysis of the central domains of the SINEs revealed that, in each superfamily, only a restricted region is shared among all of its members. Because the functions of the central domains of the SINE superfamilies remain unknown, such structural information of SINE superfamilies will be useful for future experimental and comparative analyses to reveal why they have been retained in metazoan genomes during evolution. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  20. The aldo-keto reductase superfamily homepage.

    PubMed

    Hyndman, David; Bauman, David R; Heredia, Vladi V; Penning, Trevor M

    2003-02-01

    The aldo-keto reductases (AKRs) are one of the three enzyme superfamilies that perform oxidoreduction on a wide variety of natural and foreign substrates. A systematic nomenclature for the AKR superfamily was adopted in 1996 and was updated in September 2000 (visit www.med.upenn.edu/akr). Investigators have been diligent in submitting sequences of functional proteins to the Web site. With the new additions, the superfamily contains 114 proteins expressed in prokaryotes and eukaryotes that are distributed over 14 families (AKR1-AKR14). The AKR1 family contains the aldose reductases, the aldehyde reductases, the hydroxysteroid dehydrogenases and steroid 5beta-reductases, and is the largest. Other families of interest include AKR6, which includes potassium channel beta-subunits, and AKR7 the aflatoxin aldehyde reductases. Two new families include AKR13 (yeast aldose reductase) and AKR14 (Escherichia coli aldehyde reductase). Crystal structures of many AKRs and their complexes with ligands are available in the PDB and accessible through the Web site. Each structure has the characteristic (alpha/beta)(8)-barrel motif of the superfamily, a conserved cofactor binding site and a catalytic tetrad, and variable loop structures that define substrate specificity. Although the majority of AKRs are monomeric proteins of about 320 amino acids in length, the AKR2, AKR6 and AKR7 family may form multimers. To expand the nomenclature to accommodate multimers, we recommend that the composition and stoichiometry be listed. For example, AKR7A1:AKR7A4 (1:3) would designate a tetramer of the composition indicated. The current nomenclature is recognized by the Human Genome Project (HUGO) and the Web site provides a link to genomic information including chromosomal localization, gene boundaries, human ESTs and SNPs and much more.

  1. Overexpression of an ABC transporter and mutations of GyrA, GyrB, and ParC in contributing to high-level ciprofloxacin resistance in Streptococcus suis type 2.

    PubMed

    Yao, Jie; Shang, Kexin; Huang, Jinhu; Ran, Wei; Kashif, Jam; Wang, Liping

    2014-04-01

    Streptococcus suis is a pathogen of zoonotic diseases. Moreover, the emergence of fluoro-quinolones (FQs) resistance in this pathogen has severe consequences for pigs and human health. In this study, the molecular mechanism of FQs resistance in S. suis type 2 (SS2) sensitive strains isolated from pigs was assessed after in vitro induction of resistance against the most frequently used FQs: ciprofloxacin, norfloxacin, and enrofloxacin. Proteome analysis, sequencing and real-time RT-PCR results strongly established an overexpression of an ABC transporter protein (other than SatAB) and topoisomerase mutations in GyrA (Ser81Arg), GyrB (Glu354Lys), and ParC (Ser79Phe) in contributing to high level ciprofloxacin resistance in SS2. Due to the overexpression of the ABC transporter, intracellular ciprofloxacin concentrations were significantly lower in the resistant strains than those of sensitive strains after 20, 35, and 60 min exposures to ciprofloxacin (p < 0.05). It was concluded that improper use of FQs is one of the main causes of the emergence of this zoonotic pathogen as a multiresistant organism against commonly used antibiotics. The existence of an efflux-like protein is an incentive to find new drug targets to avoid the spread of FQs-resistant S. suis isolates in pigs and the human population.

  2. Functional Expression of P-glycoprotein and Organic Anion Transporting Polypeptides at the Blood-Brain Barrier: Understanding Transport Mechanisms for Improved CNS Drug Delivery?

    PubMed

    Abdullahi, Wazir; Davis, Thomas P; Ronaldson, Patrick T

    2017-07-01

    Drug delivery to the central nervous system (CNS) is greatly limited by the blood-brain barrier (BBB). Physical and biochemical properties of the BBB have rendered treatment of CNS diseases, including those with a hypoxia/reoxygenation (H/R) component, extremely difficult. Targeting endogenous BBB transporters from the ATP-binding cassette (ABC) superfamily (i.e., P-glycoprotein (P-gp)) or from the solute carrier (SLC) family (i.e., organic anion transporting polypeptides (OATPs in humans; Oatps in rodents)) has been suggested as a strategy that can improve delivery of drugs to the brain. With respect to P-gp, direct pharmacological inhibition using small molecules or selective regulation by targeting intracellular signaling pathways has been explored. These approaches have been largely unsuccessful due to toxicity issues and unpredictable pharmacokinetics. Therefore, our laboratory has proposed that optimization of CNS drug delivery, particularly for treatment of diseases with an H/R component, can be achieved by targeting Oatp isoforms at the BBB. As the major drug transporting Oatp isoform, Oatp1a4 has demonstrated blood-to-brain transport of substrate drugs with neuroprotective properties. Furthermore, our laboratory has shown that targeting Oatp1a4 regulation (i.e., TGF-β signaling mediated via the ALK-1 and ALK-5 transmembrane receptors) represents an opportunity to control Oatp1a4 functional expression for the purpose of delivering therapeutics to the CNS. In this review, we will discuss limitations of targeting P-gp-mediated transport activity and the advantages of targeting Oatp-mediated transport. Through this discussion, we will also provide critical information on novel approaches to improve CNS drug delivery by targeting endogenous uptake transporters expressed at the BBB.

  3. Loss of plastoglobule kinases ABC1K1 and ABC1K3 causes conditional degreening, modified prenyl-lipids, and recruitment of the jasmonic acid pathway

    USDA-ARS?s Scientific Manuscript database

    Plastoglobules (PGs) are plastid lipid-protein particles. This study examines the function of PG-localized kinases ABC1K1 and ABC1K3 in Arabidopsis thaliana. Several lines of evidence suggested that ABC1K1 and ABC1K3 form a protein complex. Null mutants for both genes (abc1k1 and abc1k3) and the dou...

  4. A Mutation within the Extended X Loop Abolished Substrate-induced ATPase Activity of the Human Liver ATP-binding Cassette (ABC) Transporter MDR3*

    PubMed Central

    Kluth, Marianne; Stindt, Jan; Dröge, Carola; Linnemann, Doris; Kubitz, Ralf; Schmitt, Lutz

    2015-01-01

    The human multidrug resistance protein 3 (MDR3/ABCB4) belongs to the ubiquitous family of ATP-binding cassette (ABC) transporters and is located in the canalicular membrane of hepatocytes. There it flops the phospholipids of the phosphatidylcholine (PC) family from the inner to the outer leaflet. Here, we report the characterization of wild type MDR3 and the Q1174E mutant, which was identified previously in a patient with progressive familial intrahepatic cholestasis type 3 (PFIC-3). We expressed different variants of MDR3 in the yeast Pichia pastoris, purified the proteins via tandem affinity chromatography, and determined MDR3-specific ATPase activity in the presence or absence of phospholipids. The ATPase activity of wild type MDR3 was stimulated 2-fold by liver PC or 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine lipids. Furthermore, the cross-linking of MDR3 with a thiol-reactive fluorophore blocked ATP hydrolysis and exhibited no PC stimulation. Similarly, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin lipids did not induce an increase of wild type MDR3 ATPase activity. The phosphate analogues beryllium fluoride and aluminum fluoride led to complete inhibition of ATPase activity, whereas orthovanadate inhibited exclusively the PC-stimulated ATPase activity of MDR3. The Q1174E mutation is located in the nucleotide-binding domain in direct proximity of the leucine of the ABC signature motif and extended the X loop, which is found in ABC exporters. Our data on the Q1174E mutant demonstrated basal ATPase activity, but PC lipids were incapable of stimulating ATPase activity highlighting the role of the extended X loop in the cross-talk of the nucleotide-binding domain and the transmembrane domain. PMID:25533467

  5. The trehalose-specific transporter LpqY-SugABC is required for antimicrobial and anti-biofilm activity of trehalose analogues in Mycobacterium smegmatis.

    PubMed

    Wolber, Jeffrey M; Urbanek, Bailey L; Meints, Lisa M; Piligian, Brent F; Lopez-Casillas, Irene C; Zochowski, Kailey M; Woodruff, Peter J; Swarts, Benjamin M

    2017-10-10

    Mycobacteria, including the bacterial pathogen that causes human tuberculosis, possess distinctive pathways for synthesizing and utilizing the non-mammalian disaccharide trehalose. Trehalose metabolism is essential for mycobacterial viability and has been linked to in vitro biofilm formation, which may bear relevance to in vivo drug tolerance. Previous research has shown that some trehalose analogues bearing modifications at the 6-position inhibit growth of various mycobacterial species. In this work, 2-, 5-, and 6-position-modified trehalose analogues were synthesized using our previously reported one-step chemoenzymatic method and shown to inhibit growth and biofilm formation in the two-to three-digit micromolar range in Mycobacterium smegmatis. The trehalose-specific ABC transporter LpqY-SugABC was essential for antimicrobial and anti-biofilm activity, suggesting that inhibition by monosubstituted trehalose analogues requires cellular uptake and does not proceed via direct action on extracellular targets such as antigen 85 acyltransferases or trehalose dimycolate hydrolase. Although the potency of the described compounds in in vitro growth and biofilm assays is moderate, this study reports the first trehalose-based mycobacterial biofilm inhibitors and reinforces the concept of exploiting unique sugar uptake pathways to deliver inhibitors and other chemical cargo to mycobacteria. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Nitrate transporter genes in apple and the effect of water deficit on their expression

    USDA-ARS?s Scientific Manuscript database

    Nitrogen transporters are members of a large superfamily, the Major Facilitator Superfamily (MFS). This family is ubiquitous and diverse, and includes proteins that facilitate the transport of a wide range of substrates across the cytoplasmic or intracellular membranes. Among the proteins encoded ...

  7. 4th International Consensus Conference on Advanced Breast Cancer (ABC4), Lisbon, November 4, 2017 : ABC4 Consensus: Assessment by a Panel of German Experts.

    PubMed

    Untch, Michael; Würstlein, Rachel; Marschner, Norbert; Lüftner, Diana; Augustin, Doris; Briest, Susanne; Ettl, Johannes; Haidinger, Renate; Müller, Lothar; Müller, Volkmar; Ruckhäberle, Eugen; Harbeck, Nadia; Thomssen, Christoph

    2018-05-01

    The fourth international advanced breast cancer consensus conference (ABC4) on the diagnosis and treatment of advanced breast cancer (ABC) headed by Professor Fatima Cardoso was once again held in Lisbon on November 2 - 4, 2017. To simplify matters, the abbreviation ABC will be used hereinafter in the text. In clinical practice, the abbreviation corresponds to metastatic breast cancer or locally far-advanced disease. This year the focus was on new developments in the treatment of ABC. Topics discussed included the importance of CDK4/6 inhibition in hormone receptor (HR)-positive ABC, the use of dual antibody blockade to treat HER2-positive ABC, PARP inhibition in triple-negative ABC and the potential therapeutic outcomes. Another major area discussed at the conference was BRCA-associated breast cancer, the treatment of cerebral metastasis, and individualized treatment decisions based on molecular testing (so-called precision medicine). As in previous years, close cooperation with representatives from patient organizations from around the world is an important aspect of the ABC conference. This cooperation was reinforced and expanded at the ABC4 conference. A global alliance was founded at the conclusion of the consensus conference, which aims to promote and coordinate the measures considered necessary by patient advocates worldwide. Because the panel of experts was composed of specialists from all over the world, it was inevitable that the ABC consensus also reflected country-specific features. As in previous years, a team of German breast cancer specialists who closely followed the consensus voting of the ABC panelists in Lisbon and intensively discussed the votes has therefore commented on the consensus in the context of the current German guidelines on the diagnosis and treatment of breast cancer 1 ,  2 used in clinical practice in Germany. The ABC consensus is based on the votes of the ABC panelists in Lisbon.

  8. eUnaG: a new ligand-inducible fluorescent reporter to detect drug transporter activity in live cells

    PubMed Central

    Yeh, Johannes T.-H.; Nam, Kwangho; Yeh, Joshua T.-H.; Perrimon, Norbert

    2017-01-01

    The absorption, distribution, metabolism and excretion (ADME) of metabolites and toxic organic solutes are orchestrated by the ATP-binding cassette (ABC) transporters and the organic solute carrier family (SLC) proteins. A large number of ABC and SLC transpoters exist; however, only a small number have been well characterized. To facilitate the analysis of these transporters, which is important for drug safety and physiological studies, we developed a sensitive genetically encoded bilirubin (BR)-inducible fluorescence sensor (eUnaG) to detect transporter-coupled influx/efflux of organic compounds. This sensor can be used in live cells to measure transporter activity, as excretion of BR depends on ABC and SLC transporters. Applying eUnaG in functional RNAi screens, we characterize l(2)03659 as a Drosophila multidrug resistant-associated ABC transporter. PMID:28176814

  9. The minimal-ABC trees with B1-branches.

    PubMed

    Dimitrov, Darko; Du, Zhibin; Fonseca, Carlos M da

    2018-01-01

    The atom-bond connectivity index (or, for short, ABC index) is a molecular structure descriptor bridging chemistry to graph theory. It is probably the most studied topological index among all numerical parameters of a graph that characterize its topology. For a given graph G = (V, E), the ABC index of G is defined as [Formula: see text], where di denotes the degree of the vertex i, and ij is the edge incident to the vertices i and j. A combination of physicochemical and the ABC index properties are commonly used to foresee the bioactivity of different chemical composites. Additionally, the applicability of the ABC index in chemical thermodynamics and other areas of chemistry, such as in dendrimer nanostars, benzenoid systems, fluoranthene congeners, and phenylenes is well studied in the literature. While finding of the graphs with the greatest ABC-value is a straightforward assignment, the characterization of the tree(s) with minimal ABC index is a problem largely open and has recently given rise to numerous studies and conjectures. A B1-branch of a graph is a pendent path of order 2. In this paper, we provide an important step forward to the full characterization of these minimal trees. Namely, we show that a minimal-ABC tree contains neither 4 nor 3 B1-branches. The case when the number of B1-branches is 2 is also considered.

  10. Yang-Mills theory and the ABC conjecture

    NASA Astrophysics Data System (ADS)

    He, Yang-Hui; Hu, Zhi; Probst, Malte; Read, James

    2018-05-01

    We establish a precise correspondence between the ABC Conjecture and 𝒩 = 4 super-Yang-Mills theory. This is achieved by combining three ingredients: (i) Elkies’ method of mapping ABC-triples to elliptic curves in his demonstration that ABC implies Mordell/Faltings; (ii) an explicit pair of elliptic curve and associated Belyi map given by Khadjavi-Scharaschkin; and (iii) the fact that the bipartite brane-tiling/dimer model for a gauge theory with toric moduli space is a particular dessin d’enfant in the sense of Grothendieck. We explore this correspondence for the highest quality ABC-triples as well as large samples of random triples. The conjecture itself is mapped to a statement about the fundamental domain of the toroidal compactification of the string realization of 𝒩 = 4 SYM.

  11. Low levels of graphene and graphene oxide inhibit cellular xenobiotic defense system mediated by efflux transporters.

    PubMed

    Liu, Su; Jiang, Wei; Wu, Bing; Yu, Jing; Yu, Haiyan; Zhang, Xu-Xiang; Torres-Duarte, Cristina; Cherr, Gary N

    2016-01-01

    Low levels of graphene and graphene oxide (GO) are considered to be environmentally safe. In this study, we analyzed the potential effects of graphene and GO at relatively low concentrations on cellular xenobiotic defense system mediated by efflux transporters. The results showed that graphene (<0.5 μg/mL) and GO (<20 μg/mL) did not decrease cell viability, generate reactive oxygen species, or disrupt mitochondrial function. However, graphene and GO at the nontoxic concentrations could increase calcein-AM (CAM, an indicator of membrane ATP-binding cassette (ABC) transporter) activity) accumulation, indicating inhibition of ABC transporters' efflux capabilities. This inhibition was observed even at 0.005 μg/mL graphene and 0.05 μg/mL GO, which are 100 times and 400 times lower than their lowest toxic concentration from cytotoxicity experiments, respectively. The inhibition of ABC transporters significantly increased the toxicity of paraquat and arsenic, known substrates of ABC transporters. The inhibition of ABC transporters was found to be based on graphene and GO damaging the plasma membrane structure and fluidity, thus altering functions of transmembrane ABC transporters. This study demonstrates that low levels of graphene and GO are not environmentally safe since they can significantly make cell more susceptible to other xenobiotics, and this chemosensitizing activity should be considered in the risk assessment of graphene and GO.

  12. A mutation within the extended X loop abolished substrate-induced ATPase activity of the human liver ATP-binding cassette (ABC) transporter MDR3.

    PubMed

    Kluth, Marianne; Stindt, Jan; Dröge, Carola; Linnemann, Doris; Kubitz, Ralf; Schmitt, Lutz

    2015-02-20

    The human multidrug resistance protein 3 (MDR3/ABCB4) belongs to the ubiquitous family of ATP-binding cassette (ABC) transporters and is located in the canalicular membrane of hepatocytes. There it flops the phospholipids of the phosphatidylcholine (PC) family from the inner to the outer leaflet. Here, we report the characterization of wild type MDR3 and the Q1174E mutant, which was identified previously in a patient with progressive familial intrahepatic cholestasis type 3 (PFIC-3). We expressed different variants of MDR3 in the yeast Pichia pastoris, purified the proteins via tandem affinity chromatography, and determined MDR3-specific ATPase activity in the presence or absence of phospholipids. The ATPase activity of wild type MDR3 was stimulated 2-fold by liver PC or 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine lipids. Furthermore, the cross-linking of MDR3 with a thiol-reactive fluorophore blocked ATP hydrolysis and exhibited no PC stimulation. Similarly, phosphatidylethanolamine, phosphatidylserine, and sphingomyelin lipids did not induce an increase of wild type MDR3 ATPase activity. The phosphate analogues beryllium fluoride and aluminum fluoride led to complete inhibition of ATPase activity, whereas orthovanadate inhibited exclusively the PC-stimulated ATPase activity of MDR3. The Q1174E mutation is located in the nucleotide-binding domain in direct proximity of the leucine of the ABC signature motif and extended the X loop, which is found in ABC exporters. Our data on the Q1174E mutant demonstrated basal ATPase activity, but PC lipids were incapable of stimulating ATPase activity highlighting the role of the extended X loop in the cross-talk of the nucleotide-binding domain and the transmembrane domain. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. ATP-binding cassette exporters: structure and mechanism with a focus on P-glycoprotein and MRP1.

    PubMed

    Arana, Maite Rocío; Altenberg, Guillermo

    2017-10-12

    The majority of proteins that belong to the ATP-binding cassette (ABC) superfamily are transporters that mediate the efflux of substrates from cells. These exporters include multidrug resistance proteins of the ABCB and ABCC subfamilies, such as P-glycoprotein (Pgp) and MRP1, respectively. These proteins are not only involved in the resistance of cancer to cytotoxic agents, but also in the protection from endo and xenobiotics, and the determination of drug pharmacokinetics, as well as in the pathophysiology of a variety of disorders. Here, we present a review of the information available on ABC exporters, with a focus on Pgp, MRP1 and related proteins. We describe tissue localization and function of these transporters in health and disease, and discuss the mechanisms of substrate transport. We also correlate recent structural information with the function of the exporters, and discuss details of their molecular mechanism with a focus on the nucleotide-binding domains. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  14. Hijacking membrane transporters for arsenic phytoextraction

    PubMed Central

    LeBlanc, Melissa S.; McKinney, Elizabeth C.; Meagher, Richard B.; Smith, Aaron P.

    2012-01-01

    Arsenic is a toxic metalloid and recognized carcinogen. Arsenate and arsenite are the most common arsenic species available for uptake by plants. As an inorganic phosphate (Pi) analog, arsenate is acquired by plant roots through endogenous Pi transport systems. Inside the cell, arsenate is reduced to the thiol-reactive form arsenite. Glutathione (GSH)-conjugates of arsenite may be extruded from the cell or sequestered in vacuoles by members of the ATP-binding cassette (ABC) family of transporters. In the present study we sought to enhance both plant arsenic uptake through Pi transporter overexpression, and plant arsenic tolerance through ABC transporter overexpression. We demonstrate that Arabidopsis thaliana plants overexpressing the high-affinity Pi transporter family members, AtPht1;1 or AtPht1;7, are hypersensitive to arsenate due to increased arsenate uptake. These plants do not exhibit increased sensitivity to arsenite. Co-overexpression of the yeast ABC transporter YCF1 in combination with AtPht1;1 or AtPht1;7 suppresses the arsenate-sensitive phenotype while further enhancing arsenic uptake. Taken together, our results support an arsenic transport mechanism in which arsenate uptake is increased through Pi transporter overexpression, and arsenic tolerance is enhanced through YCF1-mediated vacuolar sequestration. This work substantiates the viability of coupling enhanced uptake and vacuolar sequestration as a means for developing a prototypical engineered arsenic hyperaccumulator. PMID:23108027

  15. High-Affinity Vanadate Transport System in the Cyanobacterium Anabaena variabilis ATCC 29413

    PubMed Central

    Pratte, Brenda S.; Thiel, Teresa

    2006-01-01

    High-affinity vanadate transport systems have not heretofore been identified in any organism. Anabaena variabilis, which can fix nitrogen by using an alternative V-dependent nitrogenase, transported vanadate well. The concentration of vanadate giving half-maximum V-nitrogenase activity when added to V-starved cells was about 3 × 10−9 M. The genes for an ABC-type vanadate transport system, vupABC, were found in A. variabilis about 5 kb from the major cluster of genes encoding the V-nitrogenase, and like those genes, the vupABC genes were repressed by molybdate; however, unlike the V-nitrogenase genes the vanadate transport genes were expressed in vegetative cells. A vupB mutant failed to grow by using V-nitrogenase unless high levels of vanadate were provided, suggesting that there was also a low-affinity vanadate transport system that functioned in the vupB mutant. The vupABC genes belong to a family of putative metal transport genes that include only one other characterized transport system, the tungstate transport genes of Eubacterium acidaminophilum. Similar genes are not present in the complete genomes of other bacterial strains that have a V-nitrogenase, including Azotobacter vinelandii, Rhodopseudomonas palustris, and Methanosarcina barkeri. PMID:16385036

  16. Multiple ABC transporters are involved in the acquisition of petrobactin in Bacillus anthracis

    PubMed Central

    Dixon, Shandee D.; Janes, Brian K.; Bourgis, Alexandra; Carlson, Paul E.; Hanna, Philip C.

    2012-01-01

    Summary In Bacillus anthracis the siderophore petrobactin is vital for iron acquisition and virulence. The petrobactin-binding receptor FpuA is required for these processes. Here additional components of petrobactin reacquisition are described. To identify these proteins, mutants of candidate permease and ATPase genes were generated allowing for characterization of multiple petrobactin ATP-binding cassette (ABC)-import systems. Either of two distinct permeases, FpuB or FatCD, are required for iron acquisition and play redundant roles in petrobactin transport. A mutant strain lacking both permeases, ΔfpuBΔfatCD, was incapable of using petrobactin as an iron source and exhibited attenuated virulence in a murine model of inhalational anthrax infection. ATPase mutants were generated in either of the permease mutant backgrounds to identify the ATPase(s) interacting with each individual permease channel. Mutants lacking the FpuB permease and FatE ATPase (ΔfpuBΔfatE) and a mutant lacking the distinct ATPases FpuC and FpuD generated in the ΔfatCD background (ΔfatCDΔfpuCΔfpuD) displayed phenotypic characteristics of a mutant deficient in petrobactin import. A mutant lacking all three of the identified ATPases (ΔfatEΔfpuCΔfpuD) exhibited the same growth defect in iron-depleted conditions. Taken together, these results provide the first description of the permease and ATPase proteins required for the import of petrobactin in B. anthracis. PMID:22429808

  17. ABC1 Consensus Conference - a German Perspective: First International Consensus Conference on Advanced Breast Cancer (ABC1), Lisbon, November 5, 2011.

    PubMed

    Thomssen, Christoph; Marschner, Norbert; Untch, Michael; Decker, Thomas; Hegewisch-Becker, Susanna; Jackisch, Christian; Janni, Wolfgang; Hans-Joachim, Lück; von Minckwitz, Gunter; Scharl, Anton; Schneeweiss, Andreas; Tesch, Hans; Welt, Anja; Harbeck, Nadia

    2012-02-01

    A group of German breast cancer experts (medical oncologists and gynaecologists) reviewed and commented on the results of the first international 'Advanced Breast Cancer First Consensus Conference' (ABC1) for the diagnosis and treatment of advanced breast cancer. The ABC1 Conference is an initiative of the European School of Oncology (ESO) Metastatic Breast Cancer Task Force in cooperation with the EBCC (European Breast Cancer Conference), ESMO (European Society of Medical Oncology) and the American JNCI (Journal of the National Cancer Institute). The main focus of the ABC1 Conference was metastatic breast cancer (stage IV). The ABC1 consensus is based on the vote of 33 breast cancer experts from different countries and has been specified as a guideline for therapeutic practice by the German expert group. It is the objective of the ABC1 consensus as well as of the German comments to provide an internationally standardized and evidence-based foundation for qualified decision-making in the treatment of metastatic breast cancer.

  18. Novel actin crosslinker superfamily member identified by a two step degenerate PCR procedure.

    PubMed

    Byers, T J; Beggs, A H; McNally, E M; Kunkel, L M

    1995-07-24

    Actin-crosslinking proteins link F-actin into the bundles and networks that constitute the cytoskeleton. Dystrophin, beta-spectrin, alpha-actinin, ABP-120, ABP-280, and fimbrin share homologous actin-binding domains and comprise an actin crosslinker superfamily. We have identified a novel member of this superfamily (ACF7) using a degenerate primer-mediated PCR strategy that was optimized to resolve less-abundant superfamily sequences. The ACF7 gene is on human chromosome 1 and hybridizes to high molecular weight bands on northern blots. Sequence comparisons argue that ACF7 does not fit into one of the existing families, but represents a new class within the superfamily.

  19. ABCE1 Is a Highly Conserved RNA Silencing Suppressor

    PubMed Central

    Kärblane, Kairi; Gerassimenko, Jelena; Nigul, Lenne; Piirsoo, Alla; Smialowska, Agata; Vinkel, Kadri; Kylsten, Per; Ekwall, Karl; Swoboda, Peter; Truve, Erkki; Sarmiento, Cecilia

    2015-01-01

    ATP-binding cassette sub-family E member 1 (ABCE1) is a highly conserved protein among eukaryotes and archaea. Recent studies have identified ABCE1 as a ribosome-recycling factor important for translation termination in mammalian cells, yeast and also archaea. Here we report another conserved function of ABCE1. We have previously described AtRLI2, the homolog of ABCE1 in the plant Arabidopsis thaliana, as an endogenous suppressor of RNA silencing. In this study we show that this function is conserved: human ABCE1 is able to suppress RNA silencing in Nicotiana benthamiana plants, in mammalian HEK293 cells and in the worm Caenorhabditis elegans. Using co-immunoprecipitation and mass spectrometry, we found a number of potential ABCE1-interacting proteins that might support its function as an endogenous suppressor of RNA interference. The interactor candidates are associated with epigenetic regulation, transcription, RNA processing and mRNA surveillance. In addition, one of the identified proteins is translin, which together with its binding partner TRAX supports RNA interference. PMID:25659154

  20. Dimeric isoxazolyl-1,4-dihydropyridines have enhanced binding at the multi-drug resistance transporter.

    PubMed

    Steiger, Scott A; Li, Chun; Backos, Donald S; Reigan, Philip; Natale, N R

    2017-06-15

    A series of dimeric isoxazolyl-1,4-dihydropyridines (IDHPs) were prepared by click chemistry and examined for their ability to bind the multi-drug resistance transporter (MDR-1), a member of the ATP-binding cassette superfamily (ABC). Eight compounds in the present study exhibited single digit micromolar binding to this efflux transporter. One monomeric IDHP m-Br-1c, possessed submicromolar binding of 510nM at MDR-1. Three of the dimeric IDHPs possessed <1.5µM activity, and 4b and 4c were observed to have superior binding selectivity compared to their corresponding monomers verses the voltage gated calcium channel (VGCC). The dimer with the best combination of activity and selectivity for MDR-1 was analog 4c containing an m-Br phenyl moiety in the 3-position of the isoxazole, and a tether with five ethyleneoxy units, referred to herein as Isoxaquidar. Two important controls, mono-triazole 5 and pyridine 6, also were examined, indicating that the triazole - incorporated as part of the click assembly as a spacer - contributes to MDR-1 binding. Compounds were also assayed at the allosteric site of the mGluR5 receptor, as a GPCR 7TM control, indicating that the p-Br IDHPs 4d, 4e and 4f with tethers of from n=2 to 5 ethylenedioxy units, had sub-micromolar affinities with 4d being the most efficacious at 193nM at mGluR5. The results are interpreted using a docking study using a human ABC as our current working hypothesis, and suggest that the distinct SARs emerging for these three divergent classes of biomolecular targets may be tunable, and amenable to the development of further selectivity. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. ATP-binding cassette transporters in tumor endothelial cells and resistance to metronomic chemotherapy.

    PubMed

    Hida, Kyoko; Kikuchi, Hiroshi; Maishi, Nako; Hida, Yasuhiro

    2017-08-01

    Drug resistance is a major problem in anticancer therapy. ATP-binding cassette (ABC) transporters have a role in the multidrug resistance. A new regimen of chemotherapy has been proposed, called "metronomic chemotherapy". Metronomic chemotherapy is the frequent, regular administration of drug doses designed to maintain low, but active, concentrations of chemotherapeutic drugs over prolonged periods of time, without causing serious toxicities. Metronomic chemotherapy regimens were developed to optimize the antitumor efficacy of agents that target the tumor vasculature instead of tumor cells, and to reduce toxicity of antineoplastic drugs [1]. Nevertheless, recent studies revealed that ABC transporters are expressed at a higher level in the endothelium in the tumor. To avoid resistance to metronomic anti-angiogenic chemotherapy, ABC transporter inhibition of tumor endothelial cells may be a promising strategy. In this mini-review, we discuss the possible mechanism of resistance to metronomic chemotherapy from the viewpoint of tumor endothelial cell biology, focusing on ABC transporters. Copyright © 2017. Published by Elsevier B.V.

  2. Functions of the poly(ADP-ribose) polymerase superfamily in plants.

    PubMed

    Lamb, Rebecca S; Citarelli, Matteo; Teotia, Sachin

    2012-01-01

    Poly(ADP-ribosyl)ation is the covalent attachment of ADP-ribose subunits from NAD(+) to target proteins and was first described in plants in the 1970s. This post-translational modification is mediated by poly(ADP-ribose) polymerases (PARPs) and removed by poly(ADP-ribose) glycohydrolases (PARGs). PARPs have important functions in many biological processes including DNA repair, epigenetic regulation and transcription. However, these roles are not always associated with enzymatic activity. The PARP superfamily has been well studied in animals, but remains under-investigated in plants. Although plants lack the variety of PARP superfamily members found in mammals, they do encode three different types of PARP superfamily proteins, including a group of PARP-like proteins, the SRO family, that are plant specific. In plants, members of the PARP family and/or poly(ADP-ribosyl)ation have been linked to DNA repair, mitosis, innate immunity and stress responses. In addition, members of the SRO family have been shown to be necessary for normal sporophytic development. In this review, we summarize the current state of plant research into poly(ADP-ribosyl)ation and the PARP superfamily in plants.

  3. Arabidopsis thaliana NIP7;1: An Anther-Specific Boric Acid Transporter of the Aquaporin Superfamily Regulated by an Unusual Tyrosine in Helix 2 of the Transport Pore

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Li, Tian; Choi, Won-Gyu; Baudry, Jerome Y

    Plant nodulin-26 intrinsic proteins (NIPs) are members of the aquaporin superfamily that serve as multifunctional transporters of uncharged metabolites. In Arabidopsis thaliana, a specific NIP pore subclass, known as the NIP II proteins, is represented by AtNIP5;1 and AtNIP6;1, which encode channel proteins expressed in roots and leaf nodes, respectively, that participate in the transport of the critical cell wall nutrient boric acid. Modeling of the protein encoded by the AtNIP7;1 gene shows that it is a third member of the NIP II pore subclass in Arabidopsis. However, unlike AtNIP5;1 and AtNIP6;1 proteins, which form constitutive boric acid channels, AtNIP7;1more » forms a channel with an extremely low intrinsic boric acid transport activity. Molecular modeling and molecular dynamics simulations of AtNIP7;1 suggest that a conserved tyrosine residue (Tyr81) located in transmembrane helix 2 adjacent to the aromatic arginine (ar/R) pore selectivity region stabilizes a closed pore conformation through interaction with the canonical Arg220 in ar/R region. Substitution of Tyr81 with a Cys residue, characteristic of established NIP boric acid channels, results in opening of the AtNIP7;1 pore that acquires a robust, transport activity for boric acid as well as other NIP II test solutes (glycerol and urea). Substitution of a Phe for Tyr81 also opens the channel, supporting the prediction from MD simulations that hydrogen bond interaction between the Tyr81 phenol group and the ar/R Arg may contribute to the stabilization of a closed pore state. Expression analyses show that AtNIP7;1 is selectively expressed in developing anther tissues of young floral buds of A. thaliana, principally in developing pollen grains of stage 9 11 anthers. Because boric acid is both an essential nutrient as well as a toxic compound at high concentrations, it is proposed that Tyr81 modulates transport and may provide an additional level of regulation for this transporter in male gametophyte

  4. Arabidopsis thaliana NIP7;1: an anther-specific boric acid transporter of the aquaporin superfamily regulated by an unusual tyrosine in helix 2 of the transport pore.

    PubMed

    Li, Tian; Choi, Won-Gyu; Wallace, Ian S; Baudry, Jerome; Roberts, Daniel M

    2011-08-09

    Plant nodulin-26 intrinsic proteins (NIPs) are members of the aquaporin superfamily that serve as multifunctional transporters of uncharged metabolites. In Arabidopsis thaliana, a specific NIP pore subclass, known as the NIP II proteins, is represented by AtNIP5;1 and AtNIP6;1, which encode channel proteins expressed in roots and leaf nodes, respectively, that participate in the transport of the critical cell wall nutrient boric acid. Modeling of the protein encoded by the AtNIP7;1 gene shows that it is a third member of the NIP II pore subclass in Arabidopsis. However, unlike AtNIP5;1 and AtNIP6;1 proteins, which form constitutive boric acid channels, AtNIP7;1 forms a channel with an extremely low intrinsic boric acid transport activity. Molecular modeling and molecular dynamics simulations of AtNIP7;1 suggest that a conserved tyrosine residue (Tyr81) located in transmembrane helix 2 adjacent to the aromatic arginine (ar/R) pore selectivity region stabilizes a closed pore conformation through interaction with the canonical Arg220 in ar/R region. Substitution of Tyr81 with a Cys residue, characteristic of established NIP boric acid channels, results in opening of the AtNIP7;1 pore that acquires a robust, transport activity for boric acid as well as other NIP II test solutes (glycerol and urea). Substitution of a Phe for Tyr81 also opens the channel, supporting the prediction from MD simulations that hydrogen bond interaction between the Tyr81 phenol group and the ar/R Arg may contribute to the stabilization of a closed pore state. Expression analyses show that AtNIP7;1 is selectively expressed in developing anther tissues of young floral buds of A. thaliana, principally in developing pollen grains of stage 9-11 anthers. Because boric acid is both an essential nutrient as well as a toxic compound at high concentrations, it is proposed that Tyr81 modulates transport and may provide an additional level of regulation for this transporter in male gametophyte development

  5. Down-regulation of a novel ABC transporter gene (Pxwhite) is associated with Cry1Ac resistance in the diamondback moth, Plutella xylostella (L.).

    PubMed

    Guo, Zhaojiang; Kang, Shi; Zhu, Xun; Xia, Jixing; Wu, Qingjun; Wang, Shaoli; Xie, Wen; Zhang, Youjun

    2015-04-01

    Biopesticides or transgenic crops based on Cry toxins from the soil bacterium Bacillus thuringiensis (Bt) effectively control agricultural insect pests. The sustainable use of Bt biopesticides and Bt crops is threatened, however, by the development of Cry resistance in the target pests. The diamondback moth, Plutella xylostella (L.), is the first pest that developed resistance to a Bt biopesticide in the field, and a recent study has shown that the resistance of P. xylostella to Cry1Ac is caused by a mutation in an ATP-binding cassette (ABC) transporter gene (ABCC2). In this study, we report that down-regulation of a novel ABC transporter gene from ABCG subfamily (Pxwhite) is associated with Cry1Ac resistance in P. xylostella. The full-length cDNA sequence of Pxwhite was cloned and analyzed. Spatial-temporal expression detection revealed that Pxwhite was expressed in all tissues and developmental stages, and highest expressed in Malpighian tubule tissue and in egg stage. Sequence variation analysis of Pxwhite indicated the absence of constant non-synonymous mutations between susceptible and resistant strains, whereas midgut transcript analysis showed that Pxwhite was remarkably reduced in all resistant strains and further reduced when larvae of the moderately resistant SZ-R strain were subjected to selection with Cry1Ac toxin. Furthermore, RNA interference (RNAi)-mediated suppression of Pxwhite gene expression significantly reduced larval susceptibility to Cry1Ac toxin, and genetic linkage analysis confirmed that down-regulation of Pxwhite gene is tightly linked to Cry1Ac resistance in P. xylostella. To our knowledge, this is the first report indicating that Pxwhite gene is involved in Cry1Ac resistance in P. xylostella. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Computer simulations of transport through membranes: passive diffusion, pores, channels and transporters.

    PubMed

    Tieleman, D Peter

    2006-10-01

    A key function of biological membranes is to provide mechanisms for the controlled transport of ions, nutrients, metabolites, peptides and proteins between a cell and its environment. We are using computer simulations to study several processes involved in transport. In model membranes, the distribution of small molecules can be accurately calculated; we are making progress towards understanding the factors that determine the partitioning behaviour in the inhomogeneous lipid environment, with implications for drug distribution, membrane protein folding and the energetics of voltage gating. Lipid bilayers can be simulated at a scale that is sufficiently large to study significant defects, such as those caused by electroporation. Computer simulations of complex membrane proteins, such as potassium channels and ATP-binding cassette (ABC) transporters, can give detailed information about the atomistic dynamics that form the basis of ion transport, selectivity, conformational change and the molecular mechanism of ATP-driven transport. This is illustrated in the present review with recent simulation studies of the voltage-gated potassium channel KvAP and the ABC transporter BtuCD.

  7. Characterization of the intronic portion of cadherin superfamily members, common cancer orchestrators

    PubMed Central

    Oliveira, Patrícia; Sanges, Remo; Huntsman, David; Stupka, Elia; Oliveira, Carla

    2012-01-01

    Cadherins are cell–cell adhesion proteins essential for the maintenance of tissue architecture and integrity, and their impairment is often associated with human cancer. Knowledge regarding regulatory mechanisms associated with cadherin misexpression in cancer is scarce. Specific features of the intronic-structure and intronic-based regulatory mechanisms in the cadherin superfamily are unidentified. This study aims at systematically characterizing the intronic portion of cadherin superfamily members and the identification of intronic regions constituting putative targets/triggers of regulation, using a bioinformatic approach and biological data mining. Our study demonstrates that the cadherin superfamily genes harbour specific characteristics in comparison to all non-cadherin genes, both from the genomic and transcriptional standpoints. Cadherin superfamily genes display higher average total intron number and significantly longer introns than other genes and across the entire vertebrate lineage. Moreover, in the human genome, we observed an uncommon high frequency of MIR (mammalian-wide interspersed repeats) and MaLR (mammalian-wide interspersed repeats, a subtype of LTR) regulatory-associated repetitive elements at 5′-located introns, concomitantly with increased de novo intronic transcription. Using this approach, we identified cadherin intronic-specific sites that may constitute novel targets/triggers of cadherin superfamily expression regulation. These findings pinpoint the need to identify mechanisms affecting particularly MIR and MaLR elements located in introns 2 and 3 of human cadherin genes, possibly important in the expression modulation of this superfamily in homeostasis and cancer. PMID:22317972

  8. Role of Conserved Glycine in Zinc-dependent Medium Chain Dehydrogenase/Reductase Superfamily*

    PubMed Central

    Tiwari, Manish Kumar; Singh, Raushan Kumar; Singh, Ranjitha; Jeya, Marimuthu; Zhao, Huimin; Lee, Jung-Kul

    2012-01-01

    The medium-chain dehydrogenase/reductase (MDR) superfamily consists of a large group of enzymes with a broad range of activities. Members of this superfamily are currently the subject of intensive investigation, but many aspects, including the zinc dependence of MDR superfamily proteins, have not yet have been adequately investigated. Using a density functional theory-based screening strategy, we have identified a strictly conserved glycine residue (Gly) in the zinc-dependent MDR superfamily. To elucidate the role of this conserved Gly in MDR, we carried out a comprehensive structural, functional, and computational analysis of four MDR enzymes through a series of studies including site-directed mutagenesis, isothermal titration calorimetry, electron paramagnetic resonance (EPR), quantum mechanics, and molecular mechanics analysis. Gly substitution by other amino acids posed a significant threat to the metal binding affinity and activity of MDR superfamily enzymes. Mutagenesis at the conserved Gly resulted in alterations in the coordination of the catalytic zinc ion, with concomitant changes in metal-ligand bond length, bond angle, and the affinity (Kd) toward the zinc ion. The Gly mutants also showed different spectroscopic properties in EPR compared with those of the wild type, indicating that the binding geometries of the zinc to the zinc binding ligands were changed by the mutation. The present results demonstrate that the conserved Gly in the GHE motif plays a role in maintaining the metal binding affinity and the electronic state of the catalytic zinc ion during catalysis of the MDR superfamily enzymes. PMID:22500022

  9. Phylogenomic evolutionary surveys of subtilase superfamily genes in fungi.

    PubMed

    Li, Juan; Gu, Fei; Wu, Runian; Yang, JinKui; Zhang, Ke-Qin

    2017-03-30

    Subtilases belong to a superfamily of serine proteases which are ubiquitous in fungi and are suspected to have developed distinct functional properties to help fungi adapt to different ecological niches. In this study, we conducted a large-scale phylogenomic survey of subtilase protease genes in 83 whole genome sequenced fungal species in order to identify the evolutionary patterns and subsequent functional divergences of different subtilase families among the main lineages of the fungal kingdom. Our comparative genomic analyses of the subtilase superfamily indicated that extensive gene duplications, losses and functional diversifications have occurred in fungi, and that the four families of subtilase enzymes in fungi, including proteinase K-like, Pyrolisin, kexin and S53, have distinct evolutionary histories which may have facilitated the adaptation of fungi to a broad array of life strategies. Our study provides new insights into the evolution of the subtilase superfamily in fungi and expands our understanding of the evolution of fungi with different lifestyles.

  10. Large-Scale Analysis Exploring Evolution of Catalytic Machineries and Mechanisms in Enzyme Superfamilies.

    PubMed

    Furnham, Nicholas; Dawson, Natalie L; Rahman, Syed A; Thornton, Janet M; Orengo, Christine A

    2016-01-29

    Enzymes, as biological catalysts, form the basis of all forms of life. How these proteins have evolved their functions remains a fundamental question in biology. Over 100 years of detailed biochemistry studies, combined with the large volumes of sequence and protein structural data now available, means that we are able to perform large-scale analyses to address this question. Using a range of computational tools and resources, we have compiled information on all experimentally annotated changes in enzyme function within 379 structurally defined protein domain superfamilies, linking the changes observed in functions during evolution to changes in reaction chemistry. Many superfamilies show changes in function at some level, although one function often dominates one superfamily. We use quantitative measures of changes in reaction chemistry to reveal the various types of chemical changes occurring during evolution and to exemplify these by detailed examples. Additionally, we use structural information of the enzymes active site to examine how different superfamilies have changed their catalytic machinery during evolution. Some superfamilies have changed the reactions they perform without changing catalytic machinery. In others, large changes of enzyme function, in terms of both overall chemistry and substrate specificity, have been brought about by significant changes in catalytic machinery. Interestingly, in some superfamilies, relatives perform similar functions but with different catalytic machineries. This analysis highlights characteristics of functional evolution across a wide range of superfamilies, providing insights that will be useful in predicting the function of uncharacterised sequences and the design of new synthetic enzymes. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  11. Comparative genomics of transport proteins in developmental bacteria: Myxococcus xanthus and Streptomyces coelicolor

    PubMed Central

    2013-01-01

    Background Two of the largest fully sequenced prokaryotic genomes are those of the actinobacterium, Streptomyces coelicolor (Sco), and the δ-proteobacterium, Myxococcus xanthus (Mxa), both differentiating, sporulating, antibiotic producing, soil microbes. Although the genomes of Sco and Mxa are the same size (~9 Mbp), Sco has 10% more genes that are on average 10% smaller than those in Mxa. Results Surprisingly, Sco has 93% more identifiable transport proteins than Mxa. This is because Sco has amplified several specific types of its transport protein genes, while Mxa has done so to a much lesser extent. Amplification is substrate- and family-specific. For example, Sco but not Mxa has amplified its voltage-gated ion channels but not its aquaporins and mechano-sensitive channels. Sco but not Mxa has also amplified drug efflux pumps of the DHA2 Family of the Major Facilitator Superfamily (MFS) (49 versus 6), amino acid transporters of the APC Family (17 versus 2), ABC-type sugar transport proteins (85 versus 6), and organic anion transporters of several families. Sco has not amplified most other types of transporters. Mxa has selectively amplified one family of macrolid exporters relative to Sco (16 versus 1), consistent with the observation that Mxa makes more macrolids than does Sco. Conclusions Except for electron transport carriers, there is a poor correlation between the types of transporters found in these two organisms, suggesting that their solutions to differentiative and metabolic needs evolved independently. A number of unexpected and surprising observations are presented, and predictions are made regarding the physiological functions of recognizable transporters as well as the existence of yet to be discovered transport systems in these two important model organisms and their relatives. The results provide insight into the evolutionary processes by which two dissimilar prokaryotes evolved complexity, particularly through selective chromosomal gene

  12. P-type ATPase superfamily: evidence for critical roles for kingdom evolution.

    PubMed

    Okamura, Hideyuki; Denawa, Masatsugu; Ohniwa, Ryosuke; Takeyasu, Kunio

    2003-04-01

    The P-type ATPase has become a protein superfamily. On the basis of sequence similarities, the phylogenetic analyses, and substrate specificities, this superfamily can be classified into 5 families and 11 subfamilies. A comparative phylogenetic analysis demonstrates the relationship between the molecular evolution of these subfamilies and the establishment of the kingdoms of living things.

  13. Heterologous expression of the yeast Tpo1p or Pdr5p membrane transporters in Arabidopsis confers plant xenobiotic tolerance.

    PubMed

    Remy, Estelle; Niño-González, María; Godinho, Cláudia P; Cabrito, Tânia R; Teixeira, Miguel C; Sá-Correia, Isabel; Duque, Paula

    2017-07-03

    Soil contamination is a major hindrance for plant growth and development. The lack of effective strategies to remove chemicals released into the environment has raised the need to increase plant resilience to soil pollutants. Here, we investigated the ability of two Saccharomyces cerevisiae plasma-membrane transporters, the Major Facilitator Superfamily (MFS) member Tpo1p and the ATP-Binding Cassette (ABC) protein Pdr5p, to confer Multiple Drug Resistance (MDR) in Arabidopsis thaliana. Transgenic plants expressing either of the yeast transporters were undistinguishable from the wild type under control conditions, but displayed tolerance when challenged with the herbicides 2,4-D and barban. Plants expressing ScTPO1 were also more resistant to the herbicides alachlor and metolachlor as well as to the fungicide mancozeb and the Co 2+ , Cu 2+ , Ni 2+ , Al 3+ and Cd 2+ cations, while ScPDR5-expressing plants exhibited tolerance to cycloheximide. Yeast mutants lacking Tpo1p or Pdr5p showed increased sensitivity to most of the agents tested in plants. Our results demonstrate that the S. cerevisiae Tpo1p and Pdr5p transporters are able to mediate resistance to a broad range of compounds of agricultural interest in yeast as well as in Arabidopsis, underscoring their potential in future biotechnological applications.

  14. PASS2: an automated database of protein alignments organised as structural superfamilies.

    PubMed

    Bhaduri, Anirban; Pugalenthi, Ganesan; Sowdhamini, Ramanathan

    2004-04-02

    The functional selection and three-dimensional structural constraints of proteins in nature often relates to the retention of significant sequence similarity between proteins of similar fold and function despite poor sequence identity. Organization of structure-based sequence alignments for distantly related proteins, provides a map of the conserved and critical regions of the protein universe that is useful for the analysis of folding principles, for the evolutionary unification of protein families and for maximizing the information return from experimental structure determination. The Protein Alignment organised as Structural Superfamily (PASS2) database represents continuously updated, structural alignments for evolutionary related, sequentially distant proteins. An automated and updated version of PASS2 is, in direct correspondence with SCOP 1.63, consisting of sequences having identity below 40% among themselves. Protein domains have been grouped into 628 multi-member superfamilies and 566 single member superfamilies. Structure-based sequence alignments for the superfamilies have been obtained using COMPARER, while initial equivalencies have been derived from a preliminary superposition using LSQMAN or STAMP 4.0. The final sequence alignments have been annotated for structural features using JOY4.0. The database is supplemented with sequence relatives belonging to different genomes, conserved spatially interacting and structural motifs, probabilistic hidden markov models of superfamilies based on the alignments and useful links to other databases. Probabilistic models and sensitive position specific profiles obtained from reliable superfamily alignments aid annotation of remote homologues and are useful tools in structural and functional genomics. PASS2 presents the phylogeny of its members both based on sequence and structural dissimilarities. Clustering of members allows us to understand diversification of the family members. The search engine has been

  15. Searching for the fastest dynamo: laminar ABC flows.

    PubMed

    Alexakis, Alexandros

    2011-08-01

    The growth rate of the dynamo instability as a function of the magnetic Reynolds number R(M) is investigated by means of numerical simulations for the family of the Arnold-Beltrami-Childress (ABC) flows and for two different forcing scales. For the ABC flows that are driven at the largest available length scale, it is found that, as the magnetic Reynolds number is increased: (a) The flow that results first in a dynamo is the 2 1/2-dimensional flow for which A=B and C=0 (and all permutations). (b) The second type of flow that results in a dynamo is the one for which A=B≃2C/5 (and permutations). (c) The most symmetric flow, A=B=C, is the third type of flow that results in a dynamo. (d) As R(M) is increased, the A=B=C flow stops being a dynamo and transitions from a local maximum to a third-order saddle point. (e) At larger R(M), the A=B=C flow reestablishes itself as a dynamo but remains a saddle point. (f) At the largest examined R(M), the growth rate of the 2 1/2-dimensional flows starts to decay, the A=B=C flow comes close to a local maximum again, and the flow A=B≃2C/5 (and permutations) results in the fastest dynamo with growth rate γ≃0.12 at the largest examined R(M). For the ABC flows that are driven at the second largest available length scale, it is found that (a) the 2 1/2-dimensional flows A=B,C=0 (and permutations) are again the first flows that result in a dynamo with a decreased onset. (b) The most symmetric flow, A=B=C, is the second type of flow that results in a dynamo. It is, and it remains, a local maximum. (c) At larger R(M), the flow A=B≃2C/5 (and permutations) appears as the third type of flow that results in a dynamo. As R(M) is increased, it becomes the flow with the largest growth rate. The growth rates appear to have some correlation with the Lyapunov exponents, but constructive refolding of the field lines appears equally important in determining the fastest dynamo flow.

  16. Attenuated mutant strain of Salmonella Typhimurium lacking the ZnuABC transporter contrasts tumor growth promoting anti-cancer immune response.

    PubMed

    Chirullo, Barbara; Ammendola, Serena; Leonardi, Leonardo; Falcini, Roberto; Petrucci, Paola; Pistoia, Claudia; Vendetti, Silvia; Battistoni, Andrea; Pasquali, Paolo

    2015-07-10

    Salmonella Typhimurium has been shown to be highly effective as antitumor agent. The aim of this study was to investigate the tumor targeting efficacy and the mechanism of action of a specific attenuated mutant strain of Salmonella Typhimurium (STM) devoid of the whole operon coding for the high-affinity zinc transporter ZnuABC, which is required for bacterial growth in environments poor in zinc and for conferring full virulence to different Gram-negative pathogens.We showed that STM is able to penetrate and replicate into tumor cells in in vitro and in vivo models. The subcutaneous administration of STM in mammary adenocarcinoma mouse model led to both reduction of tumor growth and increase in life expectancy of STM treated mice. Moreover, investigating the potential mechanism behind the favorable clinical outcomes, we provide evidence that STM stimulates a potent inflammatory response and a specific immune pattern, recruiting a large number of innate and adaptive immune cells capable to contrast the immunosuppressive environment generated by tumors.

  17. Ancient expansion of the ribonuclease A superfamily revealed by genomic analysis of placental and marsupial mammals.

    PubMed

    Cho, Soochin; Zhang, Jianzhi

    2006-05-24

    Members of the ribonuclease (RNase) A superfamily participate in a diverse array of biological processes, including digestion, angiogenesis, innate immunity, and possibly male reproduction. The superfamily is vertebrate-specific, with 13-20 highly divergent members in primates and rodents, but only a few members in chicken and fish. This has led to the proposal that the superfamily started off from a progenitor with structural similarities to angiogenin and that the superfamily underwent a dramatic expansion during mammalian evolution. To date this evolutionary expansion and understand the functional diversification of the superfamily, we here determine its entire repertoire in the sequenced genomes of dog, cow, and opossum. We identified 7, 20, and 21 putatively functional RNase genes from these three species, respectively. Many of the identified genes are highly divergent from all previously known RNase genes, thus representing new lineages within the superfamily. Phylogenetic analysis indicates that the superfamily expansion predated the separation of placental and marsupial mammals and that differential gene loss and duplication occurred in different species, generating a great variation in gene number and content among extant mammals.

  18. Whole genome re-sequencing identifies a mutation in an ABC transporter (mdr2) in a Plasmodium chabaudi clone with altered susceptibility to antifolate drugs☆

    PubMed Central

    Martinelli, Axel; Henriques, Gisela; Cravo, Pedro; Hunt, Paul

    2011-01-01

    In malaria parasites, mutations in two genes of folate biosynthesis encoding dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) modify responses to antifolate therapies which target these enzymes. However, the involvement of other genes which modify the availability of exogenous folate, for example, has been proposed. Here, we used short-read whole-genome re-sequencing to determine the mutations in a clone of the rodent malaria parasite, Plasmodium chabaudi, which has altered susceptibility to both sulphadoxine and pyrimethamine. This clone bears a previously identified S106N mutation in dhfr and no mutation in dhps. Instead, three additional point mutations in genes on chromosomes 2, 13 and 14 were identified. The mutated gene on chromosome 13 (mdr2 K392Q) encodes an ABC transporter. Because Quantitative Trait Locus analysis previously indicated an association of genetic markers on chromosome 13 with responses to individual and combined antifolates, MDR2 is proposed to modulate antifolate responses, possibly mediated by the transport of folate intermediates. PMID:20858498

  19. ABCE1 is essential for S phase progression in human cells

    PubMed Central

    Toompuu, Marina; Kärblane, Kairi; Pata, Pille; Truve, Erkki; Sarmiento, Cecilia

    2016-01-01

    ABSTRACT ABCE1 is a highly conserved protein universally present in eukaryotes and archaea, which is crucial for the viability of different organisms. First identified as RNase L inhibitor, ABCE1 is currently recognized as an essential translation factor involved in several stages of eukaryotic translation and ribosome biogenesis. The nature of vital functions of ABCE1, however, remains unexplained. Here, we study the role of ABCE1 in human cell proliferation and its possible connection to translation. We show that ABCE1 depletion by siRNA results in a decreased rate of cell growth due to accumulation of cells in S phase, which is accompanied by inefficient DNA synthesis and reduced histone mRNA and protein levels. We infer that in addition to the role in general translation, ABCE1 is involved in histone biosynthesis and DNA replication and therefore is essential for normal S phase progression. In addition, we analyze whether ABCE1 is implicated in transcript-specific translation via its association with the eIF3 complex subunits known to control the synthesis of cell proliferation-related proteins. The expression levels of a few such targets regulated by eIF3A, however, were not consistently affected by ABCE1 depletion. PMID:26985706

  20. Comparison of molecular dynamics and superfamily spaces of protein domain deformation.

    PubMed

    Velázquez-Muriel, Javier A; Rueda, Manuel; Cuesta, Isabel; Pascual-Montano, Alberto; Orozco, Modesto; Carazo, José-María

    2009-02-17

    It is well known the strong relationship between protein structure and flexibility, on one hand, and biological protein function, on the other hand. Technically, protein flexibility exploration is an essential task in many applications, such as protein structure prediction and modeling. In this contribution we have compared two different approaches to explore the flexibility space of protein domains: i) molecular dynamics (MD-space), and ii) the study of the structural changes within superfamily (SF-space). Our analysis indicates that the MD-space and the SF-space display a significant overlap, but are still different enough to be considered as complementary. The SF-space space is wider but less complex than the MD-space, irrespective of the number of members in the superfamily. Also, the SF-space does not sample all possibilities offered by the MD-space, but often introduces very large changes along just a few deformation modes, whose number tend to a plateau as the number of related folds in the superfamily increases. Theoretically, we obtained two conclusions. First, that function restricts the access to some flexibility patterns to evolution, as we observe that when a superfamily member changes to become another, the path does not completely overlap with the physical deformability. Second, that conformational changes from variation in a superfamily are larger and much simpler than those allowed by physical deformability. Methodologically, the conclusion is that both spaces studied are complementary, and have different size and complexity. We expect this fact to have application in fields as 3D-EM/X-ray hybrid models or ab initio protein folding.

  1. CREATING AN IPHONE APPLICATION FOR COLLECTING CONTINUOUS ABC DATA

    PubMed Central

    Whiting, Seth W; Dixon, Mark R

    2012-01-01

    This paper provides an overview and task analysis for creating a continuous ABC data-collection application using Xcode on a Mac computer. Behavior analysts can program an ABC data collection system, complete with a customized list of target clients, antecedents, behaviors, and consequences to be recorded, and have the data automatically sent to an e-mail account after observations have concluded. Further suggestions are provided to customize the ABC data- collection system for individual preferences and clinical needs. PMID:23060682

  2. ABCs of Being Smart: S Is for Supporting

    ERIC Educational Resources Information Center

    Foster, Joanne

    2014-01-01

    Joanne Foster's article "R We There Yet?" was first published in "Parenting for High Potential" ("PHP") in 2006, which became the springboard for the "ABCs of Being Smart" series of columns. At that time, Foster invited "PHP" readers to think about their own versions of the "ABCs of Being…

  3. Suppression of c-Myc is involved in multi-walled carbon nanotubes' down-regulation of ATP-binding cassette transporters in human colon adenocarcinoma cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wang, Zhaojing; Xu, Yonghong; Meng, Xiangning

    Over-expression of ATP-binding cassette (ABC) transporters, a large family of integral membrane proteins that decrease cellular drug uptake and accumulation by active extrusion, is one of the major causes of cancer multi-drug resistance (MDR) that frequently leads to failure of chemotherapy. Carbon nanotubes (CNTs)-based drug delivery devices hold great promise in enhancing the efficacy of cancer chemotherapy. However, CNTs' effects on the ABC transporters remain under-investigated. In this study, we found that multiwalled carbon nanotubes (MWCNTs) reduced transport activity and expression of ABC transporters including ABCB1/Pgp and ABCC4/MRP4 in human colon adenocarcinoma Caco-2 cells. Proto-oncogene c-Myc, which directly regulates ABCmore » gene expression, was concurrently decreased in MWCNT-treated cells and forced over-expression of c-Myc reversed MWCNTs' inhibitory effects on ABCB1 and ABCC4 expression. MWCNT-cell membrane interaction and cell membrane oxidative damage were observed. However, antioxidants such as vitamin C, β-mecaptoethanol and dimethylthiourea failed to antagonize MWCNTs' down-regulation of ABC transporters. These data suggest that MWCNTs may act on c-Myc, but not through oxidative stress, to down-regulate ABC transporter expression. Our findings thus shed light on CNTs' novel cellular effects that may be utilized to develop CNTs-based drug delivery devices to overcome ABC transporter-mediated cancer chemoresistance.« less

  4. Phase 0 and phase III transport in various organs: combined concept of phases in xenobiotic transport and metabolism.

    PubMed

    Döring, Barbara; Petzinger, Ernst

    2014-08-01

    The historical phasing concept of drug metabolism and elimination was introduced to comprise the two phases of metabolism: phase I metabolism for oxidations, reductions and hydrolyses, and phase II metabolism for synthesis. With this concept, biological membrane barriers obstructing the accessibility of metabolism sites in the cells for drugs were not considered. The concept of two phases was extended to a concept of four phases when drug transporters were detected that guided drugs and drug metabolites in and out of the cells. In particular, water soluble or charged drugs are virtually not able to overcome the phospholipid membrane barrier. Drug transporters belong to two main clusters of transporter families: the solute carrier (SLC) families and the ATP binding cassette (ABC) carriers. The ABC transporters comprise seven families with about 20 carriers involved in drug transport. All of them operate as pumps at the expense of ATP splitting. Embedded in the former phase concept, the term "phase III" was introduced by Ishikawa in 1992 for drug export by ABC efflux pumps. SLC comprise 52 families, from which many carriers are drug uptake transporters. Later on, this uptake process was referred to as the "phase 0 transport" of drugs. Transporters for xenobiotics in man and animal are most expressed in liver, but they are also present in extra-hepatic tissues such as in the kidney, the adrenal gland and lung. This review deals with the function of drug carriers in various organs and their impact on drug metabolism and elimination.

  5. Evolutionary and molecular foundations of multiple contemporary functions of the nitroreductase superfamily

    PubMed Central

    Akiva, Eyal; Copp, Janine N.; Tokuriki, Nobuhiko; Babbitt, Patricia C.

    2017-01-01

    Insight regarding how diverse enzymatic functions and reactions have evolved from ancestral scaffolds is fundamental to understanding chemical and evolutionary biology, and for the exploitation of enzymes for biotechnology. We undertook an extensive computational analysis using a unique and comprehensive combination of tools that include large-scale phylogenetic reconstruction to determine the sequence, structural, and functional relationships of the functionally diverse flavin mononucleotide-dependent nitroreductase (NTR) superfamily (>24,000 sequences from all domains of life, 54 structures, and >10 enzymatic functions). Our results suggest an evolutionary model in which contemporary subgroups of the superfamily have diverged in a radial manner from a minimal flavin-binding scaffold. We identified the structural design principle for this divergence: Insertions at key positions in the minimal scaffold that, combined with the fixation of key residues, have led to functional specialization. These results will aid future efforts to delineate the emergence of functional diversity in enzyme superfamilies, provide clues for functional inference for superfamily members of unknown function, and facilitate rational redesign of the NTR scaffold. PMID:29078300

  6. Regulation of ATP-binding Cassette Transporters and Cholesterol Efflux by Glucose in Primary Human Monocytes and Murine Bone Marrow-derived Macrophages

    PubMed Central

    Spartano, N. L.; Lamon-Fava, S.; Matthan, N. R.; Ronxhi, J.; Greenberg, A. S.; Obin, M. S.; Lichtenstein, A. H.

    2014-01-01

    Purpose Individuals with type 2 diabetes mellitus are at increased risk of developing atherosclerosis. This may be partially attributable to suppression of macrophage ATP-binding cassette (ABC) transporter mediated cholesterol efflux by sustained elevated blood glucose concentrations. 2 models were used to assess this potential relationship: human monocytes/leukocytes and murine bone marrow-derived macrophages (BMDM). Methods 10 subjects (4 F/6 M, 50–85 years, BMI 25–35 kg/m2) underwent an oral glucose challenge. Baseline and 1- and 2-h post-challenge ABC-transporter mRNA expression was determined in monocytes, leukocytes and peripheral blood mononuclear cells (PBMC). In a separate study, murine-BMDM were exposed to 5 mmol/L D-glucose (control) or additional 20 mmol/L D-or L-glucose and 25 ug/mL oxidized low density lipoprotein (oxLDL). High density lipoprotein (HDL)-mediated cholesterol efflux and ABC-transporter (ABCA1 and ABCG1) expression were determined. Results Baseline ABCA1and ABCG1 expression was lower (> 50 %) in human monocytes and PBMC than leukocytes (p < 0.05). 1 h post-challenge leukocyte ABCA1 and ABCG1 expression increased by 37 % and 30 %, respectively (p < 0.05), and began to return to baseline thereafter. There was no significant change in monocyte ABC-transporter expression. In murine BMDM, higher glucose concentrations suppressed HDL-mediated cholesterol efflux (10 %; p < 0.01) without significantly affecting ABCA1 and ABCG1 expression. Data demonstrate that leukocytes are not a reliable indicator of monocyte ABC-transporter expression. Conclusions Human monocyte ABC-transporter gene expression was unresponsive to a glucose challenge. Correspondingly, in BMDM, hyperglycemia attenuated macrophage cholesterol efflux in the absence of altered ABC-transporter expression, suggesting that hyperglycemia, per se, suppresses cholesterol transporter activity. This glucose-related impairment in cholesterol efflux may potentially contribute to

  7. A strategy for detecting the conservation of folding-nucleus residues in protein superfamilies.

    PubMed

    Michnick, S W; Shakhnovich, E

    1998-01-01

    Nucleation-growth theory predicts that fast-folding peptide sequences fold to their native structure via structures in a transition-state ensemble that share a small number of native contacts (the folding nucleus). Experimental and theoretical studies of proteins suggest that residues participating in folding nuclei are conserved among homologs. We attempted to determine if this is true in proteins with highly diverged sequences but identical folds (superfamilies). We describe a strategy based on comparisons of residue conservation in natural superfamily sequences with simulated sequences (generated with a Monte-Carlo sequence design strategy) for the same proteins. The basic assumptions of the strategy were that natural sequences will conserve residues needed for folding and stability plus function, the simulated sequences contain no functional conservation, and nucleus residues make native contacts with each other. Based on these assumptions, we identified seven potential nucleus residues in ubiquitin superfamily members. Non-nucleus conserved residues were also identified; these are proposed to be involved in stabilizing native interactions. We found that all superfamily members conserved the same potential nucleus residue positions, except those for which the structural topology is significantly different. Our results suggest that the conservation of the nucleus of a specific fold can be predicted by comparing designed simulated sequences with natural highly diverged sequences that fold to the same structure. We suggest that such a strategy could be used to help plan protein folding and design experiments, to identify new superfamily members, and to subdivide superfamilies further into classes having a similar folding mechanism.

  8. Kinetic control of TolC recruitment by multidrug efflux complexes.

    PubMed

    Tikhonova, Elena B; Dastidar, Vishakha; Rybenkov, Valentin V; Zgurskaya, Helen I

    2009-09-22

    In Gram-negative pathogens, multidrug efflux pumps that provide clinically significant levels of antibiotic resistance function as three-component complexes. They are composed of the inner membrane transporters belonging to one of three superfamilies of proteins, RND, ABC, or MF; periplasmic proteins belonging to the membrane fusion protein (MFP) family; and outer membrane channels exemplified by the Escherichia coli TolC. The three-component complexes span the entire two-membrane envelope of Gram-negative bacteria and expel toxic molecules from the cytoplasmic membrane to the medium. The architecture of these complexes is expected to vary significantly because of the structural diversity of the inner membrane transporters. How the three-component pumps are assembled, their architecture, and their dynamics remain unclear. In this study, we reconstituted interactions and compared binding kinetics of the E. coli TolC with AcrA, MacA, and EmrA, the periplasmic MFPs that function in multidrug efflux with transporters from the RND, ABC, and MF superfamilies, respectively. By using surface plasmon resonance, we demonstrate that TolC interactions with MFPs are highly dynamic and sensitive to pH. The affinity of TolC to MFPs decreases in the order MacA > EmrA > AcrA. We further show that MFPs are prone to oligomerization, but differ dramatically from each other in oligomerization kinetics and stability of oligomers. The propensity of MFPs to oligomerize correlates with the stability of MFP-TolC complexes and structural features of inner membrane transporters. We propose that recruitment of TolC by various MFPs is determined not only by kinetics of MFP-TolC interactions but also by oligomerization kinetics of MFPs and pH.

  9. Characterization and expression profiling of ATP-binding cassette transporter genes in the diamondback moth, Plutella xylostella (L.).

    PubMed

    Qi, Weiping; Ma, Xiaoli; He, Weiyi; Chen, Wei; Zou, Mingmin; Gurr, Geoff M; Vasseur, Liette; You, Minsheng

    2016-09-27

    ATP-binding cassette (ABC) transporters are one of the major transmembrane protein families found in all organisms and play important roles in transporting a variety of compounds across intra and extra cellular membranes. In some species, ABC transporters may be involved in the detoxification of substances such as insecticides. The diamondback moth, Plutella xylostella (L.), a destructive pest of cruciferous crops worldwide, is an important species to study as it is resistant to many types of insecticides as well as biological control Bacillus thuringiensis toxins. A total of 82 ABC genes were identified from our published P. xylostella genome, and grouped into eight subfamilies (ABCA-H) based on phylogenetic analysis. Genes of subfamilies ABCA, ABCC and ABCH were found to be expanded in P. xylostella compared with those in Bombyx mori, Manduca sexta, Heliconius melpomene, Danaus plexippus, Drosophila melanogaster, Tetranychus urticae and Homo sapiens. Phylogenetic analysis indicated that many of the ABC transporters in P. xylostella are orthologous to the well-studied ABC transporter genes in the seven other species. Transcriptome- and qRT-PCR-based analysis elucidated physiological effects of ABC gene expressions of P. xylostella which were developmental stage- and tissue-specific as well as being affected by whether or not the insects were from an insecticide-resistant strain. Two ABCC and one ABCA genes were preferentially expressed in midgut of the 4th-instar larvae of a susceptible strain (Fuzhou-S) suggesting their potential roles in metabolizing plant defensive chemicals. Most of the highly expressed genes in insecticide-resistant strains were also predominantly expressed in the tissues of Malpighian tubules and midgut. This is the most comprehensive study on identification, characterization and expression profiling of ABC transporter genes in P. xylostella to date. The diversified features and expression patterns of this gene family may be associated with

  10. Comparison of molecular dynamics and superfamily spaces of protein domain deformation

    PubMed Central

    Velázquez-Muriel, Javier A; Rueda, Manuel; Cuesta, Isabel; Pascual-Montano, Alberto; Orozco, Modesto; Carazo, José-María

    2009-01-01

    Background It is well known the strong relationship between protein structure and flexibility, on one hand, and biological protein function, on the other hand. Technically, protein flexibility exploration is an essential task in many applications, such as protein structure prediction and modeling. In this contribution we have compared two different approaches to explore the flexibility space of protein domains: i) molecular dynamics (MD-space), and ii) the study of the structural changes within superfamily (SF-space). Results Our analysis indicates that the MD-space and the SF-space display a significant overlap, but are still different enough to be considered as complementary. The SF-space space is wider but less complex than the MD-space, irrespective of the number of members in the superfamily. Also, the SF-space does not sample all possibilities offered by the MD-space, but often introduces very large changes along just a few deformation modes, whose number tend to a plateau as the number of related folds in the superfamily increases. Conclusion Theoretically, we obtained two conclusions. First, that function restricts the access to some flexibility patterns to evolution, as we observe that when a superfamily member changes to become another, the path does not completely overlap with the physical deformability. Second, that conformational changes from variation in a superfamily are larger and much simpler than those allowed by physical deformability. Methodologically, the conclusion is that both spaces studied are complementary, and have different size and complexity. We expect this fact to have application in fields as 3D-EM/X-ray hybrid models or ab initio protein folding. PMID:19220918

  11. SatR Is a Repressor of Fluoroquinolone Efflux Pump SatAB

    PubMed Central

    Escudero, Jose Antonio; San Millan, Alvaro; Montero, Natalia; Gutierrez, Belen; Ovejero, Cristina Martinez; Carrilero, Laura

    2013-01-01

    Streptococcus suis is an emerging zoonotic agent responsible for high-mortality outbreaks among the human population in China. In this species, the ABC transporter SatAB mediates fluoroquinolone resistance when overexpressed. Here, we describe and characterize satR, an open reading frame (ORF) encoding a MarR superfamily regulator that acts as a repressor of satAB. satR is cotranscribed with satAB, and its interruption entails the overexpression of the pump, leading to a clinically relevant increase in resistance to fluoroquinolones. PMID:23650171

  12. Optimization of Straight Cylindrical Turning Using Artificial Bee Colony (ABC) Algorithm

    NASA Astrophysics Data System (ADS)

    Prasanth, Rajanampalli Seshasai Srinivasa; Hans Raj, Kandikonda

    2017-04-01

    Artificial bee colony (ABC) algorithm, that mimics the intelligent foraging behavior of honey bees, is increasingly gaining acceptance in the field of process optimization, as it is capable of handling nonlinearity, complexity and uncertainty. Straight cylindrical turning is a complex and nonlinear machining process which involves the selection of appropriate cutting parameters that affect the quality of the workpiece. This paper presents the estimation of optimal cutting parameters of the straight cylindrical turning process using the ABC algorithm. The ABC algorithm is first tested on four benchmark problems of numerical optimization and its performance is compared with genetic algorithm (GA) and ant colony optimization (ACO) algorithm. Results indicate that, the rate of convergence of ABC algorithm is better than GA and ACO. Then, the ABC algorithm is used to predict optimal cutting parameters such as cutting speed, feed rate, depth of cut and tool nose radius to achieve good surface finish. Results indicate that, the ABC algorithm estimated a comparable surface finish when compared with real coded genetic algorithm and differential evolution algorithm.

  13. Translocation of the ABC transporter ABCD4 from the endoplasmic reticulum to lysosomes requires the escort protein LMBD1.

    PubMed

    Kawaguchi, Kosuke; Okamoto, Takumi; Morita, Masashi; Imanaka, Tsuneo

    2016-07-26

    We previously demonstrated that ABCD4 does not localize to peroxisomes but rather, the endoplasmic reticulum (ER), because it lacks the NH2-terminal hydrophilic region required for peroxisomal targeting. It was recently reported that mutations in ABCD4 result in a failure to release vitamin B12 from lysosomes. A similar phenotype is caused by mutations in LMBRD1, which encodes the lysosomal membrane protein LMBD1. These findings suggested to us that ABCD4 translocated from the ER to lysosomes in association with LMBD1. In this report, it is demonstrated that ABCD4 interacts with LMBD1 and then localizes to lysosomes, and this translocation depends on the lysosomal targeting ability of LMBD1. Furthermore, endogenous ABCD4 was localized to both lysosomes and the ER, and its lysosomal localization was disturbed by knockout of LMBRD1. To the best of our knowledge, this is the first report demonstrating that the subcellular localization of the ABC transporter is determined by its association with an adaptor protein.

  14. Bombyx mori ABC transporter C2 structures responsible for the receptor function of Bacillus thuringiensis Cry1Aa toxin.

    PubMed

    Tanaka, Shiho; Endo, Haruka; Adegawa, Satomi; Iizuka, Ami; Imamura, Kazuhiro; Kikuta, Shingo; Sato, Ryoichi

    2017-12-01

    Because Bombyx mori ABC transporter C2 (BmABCC2) has 1000-fold higher potential than B. mori cadherin-like protein as a receptor for Bacillus thuringiensis Cry1Aa toxin (Tanaka et al., 2013), the gate-opening ability of the latent pore under six extracellular loops (ECLs) of BmABCC2 was expected to be the reason for its higher potential (Heckel, 2012). In this study, cell swelling assays in Sf9 cells showed that BmABCC2 mutants lacking substrate-excreting activity retained receptor activity, indicating that the gate-opening activity of BmABCC2 is not responsible for Cry1Aa toxicity. The analysis of 29 BmABCC2 mutants demonstrated that 770 DYWL 773 of ECL 4 comprise a putative binding site to Cry1Aa. This suggests that specific toxicity of Cry1Aa toxin to a restricted range of lepidopteran insects is dependent on conservation and variation in the amino acid residues around 770 DYWL 773 of ECL 4 in the ABCC2. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Serum albumin promotes ATP-binding cassette transporter-dependent sterol uptake in yeast.

    PubMed

    Marek, Magdalena; Silvestro, Daniele; Fredslund, Maria D; Andersen, Tonni G; Pomorski, Thomas G

    2014-12-01

    Sterol uptake in fungi is a multistep process that involves interaction between external sterols and the cell wall, incorporation of sterol molecules into the plasma membrane, and subsequent integration into intracellular membranes for turnover. ATP-binding cassette (ABC) transporters have been implicated in sterol uptake, but key features of their activity remain to be elucidated. Here, we apply fluorescent cholesterol (NBD-cholesterol) to monitor sterol uptake under anaerobic and aerobic conditions in two fungal species, Candida glabrata (Cg) and Saccharomyces cerevisiae (Sc). We found that in both fungal species, ABC transporter-dependent uptake of cholesterol under anaerobic conditions and in mutants lacking HEM1 gene is promoted in the presence of the serum protein albumin that is able to bind the sterol molecule. Furthermore, the C. glabrata ABC transporter CgAus1p expressed in S. cerevisiae requires the presence of serum or albumin for efficient cholesterol uptake. These results suggest that albumin can serve as sterol donor in ABC transporter-dependent sterol uptake, a process potentially important for growth of C. glabrata inside infected humans. © 2014 The Authors. FEMS Yeast Research published by John Wiley & Sons Ltd on behalf of Federation of European Microbiological Societies.

  16. Transient receptor potential channel superfamily: Role in lower urinary tract function.

    PubMed

    Ogawa, Teruyuki; Imamura, Tetsuya; Nakazawa, Masaki; Hiragata, Shiro; Nagai, Takashi; Minagawa, Tomonori; Yokoyama, Hitoshi; Ishikawa, Masakuni; Domen, Takahisa; Ishizuka, Osamu

    2015-11-01

    Lower urinary tract symptoms associated with neurogenic bladder and overactive bladder syndrome are mediated in part by members of the transient receptor potential channel superfamily. The best studied member of this superfamily is the vanilloid receptor. Other transient receptor potential channels, such as the melastatin receptor and the ankyrin receptor, are also active in the pathogenesis of lower urinary tract dysfunction. However, the detailed mechanisms by which the transient receptor potential channels contribute to lower urinary tract symptoms are still not clear, and the therapeutic benefits of modulating transient receptor potential channel activity have not been proved in the clinical setting. In the present review, to better understand the pathophysiology and therapeutic potential for lower urinary tract symptoms, we summarize the presence and role of different members of the transient receptor potential channel superfamily in the lower urinary tract. © 2015 The Japanese Urological Association.

  17. Evolution of genes and repeats in the Nimrod superfamily.

    PubMed

    Somogyi, Kálmán; Sipos, Botond; Pénzes, Zsolt; Kurucz, Eva; Zsámboki, János; Hultmark, Dan; Andó, István

    2008-11-01

    The recently identified Nimrod superfamily is characterized by the presence of a special type of EGF repeat, the NIM repeat, located right after a typical CCXGY/W amino acid motif. On the basis of structural features, nimrod genes can be divided into three types. The proteins encoded by Draper-type genes have an EMI domain at the N-terminal part and only one copy of the NIM motif, followed by a variable number of EGF-like repeats. The products of Nimrod B-type and Nimrod C-type genes (including the eater gene) have different kinds of N-terminal domains, and lack EGF-like repeats but contain a variable number of NIM repeats. Draper and Nimrod C-type (but not Nimrod B-type) proteins carry a transmembrane domain. Several members of the superfamily were claimed to function as receptors in phagocytosis and/or binding of bacteria, which indicates an important role in the cellular immunity and the elimination of apoptotic cells. In this paper, the evolution of the Nimrod superfamily is studied with various methods on the level of genes and repeats. A hypothesis is presented in which the NIM repeat, along with the EMI domain, emerged by structural reorganizations at the end of an EGF-like repeat chain, suggesting a mechanism for the formation of novel types of repeats. The analyses revealed diverse evolutionary patterns in the sequences containing multiple NIM repeats. Although in the Nimrod B and Nimrod C proteins show characteristics of independent evolution, many internal NIM repeats in Eater sequences seem to have undergone concerted evolution. An analysis of the nimrod genes has been performed using phylogenetic and other methods and an evolutionary scenario of the origin and diversification of the Nimrod superfamily is proposed. Our study presents an intriguing example how the evolution of multigene families may contribute to the complexity of the innate immune response.

  18. Application of activity-based costing (ABC) for a Peruvian NGO healthcare provider.

    PubMed

    Waters, H; Abdallah, H; Santillán, D

    2001-01-01

    This article describes the application of activity-based costing (ABC) to calculate the unit costs of the services for a health care provider in Peru. While traditional costing allocates overhead and indirect costs in proportion to production volume or to direct costs, ABC assigns costs through activities within an organization. ABC uses personnel interviews to determine principal activities and the distribution of individual's time among these activities. Indirect costs are linked to services through time allocation and other tracing methods, and the result is a more accurate estimate of unit costs. The study concludes that applying ABC in a developing country setting is feasible, yielding results that are directly applicable to pricing and management. ABC determines costs for individual clinics, departments and services according to the activities that originate these costs, showing where an organization spends its money. With this information, it is possible to identify services that are generating extra revenue and those operating at a loss, and to calculate cross subsidies across services. ABC also highlights areas in the health care process where efficiency improvements are possible. Conclusions about the ultimate impact of the methodology are not drawn here, since the study was not repeated and changes in utilization patterns and the addition of new clinics affected applicability of the results. A potential constraint to implementing ABC is the availability and organization of cost information. Applying ABC efficiently requires information to be readily available, by cost category and department, since the greatest benefits of ABC come from frequent, systematic application of the methodology in order to monitor efficiency and provide feedback for management. The article concludes with a discussion of the potential applications of ABC in the health sector in developing countries.

  19. Creating an iPhone application for collecting continuous ABC data.

    PubMed

    Whiting, Seth W; Dixon, Mark R

    2012-01-01

    This paper provides an overview and task analysis for creating a continuous ABC data-collection application using Xcode on a Mac computer. Behavior analysts can program an ABC data collection system, complete with a customized list of target clients, antecedents, behaviors, and consequences to be recorded, and have the data automatically sent to an e-mail account after observations have concluded. Further suggestions are provided to customize the ABC data- collection system for individual preferences and clinical needs.

  20. ABC versus CAB for cardiopulmonary resuscitation: a prospective, randomized simulator-based trial.

    PubMed

    Marsch, Stephan; Tschan, Franziska; Semmer, Norbert K; Zobrist, Roger; Hunziker, Patrick R; Hunziker, Sabina

    2013-09-06

    After years of advocating ABC (Airway-Breathing-Circulation), current guidelines of cardiopulmonary resuscitation (CPR) recommend CAB (Circulation-Airway-Breathing). This trial compared ABC with CAB as initial approach to CPR from the arrival of rescuers until the completion of the first resuscitation cycle. 108 teams, consisting of two physicians each, were randomized to receive a graphical display of either the ABC algorithm or the CAB algorithm. Subsequently teams had to treat a simulated cardiac arrest. Data analysis was performed using video recordings obtained during simulations. The primary endpoint was the time to completion of the first resuscitation cycle of 30 compressions and two ventilations. The time to execution of the first resuscitation measure was 32 ± 12 seconds in ABC teams and 25 ± 10 seconds in CAB teams (P = 0.002). 18/53 ABC teams (34%) and none of the 55 CAB teams (P = 0.006) applied more than the recommended two initial rescue breaths which caused a longer duration of the first cycle of 30 compressions and two ventilations in ABC teams (31 ± 13 vs.23 ± 6 sec; P = 0.001). Overall, the time to completion of the first resuscitation cycle was longer in ABC teams (63 ± 17 vs. 48 ± 10 sec; P <0.0001). This randomized controlled trial found CAB superior to ABC with an earlier start of CPR and a shorter time to completion of the first 30:2 resuscitation cycle. These findings endorse the change from ABC to CAB in international resuscitation guidelines.

  1. Characterization of a lactose-responsive promoter of ATP-binding cassette (ABC) transporter gene from Lactobacillus acidophilus 05-172.

    PubMed

    Zeng, Zhu; Zuo, Fanglei; Yu, Rui; Zhang, Bo; Ma, Huiqin; Chen, Shangwu

    2017-09-01

    A novel lactose-responsive promoter of the ATP-binding cassette (ABC) transporter gene Lba1680 of Lactobacillus acidophilus strain 05-172 isolated from a traditionally fermented dairy product koumiss was characterized. In L. acidophilus 05-172, expression of Lba1680 was induced by lactose, with lactose-induced transcription of Lba1680 being 6.1-fold higher than that induced by glucose. This is in contrast to L. acidophilus NCFM, a strain isolated from human feces, in which expression of Lba1680 and Lba1679 is induced by glucose. Both gene expression and enzyme activity assays in L. paracasei transformed with a vector containing the inducible Lba1680 promoter (PLba1680) of strain 05-172 and a heme-dependent catalase gene as reporter confirmed that PLba1680 is specifically induced by lactose. Its regulatory expression could not be repressed by glucose, and was independent of cAMP receptor protein. This lactose-responsive promoter might be used in the expression of functional genes in L. paracasei incorporated into a lactose-rich environment, such as dairy products. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Repositioning of Tyrosine Kinase Inhibitors as Antagonists of ATP-Binding Cassette Transporters in Anticancer Drug Resistance

    PubMed Central

    Wang, Yi-Jun; Zhang, Yun-Kai; Kathawala, Rishil J.; Chen, Zhe-Sheng

    2014-01-01

    The phenomenon of multidrug resistance (MDR) has attenuated the efficacy of anticancer drugs and the possibility of successful cancer chemotherapy. ATP-binding cassette (ABC) transporters play an essential role in mediating MDR in cancer cells by increasing efflux of drugs from cancer cells, hence reducing the intracellular accumulation of chemotherapeutic drugs. Interestingly, small-molecule tyrosine kinase inhibitors (TKIs), such as AST1306, lapatinib, linsitinib, masitinib, motesanib, nilotinib, telatinib and WHI-P154, have been found to have the capability to overcome anticancer drug resistance by inhibiting ABC transporters in recent years. This review will focus on some of the latest and clinical developments with ABC transporters, TKIs and anticancer drug resistance. PMID:25268163

  3. Molecular evolution of miraculin-like proteins in soybean Kunitz super-family.

    PubMed

    Selvakumar, Purushotham; Gahloth, Deepankar; Tomar, Prabhat Pratap Singh; Sharma, Nidhi; Sharma, Ashwani Kumar

    2011-12-01

    Miraculin-like proteins (MLPs) belong to soybean Kunitz super-family and have been characterized from many plant families like Rutaceae, Solanaceae, Rubiaceae, etc. Many of them possess trypsin inhibitory activity and are involved in plant defense. MLPs exhibit significant sequence identity (~30-95%) to native miraculin protein, also belonging to Kunitz super-family compared with a typical Kunitz family member (~30%). The sequence and structure-function comparison of MLPs with that of a classical Kunitz inhibitor have demonstrated that MLPs have evolved to form a distinct group within Kunitz super-family. Sequence analysis of new genes along with available MLP sequences in the literature revealed three major groups for these proteins. A significant feature of Rutaceae MLP type 2 sequences is the presence of phosphorylation motif. Subtle changes are seen in putative reactive loop residues among different MLPs suggesting altered specificities to specific proteases. In phylogenetic analysis, Rutaceae MLP type 1 and type 2 proteins clustered together on separate branches, whereas native miraculin along with other MLPs formed distinct clusters. Site-specific positive Darwinian selection was observed at many sites in both the groups of Rutaceae MLP sequences with most of the residues undergoing positive selection located in loop regions. The results demonstrate the sequence and thereby the structure-function divergence of MLPs as a distinct group within soybean Kunitz super-family due to biotic and abiotic stresses of local environment.

  4. Alzheimer's disease: neuroprogesterone, epoxycholesterol, and ABC transporters as determinants of neurodesmosterol tissue levels and its role in amyloid protein processing.

    PubMed

    Javitt, Norman B

    2013-01-01

    Evidence is emerging that during the development of Alzheimer's disease (AD), changes in the synthesis and metabolism of cholesterol and progesterone are occurring that may or may not affect the progression of the disease. The concept arose from the recognition that dehydrocholesterol 24-reductase (DHCR24/Seladin-1), one of the nine enzymes in the endoplasmic reticulum that determines the transformation of lanosterol to cholesterol, is selectively reduced in late AD. As a consequence, the tissue level of desmosterol increases, affecting the expression of ABC transporters and the structure of lipid rafts, both determinants of amyloid-β processing. However, the former effect is considered beneficial and the latter detrimental to processing. Other determinants of desmosterol tissue levels are 24,25 epoxycholesterol and the ABCG1 and ABCG4 transporters. Progesterone and its metabolites are determinants of tissue levels of desmosterol and several other sterol intermediates in cholesterol synthesis. Animal models indicate marked elevations in the tissue levels of these sterols at early time frames in the progression of neurodegenerative diseases. The low level of neuroprogesterone and metabolites in AD are consonant with the low level of desmosterol and may have a role in amyloid-β processing. The sparse data that has accumulated appears to be a sufficient basis for proposing a systematic evaluation of the biologic roles of sterol intermediates in the slowly progressive neurodegeneration characteristic of AD.

  5. Lectins of beneficial microbes: system organisation, functioning and functional superfamily.

    PubMed

    Lakhtin, M; Lakhtin, V; Alyoshkin, V; Afanasyev, S

    2011-06-01

    In this review our last results and proposals with respect to general aspects of lectin studies are summarised and compared. System presence, organisation and functioning of lectins are proposed, and accents on beneficial symbiotic microbial lectins studies are presented. The proposed general principles of lectin functioning allows for a comparison of lectins with other carbohydrate-recognition systems. A new structure-functional superfamily of symbiotic microbial lectins is proposed and its main properties are described. The proposed superfamily allows for extended searches of the biological activities of any microbial member. Prospects of lectins of beneficial symbiotic microorganisms are discussed.

  6. Splice form variant and amino acid changes in MDR49 confers DDT resistance in transgenic Drosophila

    PubMed Central

    Seong, Keon Mook; Sun, Weilin; Clark, John M.; Pittendrigh, Barry R.

    2016-01-01

    The ATP-binding cassette (ABC) transporters represent a superfamily of proteins that have important physiological roles in both prokaryotes and eukaryotes. In insects, ABC transporters have previously been implicated in insecticide resistance. The 91-R strain of Drosophila melanogaster has been intensely selected with DDT over six decades. A recent selective sweeps analysis of 91-R implicated the potential role of MDR49, an ABC transporter, in DDT resistance, however, to date the details of how MDR49 may play a role in resistance have not been elucidated. In this study, we investigated the impact of structural changes and an alternative splicing event in MDR49 on DDT-resistance in 91-R, as compared to the DDT susceptible strain 91-C. We observed three amino acid differences in MDR49 when 91-R was compared with 91-C, and only one isoform (MDR49B) was implicated in DDT resistance. A transgenic Drosophila strain containing the 91-R-MDR49B isoform had a significantly higher LD50 value as compared to the 91-C-MDR49B isoform at the early time points (6 h to 12 h) during DDT exposure. Our data support the hypothesis that the MDR49B isoform, with three amino acid mutations, plays a role in the early aspects of DDT resistance in 91-R. PMID:27003579

  7. Splice form variant and amino acid changes in MDR49 confers DDT resistance in transgenic Drosophila.

    PubMed

    Seong, Keon Mook; Sun, Weilin; Clark, John M; Pittendrigh, Barry R

    2016-03-22

    The ATP-binding cassette (ABC) transporters represent a superfamily of proteins that have important physiological roles in both prokaryotes and eukaryotes. In insects, ABC transporters have previously been implicated in insecticide resistance. The 91-R strain of Drosophila melanogaster has been intensely selected with DDT over six decades. A recent selective sweeps analysis of 91-R implicated the potential role of MDR49, an ABC transporter, in DDT resistance, however, to date the details of how MDR49 may play a role in resistance have not been elucidated. In this study, we investigated the impact of structural changes and an alternative splicing event in MDR49 on DDT-resistance in 91-R, as compared to the DDT susceptible strain 91-C. We observed three amino acid differences in MDR49 when 91-R was compared with 91-C, and only one isoform (MDR49B) was implicated in DDT resistance. A transgenic Drosophila strain containing the 91-R-MDR49B isoform had a significantly higher LD50 value as compared to the 91-C-MDR49B isoform at the early time points (6 h to 12 h) during DDT exposure. Our data support the hypothesis that the MDR49B isoform, with three amino acid mutations, plays a role in the early aspects of DDT resistance in 91-R.

  8. Immunoglobulin superfamily proteins in Caenorhabditis elegans.

    PubMed

    Teichmann, S A; Chothia, C

    2000-03-10

    The predicted proteins of the genome of Caenorhabditis elegans were analysed by various sequence comparison methods to identify the repertoire of proteins that are members of the immunoglobulin superfamily (IgSF). The IgSF is one of the largest families of protein domain in this genome and likely to be one of the major families in other multicellular eukaryotes too. This is because members of the superfamily are involved in a variety of functions including cell-cell recognition, cell-surface receptors, muscle structure and, in higher organisms, the immune system. Sixty-four proteins with 488 I set IgSF domains were identified largely by using Hidden Markov models. The domain architectures of the protein products of these 64 genes are described. Twenty-one of these had been characterised previously. We show that another 25 are related to proteins of known function. The C. elegans IgSF proteins can be classified into five broad categories: muscle proteins, protein kinases and phosphatases, three categories of proteins involved in the development of the nervous system, leucine-rich repeat containing proteins and proteins without homologues of known function, of which there are 18. The 19 proteins involved in nervous system development that are not kinases or phosphatases are homologues of neuroglian, axonin, NCAM, wrapper, klingon, ICCR and nephrin or belong to the recently identified zig gene family. Out of the set of 64 genes, 22 are on the X chromosome. This study should be seen as an initial description of the IgSF repertoire in C. elegans, because the current gene definitions may contain a number of errors, especially in the case of long sequences, and there may be IgSF genes that have not yet been detected. However, the proteins described here do provide an overview of the bulk of the repertoire of immunoglobulin superfamily members in C. elegans, a framework for refinement and extension of the repertoire as gene and protein definitions improve, and the basis

  9. Conservation of Dynamics Associated with Biological Function in an Enzyme Superfamily.

    PubMed

    Narayanan, Chitra; Bernard, David N; Bafna, Khushboo; Gagné, Donald; Chennubhotla, Chakra S; Doucet, Nicolas; Agarwal, Pratul K

    2018-03-06

    Enzyme superfamily members that share common chemical and/or biological functions also share common features. While the role of structure is well characterized, the link between enzyme function and dynamics is not well understood. We present a systematic characterization of intrinsic dynamics of over 20 members of the pancreatic-type RNase superfamily, which share a common structural fold. This study is motivated by the fact that the range of chemical activity as well as molecular motions of RNase homologs spans over 10 5 folds. Dynamics was characterized using a combination of nuclear magnetic resonance experiments and computer simulations. Phylogenetic clustering led to the grouping of sequences into functionally distinct subfamilies. Detailed characterization of the diverse RNases showed conserved dynamical traits for enzymes within subfamilies. These results suggest that selective pressure for the conservation of dynamical behavior, among other factors, may be linked to the distinct chemical and biological functions in an enzyme superfamily. Copyright © 2018 Elsevier Ltd. All rights reserved.

  10. Anticipated Benefits of Care (ABC): psychometrics and predictive value in psychiatric disorders.

    PubMed

    Warden, D; Trivedi, M H; Carmody, T J; Gollan, J K; Kashner, T M; Lind, L; Crismon, M L; Rush, A J

    2010-06-01

    Attitudes and expectations about treatment have been associated with symptomatic outcomes, adherence and utilization in patients with psychiatric disorders. No measure of patients' anticipated benefits of treatment on domains of everyday functioning has previously been available. The Anticipated Benefits of Care (ABC) is a new, 10-item questionnaire used to measure patient expectations about the impact of treatment on domains of everyday functioning. The ABC was collected at baseline in adult out-patients with major depressive disorder (MDD) (n=528), bipolar disorder (n=395) and schizophrenia (n=447) in the Texas Medication Algorithm Project (TMAP). Psychometric properties of the ABC were assessed, and the association of ABC scores with treatment response at 3 months was evaluated. Evaluation of the ABC's internal consistency yielded Cronbach's alpha of 0.90-0.92 for patients across disorders. Factor analysis showed that the ABC was unidimensional for all patients and for patients with each disorder. For patients with MDD, lower anticipated benefits of treatment was associated with less symptom improvement and lower odds of treatment response [odds ratio (OR) 0.72, 95% confidence interval (CI) 0.57-0.87, p=0.0011]. There was no association between ABC and symptom improvement or treatment response for patients with bipolar disorder or schizophrenia, possibly because these patients had modest benefits with treatment. The ABC is the first self-report that measures patient expectations about the benefits of treatment on everyday functioning, filling an important gap in available assessments of attitudes and expectations about treatment. The ABC is simple, easy to use, and has acceptable psychometric properties for use in research or clinical settings.

  11. Object Detection Based on Template Matching through Use of Best-So-Far ABC

    PubMed Central

    2014-01-01

    Best-so-far ABC is a modified version of the artificial bee colony (ABC) algorithm used for optimization tasks. This algorithm is one of the swarm intelligence (SI) algorithms proposed in recent literature, in which the results demonstrated that the best-so-far ABC can produce higher quality solutions with faster convergence than either the ordinary ABC or the current state-of-the-art ABC-based algorithm. In this work, we aim to apply the best-so-far ABC-based approach for object detection based on template matching by using the difference between the RGB level histograms corresponding to the target object and the template object as the objective function. Results confirm that the proposed method was successful in both detecting objects and optimizing the time used to reach the solution. PMID:24812556

  12. Characterization of a novel domain ‘GATE’ in the ABC protein DrrA and its role in drug efflux by the DrrAB complex

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhang, Han; Rahman, Sadia; Li, Wen

    2015-03-27

    A novel domain, GATE (Glycine-loop And Transducer Element), is identified in the ABC protein DrrA. This domain shows sequence and structural conservation among close homologs of DrrA as well as distantly-related ABC proteins. Among the highly conserved residues in this domain are three glycines, G215, G221 and G231, of which G215 was found to be critical for stable expression of the DrrAB complex. Other conserved residues, including E201, G221, K227 and G231, were found to be critical for the catalytic and transport functions of the DrrAB transporter. Structural analysis of both the previously published crystal structure of the DrrA homologmore » MalK and the modeled structure of DrrA showed that G215 makes close contacts with residues in and around the Walker A motif, suggesting that these interactions may be critical for maintaining the integrity of the ATP binding pocket as well as the complex. It is also shown that G215A or K227R mutation diminishes some of the atomic interactions essential for ATP catalysis and overall transport function. Therefore, based on both the biochemical and structural analyses, it is proposed that the GATE domain, located outside of the previously identified ATP binding and hydrolysis motifs, is an additional element involved in ATP catalysis. - Highlights: • A novel domain ‘GATE’ is identified in the ABC protein DrrA. • GATE shows high sequence and structural conservation among diverse ABC proteins. • GATE is located outside of the previously studied ATP binding and hydrolysis motifs. • Conserved GATE residues are critical for stability of DrrAB and for ATP catalysis.« less

  13. The ABCs of Sex Ed.

    ERIC Educational Resources Information Center

    Sroka, Stephen R.

    2002-01-01

    Cites statistics on extent of sexually transmitted diseases and pregnancies among adolescents; describes ideological dispute over how to teach sex education; advocates teaching the ABCs of sex education: Abstinence, Be Monogamous, and Condoms. (PKP)

  14. Abrasive wear behavior of heat-treated ABC-silicon carbide

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhang, Xiao Feng; Lee, Gun Y.; Chen, Da

    2002-06-17

    Hot-pressed silicon carbide, containing aluminum, boron, and carbon additives (ABC-SiC), was subjected to three-body and two-body wear testing using diamond abrasives over a range of sizes. In general, the wear resistance of ABC-SiC, with suitable heat treatment, was superior to that of commercial SiC.

  15. The naphthoquinones, vitamin K3 and its structural analog plumbagin, are substrates of the multidrug resistance-linked ABC drug transporter ABCG2

    PubMed Central

    Shukla, Suneet; Wu, Chung-Pu; Nandigama, Krishnamachary; Ambudkar, Suresh V.

    2008-01-01

    Vitamin K3 (Menadione; 2-methyl-1,4-naphthoquinone) is a structural precursor of vitamins K1 and K2 which are essential for blood clotting. The naturally occurring structural analog of this vitamin, plumbagin (5-hydroxy-menadione), is known to modulate cellular proliferation, apoptosis, carcinogenesis, and radioresistance. We, here, report that both vitamin K3 and plumbagin are substrates of the multidrug resistance-linked ATP binding cassette (ABC) drug transporter, ABCG2. Vitamin K3 and plumbagin specifically inhibited the ABCG2-mediated efflux of mitoxantrone, but did not have any effect on the ABCB1-mediated efflux of rhodamine 123. This inhibition of ABCG2 function was due to their interaction at the substrate-binding site(s). They inhibited the binding of [125I]-Iodoarylazidoprazosin (IAAP), a substrate of ABCG2, to this transporter in a concentration-dependent manner with IC50 values of 7.3 and 22.6 μM, respectively, but had no effect on the binding of this photoaffinity analog to ABCB1. Both compounds stimulated ABCG2-mediated ATP hydrolysis and also inhibited the mitoxantrone-stimulated ATPase activity of this transporter, but did not have any significant effect on the ATPase activity of ABCB1. In a cytotoxicity assay, ABCG2-expressing HEK cells were 2.8- and 2.3-fold resistant to plumbagin and vitamin K3, respectively, compared to the control cells, suggesting that they are substrates of this transporter. Collectively, these data demonstrate for the first time that vitamin K3 is a substrate of the ABCG2 transporter. Thus, ABCG2 may have a role in the regulation of vitamin K3 levels in the body. In addition, vitamin K3 and its structural derivative, plumbagin, could potentially be used to modulate ABCG2 function. PMID:18065489

  16. A Manual for Implementation of ABC Video Duplication Projects.

    ERIC Educational Resources Information Center

    Hill, Joseph, Ed.

    The ABC (Appalachian BOCES Consortium) consists of 10 BOCES (Boards of Cooperative Educational Services) which serve the 14 southern counties of New York State designated as Appalachia. Each year since 1974, the ABC has participated in regional video duplication projects, which have yielded a total of nearly 4,000 video titles. The complexity of…

  17. Functional characterization of rainbow trout (Oncorhynchus mykiss) Abcg2a (Bcrp) transporter.

    PubMed

    Zaja, Roko; Popović, Marta; Lončar, Jovica; Smital, Tvrtko

    2016-12-01

    ABCG2 (BCRP - breast cancer resistance protein) belongs to the ATP-binding cassette (ABC) superfamily. It plays an important role in the disposition and elimination of xeno- and endobiotics and/or their metabolites in mammals. Likewise, the protective role of ABC transporters, including Abcg2, has been reported for aquatic organisms. In our previous study we have cloned the full gene sequence of rainbow trout (Oncorhynchus mykiss) Abcg2a and showed its high expression in liver and primary hepatocytes. Based on those insights, the main goal of this study was to perform a detailed functional characterization of trout Abcg2a using insect ovary cells (Spodoptera frugiperda, Sf9) as a heterologous expression system. Membrane vesicles preparations from Sf9 cells were used for the ATPase assay determinations and basic biochemical properties of fish Abcg2a versus human ABCG2 have been compared. A series of 39 physiologically and/or environmentally relevant substances was then tested on interaction with trout Abcg2a and human ABCG2. Correlation analysis reveals highly similar pattern of activation and inhibition. Significant activation of trout Abcg2a ATPase was observed for prazosin, doxorubicine, sildenafil, furosemid, propranolol, fenofibrate and pheophorbide. Pesticides showed either a weak activation (malathione) or strong (endosulfan) to weak (chlorpyrifos, fenoxycarb, DDE) inhibition of trout Abcg2a ATPase while the highest activation was obtained for benzo(a)pyrene, curcumine and testosterone. In conclusion, data from this study offer the first characterization of fish Abcg2a, reveal potent interactors among physiologically or environmentally relevant substances and point to similarities regarding strengths and interactor preferences between human ABCG2 and fish Abcg2a. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. Deciphering the Routes of invasion of Drosophila suzukii by Means of ABC Random Forest.

    PubMed

    Fraimout, Antoine; Debat, Vincent; Fellous, Simon; Hufbauer, Ruth A; Foucaud, Julien; Pudlo, Pierre; Marin, Jean-Michel; Price, Donald K; Cattel, Julien; Chen, Xiao; Deprá, Marindia; François Duyck, Pierre; Guedot, Christelle; Kenis, Marc; Kimura, Masahito T; Loeb, Gregory; Loiseau, Anne; Martinez-Sañudo, Isabel; Pascual, Marta; Polihronakis Richmond, Maxi; Shearer, Peter; Singh, Nadia; Tamura, Koichiro; Xuéreb, Anne; Zhang, Jinping; Estoup, Arnaud

    2017-04-01

    Deciphering invasion routes from molecular data is crucial to understanding biological invasions, including identifying bottlenecks in population size and admixture among distinct populations. Here, we unravel the invasion routes of the invasive pest Drosophila suzukii using a multi-locus microsatellite dataset (25 loci on 23 worldwide sampling locations). To do this, we use approximate Bayesian computation (ABC), which has improved the reconstruction of invasion routes, but can be computationally expensive. We use our study to illustrate the use of a new, more efficient, ABC method, ABC random forest (ABC-RF) and compare it to a standard ABC method (ABC-LDA). We find that Japan emerges as the most probable source of the earliest recorded invasion into Hawaii. Southeast China and Hawaii together are the most probable sources of populations in western North America, which then in turn served as sources for those in eastern North America. European populations are genetically more homogeneous than North American populations, and their most probable source is northeast China, with evidence of limited gene flow from the eastern US as well. All introduced populations passed through bottlenecks, and analyses reveal five distinct admixture events. These findings can inform hypotheses concerning how this species evolved between different and independent source and invasive populations. Methodological comparisons indicate that ABC-RF and ABC-LDA show concordant results if ABC-LDA is based on a large number of simulated datasets but that ABC-RF out-performs ABC-LDA when using a comparable and more manageable number of simulated datasets, especially when analyzing complex introduction scenarios. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  19. A lower isoelectric point increases signal sequence-mediated secretion of recombinant proteins through a bacterial ABC transporter.

    PubMed

    Byun, Hyunjong; Park, Jiyeon; Kim, Sun Chang; Ahn, Jung Hoon

    2017-12-01

    Efficient protein production for industrial and academic purposes often involves engineering microorganisms to produce and secrete target proteins into the culture. Pseudomonas fluorescens has a TliDEF ATP-binding cassette transporter, a type I secretion system, which recognizes C-terminal LARD3 signal sequence of thermostable lipase TliA. Many proteins are secreted by TliDEF in vivo when recombined with LARD3, but there are still others that cannot be secreted by TliDEF even when LARD3 is attached. However, the factors that determine whether or not a recombinant protein can be secreted through TliDEF are still unknown. Here, we recombined LARD3 with several proteins and examined their secretion through TliDEF. We found that the proteins secreted via LARD3 are highly negatively charged with highly-acidic isoelectric points (pI) lower than 5.5. Attaching oligo-aspartate to lower the pI of negatively-charged recombinant proteins improved their secretion, and attaching oligo-arginine to negatively-charged proteins blocked their secretion by LARD3. In addition, negatively supercharged green fluorescent protein (GFP) showed improved secretion, whereas positively supercharged GFP did not secrete. These results disclosed that proteins' acidic pI and net negative charge are major factors that determine their secretion through TliDEF. Homology modeling for TliDEF revealed that TliD dimer forms evolutionarily-conserved positively-charged clusters in its pore and substrate entrance site, which also partially explains the pI dependence of the TliDEF-dependent secretions. In conclusion, lowering the isoelectric point improved LARD3-mediated protein secretion, both widening the range of protein targets for efficient production via secretion and signifying an important aspect of ABC transporter-mediated secretions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. MetaABC--an integrated metagenomics platform for data adjustment, binning and clustering.

    PubMed

    Su, Chien-Hao; Hsu, Ming-Tsung; Wang, Tse-Yi; Chiang, Sufeng; Cheng, Jen-Hao; Weng, Francis C; Kao, Cheng-Yan; Wang, Daryi; Tsai, Huai-Kuang

    2011-08-15

    MetaABC is a metagenomic platform that integrates several binning tools coupled with methods for removing artifacts, analyzing unassigned reads and controlling sampling biases. It allows users to arrive at a better interpretation via series of distinct combinations of analysis tools. After execution, MetaABC provides outputs in various visual formats such as tables, pie and bar charts as well as clustering result diagrams. MetaABC source code and documentation are available at http://bits2.iis.sinica.edu.tw/MetaABC/ CONTACT: dywang@gate.sinica.edu.tw; hktsai@iis.sinica.edu.tw Supplementary data are available at Bioinformatics online.

  1. Phase behavior of model ABC triblock copolymers

    NASA Astrophysics Data System (ADS)

    Chatterjee, Joon

    The phase behavior of poly(isoprene-b-styrene- b-ethylene oxide) (ISO), a model ABC triblock copolymer has been studied. This class of materials exhibit self-assembly, forming a large array of ordered morphologies at length scales of 5-100 nm. The formation of stable three-dimensionally continuous network morphologies is of special interest in this study. Since these nanostructures considerably impact the material properties, fundamental knowledge for designing ABC systems have high technological importance for realizing applications in the areas of nanofabrication, nanoporous media, separation membranes, drug delivery and high surface area catalysts. A comprehensive framework was developed to describe the phase behavior of the ISO triblock copolymers at weak to intermediate segregation strengths spanning a wide range of composition. Phases were characterized through a combination of characterization techniques, including small angle x-ray scattering, dynamic mechanical spectroscopy, transmission electron microscopy, and birefringence measurements. Combined with previous investigations on ISO, six different stable ordered state symmetries have been identified: lamellae (LAM), Fddd orthorhombic network (O70), double gyroid (Q230), alternating gyroid (Q214), hexagonal (HEX), and body-centered cubic (BCC). The phase map was found to be somewhat asymmetric around the fI = fO isopleth. This work provides a guide for theoretical studies and gives insight into the intricate effects of various parameters on the self-assembly of ABC triblock copolymers. Experimental SAXS data evaluated with a simple scattering intensity model show that local mixing varies continuously across the phase map between states of two- and three-domain segregation. Strategies of blending homopolymers with ISO triblock copolymer were employed for studying the swelling properties of a lamellar state. Results demonstrate that lamellar domains swell or shrink depending upon the type of homopolymer that

  2. LrABCF1, a GCN-type ATP-binding cassette transporter from lilium regale, is involved in defense responses against viral and fungal pathogens

    USDA-ARS?s Scientific Manuscript database

    ATP-binding cassette (ABC) transporters are essential for membrane translocation in diverse biological processes, such as plant development and defense response. Here, a general control non-derepressible (GCN)-type ABC transporter gene, designated LrABCF1, was identified from Cucumber mosaic virus (...

  3. Purification and characterization of chondroitinase ABC from Acinetobacter sp. C26.

    PubMed

    Zhu, Changliang; Zhang, Jingliang; Zhang, Jing; Jiang, Yanhui; Shen, Zhaopeng; Guan, Huashi; Jiang, Xiaolu

    2017-02-01

    An extracellular chondroitinase ABC (ChSase ABC, EC 4.2.2.4) produced by cultivating Acinetobacter sp. C26, was purified to homogeneity from the supernatant by ammonium sulfate fractionation, Q-Sepharose Fast Flow and Sephadex G-100 chromatography. The 76kDa enzyme was purified 48.09-fold to homogeneity with specific activity of 348.64U/mg, Using the chondroitin sulfate A (CS-A) as substrate, the maximal reaction rate (Vmax) and Michaelis-Menten constant (Km) of ChSase ABC were found to be 10.471μmol/min/ml and 0.105mg/ml, respectively. The enzyme showed the highest activity at the optimal conditions of pH 6.0 and 42 ∘C, respectively. This enzyme was stable at pH 5-10, 5-9 and 5-7 at 4°C, 37°C and 42°C, respectively. Investigation about thermal stability of ChSase ABC displayed that it was stable at 37°C. ChSase ABC activity was increased in presence of Na + , K + , Mn 2+ , 1,10-phenanthrolin and strongly inhibited by Cu 2+ , Hg 2+ , Al 3+ and SDS. These properties suggested that ChSase ABC from Acinetobacter sp. C26 bring promising prospects in medical and industry applications. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Molecular evolution of the insect chemoreceptor gene superfamily in Drosophila melanogaster.

    PubMed

    Robertson, Hugh M; Warr, Coral G; Carlson, John R

    2003-11-25

    The insect chemoreceptor superfamily in Drosophila melanogaster is predicted to consist of 62 odorant receptor (Or) and 68 gustatory receptor (Gr) proteins, encoded by families of 60 Or and 60 Gr genes through alternative splicing. We include two previously undescribed Or genes and two previously undescribed Gr genes; two previously predicted Or genes are shown to be alternative splice forms. Three polymorphic pseudogenes and one highly defective pseudogene are recognized. Phylogenetic analysis reveals deep branches connecting multiple highly divergent clades within the Gr family, and the Or family appears to be a single highly expanded lineage within the superfamily. The genes are spread throughout the Drosophila genome, with some relatively recently diverged genes still clustered in the genome. The Gr5a gene on the X chromosome, which encodes a receptor for the sugar trehalose, has transposed from one such tandem cluster of six genes at cytological location 64, as has Gr61a, and all eight of these receptors might bind sugars. Analysis of intron evolution suggests that the common ancestor consisted of a long N-terminal exon encoding transmembrane domains 1-5 followed by three exons encoding transmembrane domains 6-7. As many as 57 additional introns have been acquired idiosyncratically during the evolution of the superfamily, whereas the ancestral introns and some of the older idiosyncratic introns have been lost at least 48 times independently. Altogether, these patterns of molecular evolution suggest that this is an ancient superfamily of chemoreceptors, probably dating back at least to the origin of the arthropods.

  5. Molecular evolution of the insect chemoreceptor gene superfamily in Drosophila melanogaster

    PubMed Central

    Robertson, Hugh M.; Warr, Coral G.; Carlson, John R.

    2003-01-01

    The insect chemoreceptor superfamily in Drosophila melanogaster is predicted to consist of 62 odorant receptor (Or) and 68 gustatory receptor (Gr) proteins, encoded by families of 60 Or and 60 Gr genes through alternative splicing. We include two previously undescribed Or genes and two previously undescribed Gr genes; two previously predicted Or genes are shown to be alternative splice forms. Three polymorphic pseudogenes and one highly defective pseudogene are recognized. Phylogenetic analysis reveals deep branches connecting multiple highly divergent clades within the Gr family, and the Or family appears to be a single highly expanded lineage within the superfamily. The genes are spread throughout the Drosophila genome, with some relatively recently diverged genes still clustered in the genome. The Gr5a gene on the X chromosome, which encodes a receptor for the sugar trehalose, has transposed from one such tandem cluster of six genes at cytological location 64, as has Gr61a, and all eight of these receptors might bind sugars. Analysis of intron evolution suggests that the common ancestor consisted of a long N-terminal exon encoding transmembrane domains 1-5 followed by three exons encoding transmembrane domains 6-7. As many as 57 additional introns have been acquired idiosyncratically during the evolution of the superfamily, whereas the ancestral introns and some of the older idiosyncratic introns have been lost at least 48 times independently. Altogether, these patterns of molecular evolution suggest that this is an ancient superfamily of chemoreceptors, probably dating back at least to the origin of the arthropods. PMID:14608037

  6. Common folds and transport mechanisms of secondary active transporters.

    PubMed

    Shi, Yigong

    2013-01-01

    Secondary active transporters exploit the electrochemical potential of solutes to shuttle specific substrate molecules across biological membranes, usually against their concentration gradient. Transporters of different functional families with little sequence similarity have repeatedly been found to exhibit similar folds, exemplified by the MFS, LeuT, and NhaA folds. Observations of multiple conformational states of the same transporter, represented by the LeuT superfamily members Mhp1, AdiC, vSGLT, and LeuT, led to proposals that structural changes are associated with substrate binding and transport. Despite recent biochemical and structural advances, our understanding of substrate recognition and energy coupling is rather preliminary. This review focuses on the common folds and shared transport mechanisms of secondary active transporters. Available structural information generally supports the alternating access model for substrate transport, with variations and extensions made by emerging structural, biochemical, and computational evidence.

  7. ATP Binding Cassette Transporter Mediates Both Heme and Pesticide Detoxification in Tick Midgut Cells

    PubMed Central

    Lara, Flavio Alves; Pohl, Paula C.; Gandara, Ana Caroline; Ferreira, Jessica da Silva; Nascimento-Silva, Maria Clara; Bechara, Gervásio Henrique; Sorgine, Marcos H. F.; Almeida, Igor C.; Vaz, Itabajara da Silva; Oliveira, Pedro L.

    2015-01-01

    In ticks, the digestion of blood occurs intracellularly and proteolytic digestion of hemoglobin takes place in a dedicated type of lysosome, the digest vesicle, followed by transfer of the heme moiety of hemoglobin to a specialized organelle that accumulates large heme aggregates, called hemosomes. In the present work, we studied the uptake of fluorescent metalloporphyrins, used as heme analogs, and amitraz, one of the most regularly used acaricides to control cattle tick infestations, by Rhipicephalus (Boophilus) microplus midgut cells. Both compounds were taken up by midgut cells in vitro and accumulated inside the hemosomes. Transport of both molecules was sensitive to cyclosporine A (CsA), a well-known inhibitor of ATP binding cassette (ABC) transporters. Rhodamine 123, a fluorescent probe that is also a recognized ABC substrate, was similarly directed to the hemosome in a CsA-sensitive manner. Using an antibody against conserved domain of PgP-1-type ABC transporter, we were able to immunolocalize PgP-1 in the digest vesicle membranes. Comparison between two R. microplus strains that were resistant and susceptible to amitraz revealed that the resistant strain detoxified both amitraz and Sn-Pp IX more efficiently than the susceptible strain, a process that was also sensitive to CsA. A transcript containing an ABC transporter signature exhibited 2.5-fold increased expression in the amitraz-resistant strain when compared with the susceptible strain. RNAi-induced down-regulation of this ABC transporter led to the accumulation of metalloporphyrin in the digestive vacuole, interrupting heme traffic to the hemosome. This evidence further confirms that this transcript codes for a heme transporter. This is the first report of heme transport in a blood-feeding organism. While the primary physiological function of the hemosome is to detoxify heme and attenuate its toxicity, we suggest that the use of this acaricide detoxification pathway by ticks may represent a new

  8. Overexpression of Both ERG11 and ABC2 Genes Might Be Responsible for Itraconazole Resistance in Clinical Isolates of Candida krusei

    PubMed Central

    He, Xiaoyuan; Zhao, Mingfeng; Chen, Jinyan; Wu, Rimao; Zhang, Jianlei; Cui, Rui; Jiang, Yanyu; Chen, Jie; Cao, Xiaoli; Xing, Yi; Zhang, Yuchen; Meng, Juanxia; Deng, Qi; Sui, Tao

    2015-01-01

    Objective To study the main molecular mechanisms responsible for itraconazole resistance in clinical isolates of Candida krusei. Methods The 14α-demethylases encoded by ERG11 gene in the 16 C.krusei clinical isolates were amplified by polymerase chain reaction (PCR), and their nucleotide sequences were determined to detect point mutations. Meanwhile, ERG11 and efflux transporters (ABC1 and ABC2) genes were determined by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) for their expression in itraconazole-resistant (R), itraconazole-susceptible dose dependent (SDD) and itraconazole-susceptible (S) C.krusei at the mRNA level. Results We found 7-point mutations in ERG11 gene of all the C.krusei clinical isolates, including 6 synonymous mutations and 1 missense mutation (C44T). However, the missense mutation was found in the three groups. The mRNA levels of ERG11 gene in itraconazole-resistant isolates showed higher expression compared with itraconazole-susceptible dose dependent and itraconazole-susceptible ones (P = 0.015 and P = 0.002 respectively). ABC2 gene mRNA levels in itraconazole-resistant group was significantly higher than the other two groups, and the levels of their expression in the isolates appeared to increase with the decrease of susceptibility to itraconazole (P = 0.007 in SDD compared with S, P = 0.016 in SDD with R, and P<0.001 in S with R respectively). While ABC1 gene presented lower expression in itraconazole resistant strains. However, the mRNA levels of ERG11, ABC1 and ABC2 in a C.krusei (CK10) resistant to both itraconazole and voriconazole were expressed highest in all the itraconazole-resistant isolates. Conclusions There are ERG11 gene polymorphisms in clinical isolates of C.krusei. ERG11 gene mutations may not be involved in the development of itraconazole resistance in C.krusei. ERG11 and ABC2 overexpression might be responsible for the acquired itraconazole resistance of these clinical isolates. PMID

  9. High-throughput screening identifies Ceefourin 1 and Ceefourin 2 as highly selective inhibitors of multidrug resistance protein 4 (MRP4).

    PubMed

    Cheung, Leanna; Flemming, Claudia L; Watt, Fujiko; Masada, Nanako; Yu, Denise M T; Huynh, Tony; Conseil, Gwenaëlle; Tivnan, Amanda; Polinsky, Alexander; Gudkov, Andrei V; Munoz, Marcia A; Vishvanath, Anasuya; Cooper, Dermot M F; Henderson, Michelle J; Cole, Susan P C; Fletcher, Jamie I; Haber, Michelle; Norris, Murray D

    2014-09-01

    Multidrug resistance protein 4 (MRP4/ABCC4), a member of the ATP-binding cassette (ABC) transporter superfamily, is an organic anion transporter capable of effluxing a wide range of physiologically important signalling molecules and drugs. MRP4 has been proposed to contribute to numerous functions in both health and disease; however, in most cases these links remain to be unequivocally established. A major limitation to understanding the physiological and pharmacological roles of MRP4 has been the absence of specific small molecule inhibitors, with the majority of established inhibitors also targeting other ABC transporter family members, or inhibiting the production, function or degradation of important MRP4 substrates. We therefore set out to identify more selective and well tolerated inhibitors of MRP4 that might be used to study the many proposed functions of this transporter. Using high-throughput screening, we identified two chemically distinct small molecules, Ceefourin 1 and Ceefourin 2, that inhibit transport of a broad range of MRP4 substrates, yet are highly selective for MRP4 over other ABC transporters, including P-glycoprotein (P-gp), ABCG2 (Breast Cancer Resistance Protein; BCRP) and MRP1 (multidrug resistance protein 1; ABCC1). Both compounds are more potent MRP4 inhibitors in cellular assays than the most widely used inhibitor, MK-571, requiring lower concentrations to effect a comparable level of inhibition. Furthermore, Ceefourin 1 and Ceefourin 2 have low cellular toxicity, and high microsomal and acid stability. These newly identified inhibitors should be of great value for efforts to better understand the biological roles of MRP4, and may represent classes of compounds with therapeutic application. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Functional Interaction between the Cytoplasmic ABC Protein LptB and the Inner Membrane LptC Protein, Components of the Lipopolysaccharide Transport Machinery in Escherichia coli

    PubMed Central

    Martorana, Alessandra M.; Benedet, Mattia; Maccagni, Elisa A.; Sperandeo, Paola; Villa, Riccardo; Dehò, Gianni

    2016-01-01

    ABSTRACT The assembly of lipopolysaccharide (LPS) in the outer leaflet of the outer membrane (OM) requires the transenvelope Lpt (lipopolysaccharide transport) complex, made in Escherichia coli of seven essential proteins located in the inner membrane (IM) (LptBCFG), periplasm (LptA), and OM (LptDE). At the IM, LptBFG constitute an unusual ATP binding cassette (ABC) transporter, composed by the transmembrane LptFG proteins and the cytoplasmic LptB ATPase, which is thought to extract LPS from the IM and to provide the energy for its export across the periplasm to the cell surface. LptC is a small IM bitopic protein that binds to LptBFG and recruits LptA via its N- and C-terminal regions, and its role in LPS export is not completely understood. Here, we show that the expression level of lptB is a critical factor for suppressing lethality of deletions in the C-terminal region of LptC and the functioning of a hybrid Lpt machinery that carries Pa-LptC, the highly divergent LptC orthologue from Pseudomonas aeruginosa. We found that LptB overexpression stabilizes C-terminally truncated LptC mutant proteins, thereby allowing the formation of a sufficient amount of stable IM complexes to support growth. Moreover, the LptB level seems also critical for the assembly of IM complexes carrying Pa-LptC which is otherwise defective in interactions with the E. coli LptFG components. Overall, our data suggest that LptB and LptC functionally interact and support a model whereby LptB plays a key role in the assembly of the Lpt machinery. IMPORTANCE The asymmetric outer membrane (OM) of Gram-negative bacteria contains in its outer leaflet an unusual glycolipid, the lipopolysaccharide (LPS). LPS largely contributes to the peculiar permeability barrier properties of the OM that prevent the entry of many antibiotics, thus making Gram-negative pathogens difficult to treat. In Escherichia coli the LPS transporter (the Lpt machine) is made of seven essential proteins (LptABCDEFG) that form a

  11. A wheat ABC transporter contributes to both grain formation and mycotoxin tolerance

    PubMed Central

    Walter, Stephanie; Kahla, Amal; Arunachalam, Chanemoughasoundharam; Perochon, Alexandre; Khan, Mojibur R.; Scofield, Steven R.; Doohan, Fiona M.

    2015-01-01

    The mycotoxin deoxynivalenol (DON) acts as a disease virulence factor for Fusarium fungi, and tolerance of DON enhances wheat resistance to Fusarium head blight (FHB) disease. Two variants of an ATP-binding cassette (ABC) family C transporter gene were cloned from DON-treated wheat mRNA, namely TaABCC3.1 and TaABCC3.2. These represent two of three putative genes identified on chromosomes 3A, 3B, and 3D of the wheat genome sequence. Variant TaABCC3.1 represents the DON-responsive transcript previously associated with DON resistance in wheat. PCR-based mapping and in silico sequence analyses located TaABCC3.1 to the short arm of wheat chromosome 3B (not within the FHB resistance quantitative trait locus Fhb1). In silico analyses of microarray data indicated that TaABCC3 genes are expressed in reproductive tissue and roots, and in response to the DON producer Fusarium graminearum. Gene expression studies showed that TaABCC3.1 is activated as part of the early host response to DON and in response to the FHB defence hormone jasmonic acid. Virus-induced gene silencing (VIGS) confirmed that TaABCC3 genes contributed to DON tolerance. VIGS was performed using two independent viral construct applications: one specifically targeted TaABCC3.1 for silencing, while the other targeted this gene and the chromosome 3A homeologue. In both instances, VIGS resulted in more toxin-induced discoloration of spikelets, compared with the DON effects in non-silenced spikelets at 14 d after toxin treatment (≥2.2-fold increase, P<0.05). Silencing by both VIGS constructs enhanced head ripening, and especially so in DON-treated heads. VIGS of TaABCC3 genes also reduced the grain number by more than 28% (P<0.05), both with and without DON treatment, and the effects were greater for the construct that targeted the two homeologues. Hence, DON-responsive TaABCC3 genes warrant further study to determine their potential as disease resistance breeding targets and their function in grain formation

  12. Sustained delivery of thermostabilized chABC enhances axonal sprouting and functional recovery after spinal cord injury.

    PubMed

    Lee, Hyunjung; McKeon, Robert J; Bellamkonda, Ravi V

    2010-02-23

    Chondroitin sulfate proteoglycans (CSPGs) are a major class of axon growth inhibitors that are up-regulated after spinal cord injury (SCI) and contribute to regenerative failure. Chondroitinase ABC (chABC) digests glycosaminoglycan chains on CSPGs and can thereby overcome CSPG-mediated inhibition. But chABC loses its enzymatic activity rapidly at 37 degrees C, necessitating the use of repeated injections or local infusions for a period of days to weeks. These infusion systems are invasive, infection-prone, and clinically problematic. To overcome this limitation, we have thermostabilized chABC and developed a system for its sustained local delivery in vivo, obviating the need for chronically implanted catheters and pumps. Thermostabilized chABC remained active at 37 degrees C in vitro for up to 4 weeks. CSPG levels remained low in vivo up to 6 weeks post-SCI when thermostabilized chABC was delivered by a hydrogel-microtube scaffold system. Axonal growth and functional recovery following the sustained local release of thermostabilized chABC versus a single treatment of unstabilized chABC demonstrated significant differences in CSPG digestion. Animals treated with thermostabilized chABC in combination with sustained neurotrophin-3 delivery showed significant improvement in locomotor function and enhanced growth of cholera toxin B subunit-positive sensory axons and sprouting of serotonergic fibers. Therefore, improving chABC thermostability facilitates minimally invasive, sustained, local delivery of chABC that is potentially effective in overcoming CSPG-mediated regenerative failure. Combination therapy with thermostabilized chABC with neurotrophic factors enhances axonal regrowth, sprouting, and functional recovery after SCI.

  13. SU-E-T-401: Feasibility Study of Using ABC to Gate Lung SBRT Treatment

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Cao, D; Xie, X; Shepard, D

    2014-06-01

    Purpose: The current SBRT treatment techniques include free breathing (FB) SBRT and gated FB SBRT. Gated FB SBRT has smaller target and less lung toxicity with longer treatment time. The recent development of direct connectivity between the ABC and linac allowing for automated beam gating. In this study, we have examined the feasibility of using ABC system to gate the lung SBRT treatment. Methods: A CIRS lung phantom with a 3cm sphere-insert and a moving chest plate was used in this study. Sinusoidal motion was used for the FB pattern. An ABC signal was imported to simulate breath holds. 4D-CTmore » was taken in FB mode and average-intensity-projection (AIP) was used to create FB and 50% gated FB SBRT planning CT. A manually gated 3D CT scan was acquired for ABC gated SBRT planning.An SBRT plan was created for each treatment option. A surface-mapping system was used for 50% gating and ABC system was used for ABC gating. A manually gated CBCT scan was also performed to verify setup. Results: Among three options, the ABC gated plan has the smallest PTV of 35.94cc, which is 35% smaller comparing to that of the FB plan. Consequently, the V20 of the left lung reduced by 15% and 23% comparing to the 50% gated FB and FB plans, respectively. The FB plan took 4.7 minutes to deliver, while the 50% gated FB plan took 18.5 minutes. The ABC gated plan delivery took only 10.6 minutes. A stationary target with 3cm diameter was also obtained from the manually gated CBCT scan. Conclusion: A strategy for ABC gated lung SBRT was developed. ABC gating can significantly reduce the lung toxicity while maintaining the target coverage. Comparing to the 50% gated FB SBRT, ABC gated treatment can also provide less lung toxicity as well as improved delivery efficiency. This research is funded by Elekta.« less

  14. Creating an iPhone Application for Collecting Continuous ABC Data

    ERIC Educational Resources Information Center

    Whiting, Seth W.; Dixon, Mark R.

    2012-01-01

    This paper provides an overview and task analysis for creating a continuous ABC data- collection application using Xcode on a Mac computer. Behavior analysts can program an ABC data collection system, complete with a customized list of target clients, antecedents, behaviors, and consequences to be recorded, and have the data automatically sent to…

  15. ABCB1 and ABCC1-like transporters in immune system cells from sea urchins Echinometra lucunter and Echinus esculentus and oysters Crassostrea gasar and Crassostrea gigas.

    PubMed

    Marques-Santos, Luis Fernando; Hégaret, Hélène; Lima-Santos, Leonardo; Queiroga, Fernando Ramos; da Silva, Patricia Mirella

    2017-11-01

    ABC transporters activity and expression have been associated with the multixenobiotic resistance phenotype (MXR). The activity of these proteins leads to a reduction in the intracellular concentration of several xenobiotics, thus reducing their toxicity. However, little attention has been given to the expression of ABC transporters in marine invertebrates and few studies have investigated their role in immune system cells of sea urchins and shellfish bivalves. The aim of the present study was to investigate the activity of the ABC transporters ABCB1 and ABCC1 in immune system cells of sea urchins (coelomocytes) and oysters (hemocytes) from different climatic regions (Brazil and France). Sea urchins and oysters were collected at Paraíba coast; Brazil (Echinometra lucunter and Crassostrea gasar) and Rade of Brest; France (Echinus esculentus and Crassostrea gigas). Coelomocytes and hemocytes were stained with the ABC transporter substrate calcein-AM and dye accumulation analyzed under flow cytometry. Reversin 205 (ABCB1 transporter blocker) and MK571 (ABCC1 transporter blocker) were used as pharmacological tools to investigate ABC transporter activity. A different pattern of calcein accumulation was observed in coelomocytes: phagocytes > colorless spherulocytes > vibrate cells > red spherulocytes. The treatment with MK571 increased calcein fluorescence levels in coelomocytes from both species. However, reversin 205 treatment was not able to increase calcein fluorescence in E. esculentus coelomocytes. These data suggest that ABCC1-like transporter activity is present in both sea urchin species, but ABCB1-like transporter activity might only be present in E. lucunter coelomocytes. The activity of ABCC1-like transporter was observed in all cell types from both bivalve species. However, reversin 205 only increased calcein accumulation in hyalinocytes of the oyster C. gasar, suggesting the absence of ABCB1-like transporter activity in all other cell types

  16. Laboratory investigation of grouted coupler connection details for ABC bridge projects.

    DOT National Transportation Integrated Search

    2015-08-01

    With an ever increasing desire to utilize accelerated bridge construction (ABC) techniques, it is becoming critical that bridge : designers and contractors have confidence in typical details. The Keg Creek Bridge on US 6 in Iowa was a recent ABC exam...

  17. Automated brainstem co-registration (ABC) for MRI.

    PubMed

    Napadow, Vitaly; Dhond, Rupali; Kennedy, David; Hui, Kathleen K S; Makris, Nikos

    2006-09-01

    Group data analysis in brainstem neuroimaging is predicated on accurate co-registration of anatomy. As the brainstem is comprised of many functionally heterogeneous nuclei densely situated adjacent to one another, relatively small errors in co-registration can manifest in increased variance or decreased sensitivity (or significance) in detecting activations. We have devised a 2-stage automated, reference mask guided registration technique (Automated Brainstem Co-registration, or ABC) for improved brainstem co-registration. Our approach utilized a brainstem mask dataset to weight an automated co-registration cost function. Our method was validated through measurement of RMS error at 12 manually defined landmarks. These landmarks were also used as guides for a secondary manual co-registration option, intended for outlier individuals that may not adequately co-register with our automated method. Our methodology was tested on 10 healthy human subjects and compared to traditional co-registration techniques (Talairach transform and automated affine transform to the MNI-152 template). We found that ABC had a significantly lower mean RMS error (1.22 +/- 0.39 mm) than Talairach transform (2.88 +/- 1.22 mm, mu +/- sigma) and the global affine (3.26 +/- 0.81 mm) method. Improved accuracy was also found for our manual-landmark-guided option (1.51 +/- 0.43 mm). Visualizing individual brainstem borders demonstrated more consistent and uniform overlap for ABC compared to traditional global co-registration techniques. Improved robustness (lower susceptibility to outliers) was demonstrated with ABC through lower inter-subject RMS error variance compared with traditional co-registration methods. The use of easily available and validated tools (AFNI and FSL) for this method should ease adoption by other investigators interested in brainstem data group analysis.

  18. Novel Mutation in the ATP-Binding Cassette Transporter A3 (ABCA3) Encoding Gene Causes Respiratory Distress Syndrome in A Term Newborn in Southwest Iran

    PubMed Central

    Rezaei, Farideh; Shafiei, Mohammad; Shariati, Gholamreza; Dehdashtian, Ali; Mohebbi, Maryam; Galehdari, Hamid

    2016-01-01

    Introduction ABCA3 glycoprotein belongs to the ATP-binding cassette (ABC) superfamily of transporters, which utilize the energy derived from hydrolysis of ATP for the translocation of a wide variety of substrates across the plasma membrane. Mutations in the ABCA3 gene are knowingly causative for fatal surfactant deficiency, particularly respiratory distress syndrome (RDS) in term babies. Case Presentation In this study, Sanger sequencing of the whole ABCA3 gene (NCBI NM_001089) was performed in a neonatal boy with severe RDS. A homozygous mutation has been identified in the patient. Parents were heterozygous for the same missense mutation GGA > AGA at position 202 in exon 6 of the ABCA3 gene (c.604G > A; p.G202R). Furthermore, 70 normal individuals have been analyzed for the mentioned change with negative results. Conclusions Regarding Human Genome Mutation Database (HGMD) and other literature recherche, the detected change is a novel mutation and has not been reported before. Bioinformatics mutation predicting tools prefer it as pathogenic. PMID:27437095

  19. Anger and the ABC model underlying Rational-Emotive Behavior Therapy.

    PubMed

    Ziegler, Daniel J; Smith, Phillip N

    2004-06-01

    The ABC model underlying Ellis's Rational-Emotive Behavior Therapy predicts that people who think more irrationally should display greater trait anger than do people who think less irrationally. This study tested this prediction regarding the ABC model. 186 college students were administered the Survey of Personal Beliefs and the State-Trait Anger Expression Inventory-Second Edition to measure irrational thinking and trait anger, respectively. Students who scored higher on Overall Irrational Thinking and Low Frustration Tolerance scored significantly higher on Trait Anger than did those who scored lower on Overall Irrational Thinking and Low Frustration Tolerance. This indicates support for the ABC model, especially Ellis's construct of irrational beliefs which is central to the model.

  20. Genome-wide identification, phylogenetic analysis, and expression profiles of ATP-binding cassette transporter genes in the oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae).

    PubMed

    Xiao, Lin-Fan; Zhang, Wei; Jing, Tian-Xing; Zhang, Meng-Yi; Miao, Ze-Qing; Wei, Dan-Dan; Yuan, Guo-Rui; Wang, Jin-Jun

    2018-03-01

    The ATP-binding cassette (ABC) is the largest transporter gene family and the genes play key roles in xenobiotic resistance, metabolism, and development of all phyla. However, the specific functions of ABC gene families in insects is unclear. We report a genome-wide identification, phylogenetic, and transcriptional analysis of the ABC genes in the oriental fruit fly, Bactrocera dorsalis (Hendel). We identified a total of 47 ABC genes (BdABCs) from the transcriptomic and genomic databases of B. dorsalis and classified these genes into eight subfamilies (A-H), including 7 ABCAs, 7 ABCBs, 9 ABCCs, 2 ABCDs, 1 ABCE, 3 ABCFs, 15 ABCGs, and 3 ABCHs. Comparative phylogenetic analysis of the ABCs suggests an orthologous relationship between B. dorsalis and other insect species in which these genes have been related to pesticide resistance and essential biological processes. Comparison of transcriptome and relative expression patterns of BdABCs indicated diverse multifunctions within different B. dorsalis tissues. The expression of 4, 10, and 14 BdABCs from 18 BdABCs was significantly upregulated after exposure to LD 50 s of malathion, avermectin, and beta-cypermethrin, respectively. The maximum expression level of most BdABCs (including BdABCFs, BdABCGs, and BdABCHs) occurred at 48h post exposures, whereas BdABCEs peaked at 24h after treatment. Furthermore, RNA interference-mediated suppression of BdABCB7 resulted in increased toxicity of malathion against B. dorsalis. These data suggest that ABC transporter genes might play key roles in xenobiotic metabolism and biosynthesis in B. dorsalis. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Cystic Fibrosis Transmembrane Conductance Regulator (ABCC7) Structure

    PubMed Central

    Hunt, John F.; Wang, Chi; Ford, Robert C.

    2013-01-01

    Structural studies of the cystic fibrosis transmembrane conductance regulator (CFTR) are reviewed. Like many membrane proteins, full-length CFTR has proven to be difficult to express and purify, hence much of the structural data available is for the more tractable, independently expressed soluble domains. Therefore, this chapter covers structural data for individual CFTR domains in addition to the sparser data available for the full-length protein. To set the context for these studies, we will start by reviewing structural information on model proteins from the ATP-binding cassette (ABC) transporter superfamily, to which CFTR belongs. PMID:23378596

  2. The ABC Model and its Applicability to Basal Angiosperms

    PubMed Central

    Soltis, Douglas E.; Chanderbali, André S.; Kim, Sangtae; Buzgo, Matyas; Soltis, Pamela S.

    2007-01-01

    Background Although the flower is the central feature of the angiosperms, little is known of its origin and subsequent diversification. The ABC model has long been the unifying paradigm for floral developmental genetics, but it is based on phylogenetically derived eudicot models. Synergistic research involving phylogenetics, classical developmental studies, genomics and developmental genetics has afforded valuable new insights into floral evolution in general, and the early flower in particular. Scope and Conclusions Genomic studies indicate that basal angiosperms, and by inference the earliest angiosperms, had a rich tool kit of floral genes. Homologues of the ABCE floral organ identity genes are also present in basal angiosperm lineages; however, C-, E- and particularly B-function genes are more broadly expressed in basal lineages. There is no single model of floral organ identity that applies to all angiosperms; there are multiple models that apply depending on the phylogenetic position and floral structure of the group in question. The classic ABC (or ABCE) model may work well for most eudicots. However, modifications are needed for basal eudicots and, the focus of this paper, basal angiosperms. We offer ‘fading borders’ as a testable hypothesis for the basal-most angiosperms and, by inference, perhaps some of the earliest (now extinct) angiosperms. PMID:17616563

  3. Phylodynamic Inference with Kernel ABC and Its Application to HIV Epidemiology.

    PubMed

    Poon, Art F Y

    2015-09-01

    The shapes of phylogenetic trees relating virus populations are determined by the adaptation of viruses within each host, and by the transmission of viruses among hosts. Phylodynamic inference attempts to reverse this flow of information, estimating parameters of these processes from the shape of a virus phylogeny reconstructed from a sample of genetic sequences from the epidemic. A key challenge to phylodynamic inference is quantifying the similarity between two trees in an efficient and comprehensive way. In this study, I demonstrate that a new distance measure, based on a subset tree kernel function from computational linguistics, confers a significant improvement over previous measures of tree shape for classifying trees generated under different epidemiological scenarios. Next, I incorporate this kernel-based distance measure into an approximate Bayesian computation (ABC) framework for phylodynamic inference. ABC bypasses the need for an analytical solution of model likelihood, as it only requires the ability to simulate data from the model. I validate this "kernel-ABC" method for phylodynamic inference by estimating parameters from data simulated under a simple epidemiological model. Results indicate that kernel-ABC attained greater accuracy for parameters associated with virus transmission than leading software on the same data sets. Finally, I apply the kernel-ABC framework to study a recent outbreak of a recombinant HIV subtype in China. Kernel-ABC provides a versatile framework for phylodynamic inference because it can fit a broader range of models than methods that rely on the computation of exact likelihoods. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  4. Abc Amino Acids: Design, Synthesis, and Properties of New Photoelastic Amino Acids

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Standaert, Robert F; Park, Dr Seung Bum

    2006-01-01

    Photoisomerizable amino acids provide a direct avenue to the experimental manipulation of bioactive polypeptides, potentially allowing real-time, remote control of biological systems and enabling useful applications in nanobiotechnology. Herein, we report a new class of photoisomerizable amino acids intended to cause pronounced expansion and contraction in the polypeptide backbone, i.e., to be photoelastic. These compounds, termed Abc amino acids, employ a photoisomerizable azobiphenyl chromophore to control the relative disposition of aminomethyl and carboxyl substituents. Molecular modeling of nine Abc isomers led to the identification of one with particularly attractive properties, including the ability to induce contractions up to 13A inmore » the backbone upon transa?cis photoisomerization. This isomer, designated mpAbc, has substituents at meta and para positions on the inner (azo-linked) and outer rings, respectively. An efficient synthesis of Fmoc-protected mpAbc was executed in which the biaryl components were formed via Suzuki couplings and the azo linkage was formed via amine/nitroso condensation; protected forms of three other Abc isomers were prepared similarly. A decapeptide incorporating mpAbc was synthesized by conventional solid-phase methods and displayed characteristic azobenzene photochemical behavior with optimal conversion to the cis isomer at 360 nm and a thermal cisa?trans half life of 100 min. at 80 AoC.« less

  5. Membrane Transporters: Structure, Function and Targets for Drug Design

    NASA Astrophysics Data System (ADS)

    Ravna, Aina W.; Sager, Georg; Dahl, Svein G.; Sylte, Ingebrigt

    Current therapeutic drugs act on four main types of molecular targets: enzymes, receptors, ion channels and transporters, among which a major part (60-70%) are membrane proteins. This review discusses the molecular structures and potential impact of membrane transporter proteins on new drug discovery. The three-dimensional (3D) molecular structure of a protein contains information about the active site and possible ligand binding, and about evolutionary relationships within the protein family. Transporters have a recognition site for a particular substrate, which may be used as a target for drugs inhibiting the transporter or acting as a false substrate. Three groups of transporters have particular interest as drug targets: the major facilitator superfamily, which includes almost 4000 different proteins transporting sugars, polyols, drugs, neurotransmitters, metabolites, amino acids, peptides, organic and inorganic anions and many other substrates; the ATP-binding cassette superfamily, which plays an important role in multidrug resistance in cancer chemotherapy; and the neurotransmitter:sodium symporter family, which includes the molecular targets for some of the most widely used psychotropic drugs. Recent technical advances have increased the number of known 3D structures of membrane transporters, and demonstrated that they form a divergent group of proteins with large conformational flexibility which facilitates transport of the substrate.

  6. Step 2: Know Your Diabetes ABCs

    MedlinePlus

    ... please turn JavaScript on. Feature: Type 2 Diabetes Step 2: Know Your Diabetes ABCs Past Issues / Fall ... 2 Diabetes" Articles Diabetes Is Serious But Manageable / Step 1: Learn About Diabetes / Step 2: Know Your ...

  7. The ABCs of Student Engagement

    ERIC Educational Resources Information Center

    Parsons, Seth A.; Nuland, Leila Richey; Parsons, Allison Ward

    2014-01-01

    Student engagement is an important consideration for teachers and administrators because it is explicitly associated with achievement. What the authors call the ABC's of engagement they outline as: Affective engagement, Behavioral engagement, and Cognitive engagement. They also present "Three Things Every Teacher Needs to Know about…

  8. Direct Spectroscopic Detection of ATP Turnover Reveals Mechanistic Divergence of ABC Exporters.

    PubMed

    Collauto, Alberto; Mishra, Smriti; Litvinov, Aleksei; Mchaourab, Hassane S; Goldfarb, Daniella

    2017-08-01

    We have applied high-field (W-band) pulse electron-nuclear double resonance (ENDOR) and electron-electron double resonance (ELDOR)-detected nuclear magnetic resonance (EDNMR) to characterize the coordination sphere of the Mn 2+ co-factor in the nucleotide binding sites (NBSs) of ABC transporters. MsbA and BmrCD are two efflux transporters hypothesized to represent divergent catalytic mechanisms. Our results reveal distinct coordination of Mn 2+ to ATP and transporter residues in the consensus and degenerate NBSs of BmrCD. In contrast, the coordination of Mn 2+ at the two NBSs of MsbA is similar, which provides a mechanistic rationale for its higher rate constant of ATP hydrolysis relative to BmrCD. Direct detection of vanadate ion, trapped in a high-energy post-hydrolysis intermediate, further supports the notion of asymmetric hydrolysis by the two NBSs of BmrCD. The integrated spectroscopic approach presented here, which link energy input to conformational dynamics, can be applied to a variety of systems powered by ATP turnover. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Homologues of insulinase, a new superfamily of metalloendopeptidases.

    PubMed Central

    Rawlings, N D; Barrett, A J

    1991-01-01

    On the basis of a statistical analysis of an alignment of the amino acid sequences, a new superfamily of metalloendopeptidases is proposed, consisting of human insulinase, Escherichia coli protease III and mitochondrial processing endopeptidases from Saccharomyces and Neurospora. These enzymes do not contain the 'HEXXH' consensus sequence found in all previously recognized zinc metalloendopeptidases. PMID:2025223

  10. Implication of RuvABC and RecG in homologous recombination in Streptomyces ambofaciens.

    PubMed

    Hoff, Grégory; Bertrand, Claire; Piotrowski, Emilie; Thibessard, Annabelle; Leblond, Pierre

    2017-01-01

    Most bacterial organisms rely on homologous recombination to repair DNA double-strand breaks and for the post-replicative repair of DNA single-strand gaps. Homologous recombination can be divided into three steps: (i) a pre-synaptic step in which the DNA 3'-OH ends are processed, (ii) a recA-dependent synaptic step allowing the invasion of an intact copy and the formation of Holliday junctions, and (iii) a post-synaptic step consisting of migration and resolution of these junctions. Currently, little is known about factors involved in homologous recombination, especially for the post-synaptic step. In Escherichia coli, branch migration and resolution are performed by the RuvABC complex, but could also rely on the RecG helicase in a redundant manner. In this study, we show that recG and ruvABC are well-conserved among Streptomyces. ΔruvABC, ΔrecG and ΔruvABC ΔrecG mutant strains were constructed. ΔruvABC ΔrecG is only slightly affected by exposure to DNA damage (UV). We also show that conjugational recombination decreases in the absence of RuvABC and RecG, but that intra-chromosomal recombination is not affected. These data suggest that RuvABC and RecG are indeed involved in homologous recombination in Streptomyces ambofaciens and that alternative factors are able to take over Holliday junction in Streptomyces. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  11. Parents' Perspectives on Braille Literacy: Results from the ABC Braille Study

    ERIC Educational Resources Information Center

    Kamei-Hannan, Cheryl; Sacks, Sharon Zell

    2012-01-01

    Introduction: Parents who were the primary caretakers of children in the Alphabetic and Contracted Braille Study (ABC Braille Study) revealed their perspectives about braille literacy. Methods: A 30-item questionnaire was constructed by the ABC Braille research team, and researchers conducted telephone interviews with 31 parents who were the…

  12. Early Observations of the Type Ia Supernova iPTF 16abc

    NASA Astrophysics Data System (ADS)

    Miller, Adam; iPTF Collaboration

    2018-01-01

    Early observations of Type Ia supernovae (SNe) provide a unique probe of their progenitor systems and explosion physics. Here, we report the intermediate Palomar Transient Factory (iPTF) discovery of an extraordinarily young SN Ia, iPTF 16abc. By fitting a power law to our early light curve, we infer that first light for the SN only occurred 0.15 +0.15-0.07 d before our first detection. In the ~24 hr after discovery, iPTF 16abc rose by ~2 mag, following a near-linear rise in flux for ~3 d. Strong C II absorption is detected in the early spectra of iPTF 16abc, before disappearing after ~7 d. Unlike the extensively-observed Type Ia SN 2011fe, the (B-V)_0 colors of iPTF 16abc are blue and nearly constant in the days after explosion. We show that our early observations of iPTF 16abc cannot be explained by either SN shock breakout and the associated, subsequent cooling, or the SN ejecta colliding with a stellar companion. Instead, we argue that the early characteristics of iPTF 16abc, including: (i) the rapid, near-linear rise, (ii) the non-evolving blue colors, and (iii) the strong absorption from ionized carbon, are the result of either vigorous mixing of radioactive-Ni in the SN ejecta, or ejecta interaction with diffuse material, or a combination of the two. In the next few years, dozens of very young normal SNe Ia will be discovered, and observations similar to those presented here will constrain the white dwarf explosion mechanism.

  13. ABC-F Proteins Mediate Antibiotic Resistance through Ribosomal Protection.

    PubMed

    Sharkey, Liam K R; Edwards, Thomas A; O'Neill, Alex J

    2016-03-22

    Members of the ABC-F subfamily of ATP-binding cassette proteins mediate resistance to a broad array of clinically important antibiotic classes that target the ribosome of Gram-positive pathogens. The mechanism by which these proteins act has been a subject of long-standing controversy, with two competing hypotheses each having gained considerable support: antibiotic efflux versus ribosomal protection. Here, we report on studies employing a combination of bacteriological and biochemical techniques to unravel the mechanism of resistance of these proteins, and provide several lines of evidence that together offer clear support to the ribosomal protection hypothesis. Of particular note, we show that addition of purified ABC-F proteins to anin vitrotranslation assay prompts dose-dependent rescue of translation, and demonstrate that such proteins are capable of displacing antibiotic from the ribosomein vitro To our knowledge, these experiments constitute the first direct evidence that ABC-F proteins mediate antibiotic resistance through ribosomal protection.IMPORTANCEAntimicrobial resistance ranks among the greatest threats currently facing human health. Elucidation of the mechanisms by which microorganisms resist the effect of antibiotics is central to understanding the biology of this phenomenon and has the potential to inform the development of new drugs capable of blocking or circumventing resistance. Members of the ABC-F family, which includelsa(A),msr(A),optr(A), andvga(A), collectively yield resistance to a broader range of clinically significant antibiotic classes than any other family of resistance determinants, although their mechanism of action has been controversial since their discovery 25 years ago. Here we present the first direct evidence that proteins of the ABC-F family act to protect the bacterial ribosome from antibiotic-mediated inhibition. Copyright © 2016 Sharkey et al.

  14. Differential catalytic promiscuity of the alkaline phosphatase superfamily bimetallo core reveals mechanistic features underlying enzyme evolution

    DOE PAGES

    Sunden, Fanny; AlSadhan, Ishraq; Lyubimov, Artem; ...

    2017-10-25

    Members of enzyme superfamilies specialize in different reactions but often exhibit catalytic promiscuity for one another's reactions, consistent with catalytic promiscuity as an important driver in the evolution of new enzymes. Wanting to understand how catalytic promiscuity and other factors may influence evolution across a superfamily, we turned to the well-studied alkaline phosphatase (AP) superfamily, comparing three of its members, two evolutionarily distinct phosphatases and a phosphodiesterase. Here, we mutated distinguishing active-site residues to generate enzymes that had a common Zn 2+ bimetallo core but little sequence similarity and different auxiliary domains. We then tested the catalytic capabilities of thesemore » pruned enzymes with a series of substrates. A substantial rate enhancement of ~1011-fold for both phosphate mono- and diester hydrolysis by each enzyme indicated that the Zn 2+ bimetallo core is an effective mono/di-esterase generalist and that the bimetallo cores were not evolutionarily tuned to prefer their cognate reactions. In contrast, our pruned enzymes were ineffective sulfatases, and this limited promiscuity may have provided a driving force for founding the distinct one-metal-ion branch that contains all known AP superfamily sulfatases. Finally, our pruned enzymes exhibited 10 7–10 8-fold phosphotriesterase rate enhancements, despite absence of such enzymes within the AP superfamily. We speculate that the superfamily active-site architecture involved in nucleophile positioning prevents accommodation of the additional triester substituent. Overall, we suggest that catalytic promiscuity, and the ease or difficulty of remodeling and building onto existing protein scaffolds, have greatly influenced the course of enzyme evolution. Uncovering principles and properties of enzyme function, promiscuity, and repurposing provides lessons for engineering new enzymes.« less

  15. Differential catalytic promiscuity of the alkaline phosphatase superfamily bimetallo core reveals mechanistic features underlying enzyme evolution

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sunden, Fanny; AlSadhan, Ishraq; Lyubimov, Artem

    Members of enzyme superfamilies specialize in different reactions but often exhibit catalytic promiscuity for one another's reactions, consistent with catalytic promiscuity as an important driver in the evolution of new enzymes. Wanting to understand how catalytic promiscuity and other factors may influence evolution across a superfamily, we turned to the well-studied alkaline phosphatase (AP) superfamily, comparing three of its members, two evolutionarily distinct phosphatases and a phosphodiesterase. Here, we mutated distinguishing active-site residues to generate enzymes that had a common Zn 2+ bimetallo core but little sequence similarity and different auxiliary domains. We then tested the catalytic capabilities of thesemore » pruned enzymes with a series of substrates. A substantial rate enhancement of ~1011-fold for both phosphate mono- and diester hydrolysis by each enzyme indicated that the Zn 2+ bimetallo core is an effective mono/di-esterase generalist and that the bimetallo cores were not evolutionarily tuned to prefer their cognate reactions. In contrast, our pruned enzymes were ineffective sulfatases, and this limited promiscuity may have provided a driving force for founding the distinct one-metal-ion branch that contains all known AP superfamily sulfatases. Finally, our pruned enzymes exhibited 10 7–10 8-fold phosphotriesterase rate enhancements, despite absence of such enzymes within the AP superfamily. We speculate that the superfamily active-site architecture involved in nucleophile positioning prevents accommodation of the additional triester substituent. Overall, we suggest that catalytic promiscuity, and the ease or difficulty of remodeling and building onto existing protein scaffolds, have greatly influenced the course of enzyme evolution. Uncovering principles and properties of enzyme function, promiscuity, and repurposing provides lessons for engineering new enzymes.« less

  16. Self-assembly in the ferritin nano-cage protein superfamily.

    PubMed

    Zhang, Yu; Orner, Brendan P

    2011-01-01

    Protein self-assembly, through specific, high affinity, and geometrically constraining protein-protein interactions, can control and lead to complex cellular nano-structures. Establishing an understanding of the underlying principles that govern protein self-assembly is not only essential to appreciate the fundamental biological functions of these structures, but could also provide a basis for their enhancement for nano-material applications. The ferritins are a superfamily of well studied proteins that self-assemble into hollow cage-like structures which are ubiquitously found in both prokaryotes and eukaryotes. Structural studies have revealed that many members of the ferritin family can self-assemble into nano-cages of two types. Maxi-ferritins form hollow spheres with octahedral symmetry composed of twenty-four monomers. Mini-ferritins, on the other hand, are tetrahedrally symmetric, hollow assemblies composed of twelve monomers. This review will focus on the structure of members of the ferritin superfamily, the mechanism of ferritin self-assembly and the structure-function relations of these proteins.

  17. Brassboard Astrometric Beam Combiner (ABC) Development for the Space Interferometry Mission (SIM)

    NASA Technical Reports Server (NTRS)

    Jeganathan, Muthu; Kuan, Gary; Rud, Mike; Lin, Sean; Sutherland, Kristen; Moore, James; An, Xin

    2008-01-01

    The Astrometric Beam Combiner (ABC) is a critical element of the Space Interferometry Mission (SIM) that performs three key functions: coherently combine starlight from two siderostats; individually detect starlight for angle tracking; and disperse and detect the interferometric fringes. In addition, the ABC contains: a stimulus, cornercubes and shutters for in-orbit calibration; several tip/tilt mirror mechanisms for in-orbit alignment; and internal metrology beam launcher for pathlength monitoring. The detailed design of the brassboard ABC (which has the form, fit and function of the flight unit) is complete, procurement of long-lead items is underway, and assembly and testing is expected to be completed in Spring 2009. In this paper, we present the key requirements for the ABC, details of the completed optical and mechanical design as well as plans for assembly and alignment.

  18. fbpABC gene cluster in Neisseria meningitidis is transcribed as an operon.

    PubMed

    Khun, H H; Deved, V; Wong, H; Lee, B C

    2000-12-01

    The neisserial fbpABC locus has been proposed to constitute a single transcriptional unit. To confirm this operonic arrangement, transcription assays using reverse transcriptase PCR amplification were conducted with Neisseria meningitidis. The presence of fbpAB and fbpBC transcripts obtained by priming cDNA synthesis with an fbpC-sequence-specific oligonucleotide indicates that fbpABC is organized as a single expression unit. The ratio of fbpA to fbpABC mRNA was approximately between 10- to 20-fold, as determined by real-time quantitative PCR.

  19. fbpABC Gene Cluster in Neisseria meningitidis Is Transcribed as an Operon

    PubMed Central

    Khun, Heng H.; Deved, Vinay; Wong, Howard; Lee, B. Craig

    2000-01-01

    The neisserial fbpABC locus has been proposed to constitute a single transcriptional unit. To confirm this operonic arrangement, transcription assays using reverse transcriptase PCR amplification were conducted with Neisseria meningitidis. The presence of fbpAB and fbpBC transcripts obtained by priming cDNA synthesis with an fbpC-sequence-specific oligonucleotide indicates that fbpABC is organized as a single expression unit. The ratio of fbpA to fbpABC mRNA was approximately between 10- to 20-fold, as determined by real-time quantitative PCR. PMID:11083849

  20. Purification and biochemical characterization of NpABCG5/NpPDR5, a plant pleiotropic drug resistance transporter expressed in Nicotiana tabacum BY-2 suspension cells.

    PubMed

    Toussaint, Frédéric; Pierman, Baptiste; Bertin, Aurélie; Lévy, Daniel; Boutry, Marc

    2017-05-04

    Pleiotropic drug resistance (PDR) transporters belong to the ABCG subfamily of ATP-binding cassette (ABC) transporters and are involved in the transport of various molecules across plasma membranes. During evolution, PDR genes appeared independently in fungi and in plants from a duplication of a half-size ABC gene. The enzymatic properties of purified PDR transporters from yeast have been characterized. This is not the case for any plant PDR transporter, or, incidentally, for any purified plant ABC transporter. Yet, plant PDR transporters play important roles in plant physiology such as hormone signaling or resistance to pathogens or herbivores. Here, we describe the expression, purification, enzymatic characterization and 2D analysis by electron microscopy of NpABCG5/NpPDR5 from Nicotiana plumbaginifolia , which has been shown to be involved in the plant defense against herbivores. We constitutively expressed NpABCG5/NpPDR5, provided with a His-tag in a homologous system: suspension cells from Nicotiana tabacum (Bright Yellow 2 line). NpABCG5/NpPDR5 was targeted to the plasma membrane and was solubilized by dodecyl maltoside and purified by Ni-affinity chromatography. The ATP-hydrolyzing specific activity (27 nmol min -1  mg -1 ) was stimulated seven-fold in the presence of 0.1% asolectin. Electron microscopy analysis indicated that NpABCG5/NpPDR5 is monomeric and with dimensions shorter than those of known ABC transporters. Enzymatic data (optimal pH and sensitivity to inhibitors) confirmed that plant and fungal PDR transporters have different properties. These data also show that N. tabacum suspension cells are a convenient host for the purification and biochemical characterization of ABC transporters. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

  1. Evolutionary history, structural features and biochemical diversity of the NlpC/P60 superfamily of enzymes.

    PubMed

    Anantharaman, Vivek; Aravind, L

    2003-01-01

    Peptidoglycan is hydrolyzed by a diverse set of enzymes during bacterial growth, development and cell division. The N1pC/P60 proteins define a family of cell-wall peptidases that are widely represented in various bacterial lineages. Currently characterized members are known to hydrolyze D-gamma-glutamyl-meso-diaminopimelate or N-acetylmuramate-L-alanine linkages. Detailed analysis of the N1pC/P60 peptidases showed that these proteins define a large superfamily encompassing several diverse groups of proteins. In addition to the well characterized P60-like proteins, this superfamily includes the AcmB/LytN and YaeF/YiiX families of bacterial proteins, the amidase domain of bacterial and kinetoplastid glutathionylspermidine synthases (GSPSs), and several proteins from eukaryotes, phages, poxviruses, positive-strand RNA viruses, and certain archaea. The eukaryotic members include lecithin retinol acyltransferase (LRAT), nematode developmental regulator Egl-26, and candidate tumor suppressor H-rev107. These eukaryotic proteins, along with the bacterial YaeF/poxviral G6R family, show a circular permutation of the catalytic domain. We identified three conserved residues, namely a cysteine, a histidine and a polar residue, that are involved in the catalytic activities of this superfamily. Evolutionary analysis of this superfamily shows that it comprises four major families, with diverse domain architectures in each of them. Several related, but distinct, catalytic activities, such as murein degradation, acyl transfer and amide hydrolysis, have emerged in the N1pC/P60 superfamily. The three conserved catalytic residues of this superfamily are shown to be equivalent to the catalytic triad of the papain-like thiol peptidases. The predicted structural features indicate that the N1pC/P60 enzymes contain a fold similar to the papain-like peptidases, transglutaminases and arylamine acetyltransferases.

  2. Regulatory crosstalk by protein kinases on CFTR trafficking and activity

    NASA Astrophysics Data System (ADS)

    Farinha, Carlos Miguel; Swiatecka-Urban, Agnieszka; Brautigan, David; Jordan, Peter

    2016-01-01

    Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) is a member of the ATP binding cassette (ABC) transporter superfamily that functions as a cAMP-activated chloride ion channel in fluid-transporting epithelia. There is abundant evidence that CFTR activity (i.e. channel opening and closing) is regulated by protein kinases and phosphatases via phosphorylation and dephosphorylation. Here, we review recent evidence for the role of protein kinases in regulation of CFTR delivery to and retention in the plasma membrane. We review this information in a broader context of regulation of other transporters by protein kinases because the overall functional output of transporters involves the integrated control of both their number at the plasma membrane and their specific activity. While many details of the regulation of intracellular distribution of CFTR and other transporters remain to be elucidated, we hope that this review will motivate research providing new insights into how protein kinases control membrane transport to impact health and disease.

  3. ABCC1, an ATP Binding Cassette Protein from Grape Berry, Transports Anthocyanidin 3-O-Glucosides[W][OA

    PubMed Central

    Francisco, Rita Maria; Regalado, Ana; Ageorges, Agnès; Burla, Bo J.; Bassin, Barbara; Eisenach, Cornelia; Zarrouk, Olfa; Vialet, Sandrine; Marlin, Thérèse; Chaves, Maria Manuela; Martinoia, Enrico; Nagy, Réka

    2013-01-01

    Accumulation of anthocyanins in the exocarp of red grapevine (Vitis vinifera) cultivars is one of several events that characterize the onset of grape berry ripening (véraison). Despite our thorough understanding of anthocyanin biosynthesis and regulation, little is known about the molecular aspects of their transport. The participation of ATP binding cassette (ABC) proteins in vacuolar anthocyanin transport has long been a matter of debate. Here, we present biochemical evidence that an ABC protein, ABCC1, localizes to the tonoplast and is involved in the transport of glucosylated anthocyanidins. ABCC1 is expressed in the exocarp throughout berry development and ripening, with a significant increase at véraison (i.e., the onset of ripening). Transport experiments using microsomes isolated from ABCC1-expressing yeast cells showed that ABCC1 transports malvidin 3-O-glucoside. The transport strictly depends on the presence of GSH, which is cotransported with the anthocyanins and is sensitive to inhibitors of ABC proteins. By exposing anthocyanin-producing grapevine root cultures to buthionine sulphoximine, which reduced GSH levels, a decrease in anthocyanin concentration is observed. In conclusion, we provide evidence that ABCC1 acts as an anthocyanin transporter that depends on GSH without the formation of an anthocyanin-GSH conjugate. PMID:23723325

  4. Candida Drug Resistance Protein 1, a Major Multidrug ATP Binding Cassette Transporter of Candida albicans, Translocates Fluorescent Phospholipids in a Reconstituted System†

    PubMed Central

    Shukla, Sudhanshu; Rai, Versha; Saini, Preeti; Banerjee, Dibyendu; Menon, Anant K.; Prasad, Rajendra

    2008-01-01

    Candida albicans drug resistance protein 1 (Cdr1p), an ATP-dependent drug efflux pump, contributes to multidrug resistance in Candida-infected immunocompromised patients. Previous cell-based assays suggested that Cdr1p also acts as a phospholipid translocator. To investigate this, we reconstituted purified Cdr1p into sealed membrane vesicles. Comparison of the ATPase activities of sealed and permeabilized proteoliposomes indicated that Cdr1p was asymmetrically reconstituted such that ~70% of the molecules had their ATP binding sites accessible to the extravesicular space. Fluorescent glycerophospholipids were incorporated into the outer leaflet of the proteoliposomes, and their transport into the inner leaflet was tracked with a quenching assay using membrane-impermeant dithionite. We observed ATP-dependent transport of the fluorescent lipids into the inner leaflet of the vesicles. With ~6 molecules of Cdr1p per vesicle on average, the half-time to reach the maximal extent of transport was ~15 min. Transport was reduced in vesicles reconstituted with Cdr1p variants with impaired ATPase activity and could be competed out to different levels by a molar excess of drugs such as fluconazole and miconazole that are known to be effluxed by Cdr1p. Transport was not affected by ampicillin, a compound that is not effluxed by Cdr1p. Our results suggest a direct link between the ability of Cdr1p to translocate fluorescent phospholipids and efflux drugs. We note that only a few members of the ABC superfamily of Candida have a well-defined role as drug exporters; thus, lipid translocation mediated by Cdr1p could reflect its cellular function. PMID:17924650

  5. Expression, purification and characterization of GAPDH-ChSase ABC I from Proteus vulgaris in Escherichia coli.

    PubMed

    Li, Ye; Chen, Zhenya; Zhou, Zhao; Yuan, Qipeng

    2016-12-01

    Chondroitinases (ChSases) are a family of polysaccharide lyases that can depolymerize high molecular weight chondroitin sulfate (CS) and dermatan sulfate (DS). In this study, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which is stably expressed in different cells like normal cells and cancer cells and the expression is relatively insensitive to experimental conditions, was expressed as a fusion protein with ChSase ABC I. Results showed that the expression level and enzyme activity of GAPDH-ChSase ABC I were about 2.2 and 3.0 times higher than those of ChSase ABC I. By optimization of fermentation conditions, higher productivity of ChSase ABC I was achieved as 880 ± 61 IU/g wet cell weight compared with the reported ones. The optimal temperature and pH of GAPDH-ChSase ABC I were 40 °C and 7.5, respectively. GAPDH-ChSase ABC I had a kcat/Km of 131 ± 4.1 L/μmol s and the catalytic efficiency was decreased as compared to ChSase ABC I. The relative activity of GAPDH-ChSase ABC I remained 89% after being incubated at 30 °C for 180 min and the thermostability of ChSase ABC I was enhanced by GAPDH when it was incubated at 30, 35, 40 and 45 °C. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. ABC estimation of unit costs for emergency department services.

    PubMed

    Holmes, R L; Schroeder, R E

    1996-04-01

    Rapid evolution of the health care industry forces managers to make cost-effective decisions. Typical hospital cost accounting systems do not provide emergency department managers with the information needed, but emergency department settings are so complex and dynamic as to make the more accurate activity-based costing (ABC) system prohibitively expensive. Through judicious use of the available traditional cost accounting information and simple computer spreadsheets. managers may approximate the decision-guiding information that would result from the much more costly and time-consuming implementation of ABC.

  7. Minireview: SLCO and ABC Transporters: A Role for Steroid Transport in Prostate Cancer Progression

    PubMed Central

    Cho, Eunpi; Montgomery, R. Bruce

    2014-01-01

    Androgens play a critical role in the development and progression of prostate cancer (PCa), and androgen deprivation therapy via surgical or medical castration is front-line therapy for patients with advanced PCa. However, intratumoral testosterone levels are elevated in metastases from patients with castration-resistant disease, and residual intratumoral androgens have been implicated in mediating ligand-dependent mechanisms of androgen receptor activation. The source of residual tissue androgens present despite castration has not been fully elucidated, but proposed mechanisms include uptake and conversion of adrenal androgens, such as dehdroepiandrosterone to testosterone and dihydrotestosterone, or de novo androgen synthesis from cholesterol or progesterone precursors. In this minireview, we discuss the emerging evidence that suggests a role for specific transporters in mediating transport of steroids into or out of prostate cells, thereby influencing intratumoral androgen levels and PCa development and progression. We focus on the solute carrier and ATP binding cassette gene families, which have the most published data for a role in PCa-related steroid transport, and review the potential impact of genetic variation on steroid transport activity and PCa outcomes. Continued assessment of transport activity in PCa models and human tumor tissue is needed to better delineate the different roles these transporters play in physiologic and neoplastic settings, and in order to determine whether targeting the uptake of steroid substrates by specific transporters may be a clinically feasible therapeutic strategy. PMID:25147980

  8. The ABCs of Activity-Based Costing: A Cost Containment and Reallocation Tool.

    ERIC Educational Resources Information Center

    Turk, Frederick J.

    1992-01-01

    This article describes activity-based costing (ABC) and how this tool may help management understand the costs of major activities and identify possible alternatives. Also discussed are the traditional costing systems used by higher education and ways of applying ABC to higher education. (GLR)

  9. Applying Activity Based Costing (ABC) Method to Calculate Cost Price in Hospital and Remedy Services

    PubMed Central

    Rajabi, A; Dabiri, A

    2012-01-01

    Background Activity Based Costing (ABC) is one of the new methods began appearing as a costing methodology in the 1990’s. It calculates cost price by determining the usage of resources. In this study, ABC method was used for calculating cost price of remedial services in hospitals. Methods: To apply ABC method, Shahid Faghihi Hospital was selected. First, hospital units were divided into three main departments: administrative, diagnostic, and hospitalized. Second, activity centers were defined by the activity analysis method. Third, costs of administrative activity centers were allocated into diagnostic and operational departments based on the cost driver. Finally, with regard to the usage of cost objectives from services of activity centers, the cost price of medical services was calculated. Results: The cost price from ABC method significantly differs from tariff method. In addition, high amount of indirect costs in the hospital indicates that capacities of resources are not used properly. Conclusion: Cost price of remedial services with tariff method is not properly calculated when compared with ABC method. ABC calculates cost price by applying suitable mechanisms but tariff method is based on the fixed price. In addition, ABC represents useful information about the amount and combination of cost price services. PMID:23113171

  10. Expression, purification and thermostability of MBP-chondroitinase ABC I from Proteus vulgaris.

    PubMed

    Chen, Zhenya; Li, Ye; Yuan, Qipeng

    2015-01-01

    Chondroitinase ABC I (ChSase ABC I) which can degrade chondroitin sulfate (CS) and other glycosaminoglycan to oligosaccharide or unsaturated disaccharide, was fusionally expressed with maltose-binding protein (MBP) in Escherichia coli BL21(DE3) (E. coli BL21(DE3)) and purified for the first time in this study. The result showed that the productivity of recombinant MBP-ChSase ABC I was 3180 IU/(L fermentation liquor) with CS A as substrate, and the productivity might be the highest level when compared to the reported ones. The specific activity of recombinant MBP-ChSase ABC I was 76 IU/(mg protein) after purification. The Vmax, Km and kcat were 18.7 ± 0.3 μmol/Ls, 73.1 ± 4.1 μmol/L and 586.7 ± 10.8 s(-1), respectively. Enzyme activity of the purified enzyme remained about 78% after 210 min when the enzyme incubated at 30 °C. This study introduces a rapid method for highly expressing ChSase ABC I, and the method could be adopted in the process of industrial production. Furthermore the investigation of thermostability might lead to an important guide in clinical treatment. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Soybean (Glycine max) expansin gene superfamily origins: segmental and tandem duplication events followed by divergent selection among subfamilies

    PubMed Central

    2014-01-01

    Background Expansins are plant cell wall loosening proteins that are involved in cell enlargement and a variety of other developmental processes. The expansin superfamily contains four subfamilies; namely, α-expansin (EXPA), β-expansin (EXPB), expansin-like A (EXLA), and expansin-like B (EXLB). Although the genome sequencing of soybeans is complete, our knowledge about the pattern of expansion and evolutionary history of soybean expansin genes remains limited. Results A total of 75 expansin genes were identified in the soybean genome, and grouped into four subfamilies based on their phylogenetic relationships. Structural analysis revealed that the expansin genes are conserved in each subfamily, but are divergent among subfamilies. Furthermore, in soybean and Arabidopsis, the expansin gene family has been mainly expanded through tandem and segmental duplications; however, in rice, segmental duplication appears to be the dominant process that generates this superfamily. The transcriptome atlas revealed notable differential expression in either transcript abundance or expression patterns under normal growth conditions. This finding was consistent with the differential distribution of the cis-elements in the promoter region, and indicated wide functional divergence in this superfamily. Moreover, some critical amino acids that contribute to functional divergence and positive selection were detected. Finally, site model and branch-site model analysis of positive selection indicated that the soybean expansin gene superfamily is under strong positive selection, and that divergent selection constraints might have influenced the evolution of the four subfamilies. Conclusion This study demonstrated that the soybean expansin gene superfamily has expanded through tandem and segmental duplication. Differential expression indicated wide functional divergence in this superfamily. Furthermore, positive selection analysis revealed that divergent selection constraints might have

  12. External pH modulates EAG superfamily K+ channels through EAG-specific acidic residues in the voltage sensor

    PubMed Central

    Kazmierczak, Marcin; Zhang, Xiaofei; Chen, Bihan; Mulkey, Daniel K.; Shi, Yingtang; Wagner, Paul G.; Pivaroff-Ward, Kendra; Sassic, Jessica K.; Bayliss, Douglas A.

    2013-01-01

    The Ether-a-go-go (EAG) superfamily of voltage-gated K+ channels consists of three functionally distinct gene families (Eag, Elk, and Erg) encoding a diverse set of low-threshold K+ currents that regulate excitability in neurons and muscle. Previous studies indicate that external acidification inhibits activation of three EAG superfamily K+ channels, Kv10.1 (Eag1), Kv11.1 (Erg1), and Kv12.1 (Elk1). We show here that Kv10.2, Kv12.2, and Kv12.3 are similarly inhibited by external protons, suggesting that high sensitivity to physiological pH changes is a general property of EAG superfamily channels. External acidification depolarizes the conductance–voltage (GV) curves of these channels, reducing low threshold activation. We explored the mechanism of this high pH sensitivity in Kv12.1, Kv10.2, and Kv11.1. We first examined the role of acidic voltage sensor residues that mediate divalent cation block of voltage activation in EAG superfamily channels because protons reduce the sensitivity of Kv12.1 to Zn2+. Low pH similarly reduces Mg2+ sensitivity of Kv10.1, and we found that the pH sensitivity of Kv11.1 was greatly attenuated at 1 mM Ca2+. Individual neutralizations of a pair of EAG-specific acidic residues that have previously been implicated in divalent block of diverse EAG superfamily channels greatly reduced the pH response in Kv12.1, Kv10.2, and Kv11.1. Our results therefore suggest a common mechanism for pH-sensitive voltage activation in EAG superfamily channels. The EAG-specific acidic residues may form the proton-binding site or alternatively are required to hold the voltage sensor in a pH-sensitive conformation. The high pH sensitivity of EAG superfamily channels suggests that they could contribute to pH-sensitive K+ currents observed in vivo. PMID:23712551

  13. External pH modulates EAG superfamily K+ channels through EAG-specific acidic residues in the voltage sensor.

    PubMed

    Kazmierczak, Marcin; Zhang, Xiaofei; Chen, Bihan; Mulkey, Daniel K; Shi, Yingtang; Wagner, Paul G; Pivaroff-Ward, Kendra; Sassic, Jessica K; Bayliss, Douglas A; Jegla, Timothy

    2013-06-01

    The Ether-a-go-go (EAG) superfamily of voltage-gated K(+) channels consists of three functionally distinct gene families (Eag, Elk, and Erg) encoding a diverse set of low-threshold K(+) currents that regulate excitability in neurons and muscle. Previous studies indicate that external acidification inhibits activation of three EAG superfamily K(+) channels, Kv10.1 (Eag1), Kv11.1 (Erg1), and Kv12.1 (Elk1). We show here that Kv10.2, Kv12.2, and Kv12.3 are similarly inhibited by external protons, suggesting that high sensitivity to physiological pH changes is a general property of EAG superfamily channels. External acidification depolarizes the conductance-voltage (GV) curves of these channels, reducing low threshold activation. We explored the mechanism of this high pH sensitivity in Kv12.1, Kv10.2, and Kv11.1. We first examined the role of acidic voltage sensor residues that mediate divalent cation block of voltage activation in EAG superfamily channels because protons reduce the sensitivity of Kv12.1 to Zn(2+). Low pH similarly reduces Mg(2+) sensitivity of Kv10.1, and we found that the pH sensitivity of Kv11.1 was greatly attenuated at 1 mM Ca(2+). Individual neutralizations of a pair of EAG-specific acidic residues that have previously been implicated in divalent block of diverse EAG superfamily channels greatly reduced the pH response in Kv12.1, Kv10.2, and Kv11.1. Our results therefore suggest a common mechanism for pH-sensitive voltage activation in EAG superfamily channels. The EAG-specific acidic residues may form the proton-binding site or alternatively are required to hold the voltage sensor in a pH-sensitive conformation. The high pH sensitivity of EAG superfamily channels suggests that they could contribute to pH-sensitive K(+) currents observed in vivo.

  14. MntABC and MntH Contribute to Systemic Staphylococcus aureus Infection by Competing with Calprotectin for Nutrient Manganese

    PubMed Central

    Kehl-Fie, Thomas E.; Zhang, Yaofang; Moore, Jessica L.; Farrand, Allison J.; Hood, M. Indriati; Rathi, Subodh; Chazin, Walter J.; Caprioli, Richard M.

    2013-01-01

    During infection, vertebrates limit access to manganese and zinc, starving invading pathogens, such as Staphylococcus aureus, of these essential metals in a process termed “nutritional immunity.” The manganese and zinc binding protein calprotectin is a key component of the nutrient-withholding response, and mice lacking this protein do not sequester manganese from S. aureus liver abscesses. One potential mechanism utilized by S. aureus to minimize host-imposed manganese and zinc starvation is the expression of the metal transporters MntABC and MntH. We performed transcriptional analyses of both mntA and mntH, which revealed increased expression of both systems in response to calprotectin treatment. MntABC and MntH compete with calprotectin for manganese, which enables S. aureus growth and retention of manganese-dependent superoxide dismutase activity. Loss of MntABC and MntH results in reduced staphylococcal burdens in the livers of wild-type but not calprotectin-deficient mice, suggesting that these systems promote manganese acquisition during infection. During the course of these studies, we observed that metal content and the importance of calprotectin varies between murine organs, and infection leads to profound changes in the anatomical distribution of manganese and zinc. In total, these studies provide insight into the mechanisms utilized by bacteria to evade host-imposed nutrient metal starvation and the critical importance of restricting manganese availability during infection. PMID:23817615

  15. The ATP-binding cassette transporter Cbc (choline/betaine/carnitine) recruits multiple substrate-binding proteins with strong specificity for distinct quaternary ammonium compounds

    PubMed Central

    Chen, Chiliang; Malek, Adel A.; Wargo, Matthew J.; Hogan, Deborah A.; Beattie, Gwyn A.

    2017-01-01

    Summary We identified a choline, betaine and carnitine transporter, designated Cbc, from Pseudomonas syringae and Pseudomonas aeruginosa that is unusual among members of the ATP-binding cassette (ABC) transporter family in its use of multiple periplasmic substrate-binding proteins (SBPs) that are highly specific for their substrates. The SBP encoded by the cbcXWV operon, CbcX, binds choline with a high affinity (Km, 2.6 μM) and, although it also binds betaine (Km, 24.2 μM), CbcXWV-mediated betaine uptake did not occur in the presence of choline. The CbcX orthologue ChoX from Sinorhizobium meliloti was similar to CbcX in these binding properties. The core transporter CbcWV also interacts with the carnitine-specific SBP CaiX (Km, 24 μM) and the betaine-specific SBP BetX (Km, 0.6 μM). Unlike most ABC transporter loci, caiX, betX and cbcXWV are separated in the genome. CaiX-mediated carnitine uptake was reduced by CbcX and BetX only when they were bound by their individual ligands, providing the first in vivo evidence for a higher affinity for ligand-bound than ligand-free SBPs by an ABC transporter. These studies demonstrate not only that the Cbc transporter serves as a useful model for exploring ABC transporter component interactions, but also that the orphan SBP genes common to bacterial genomes can encode functional SBPs. PMID:19919675

  16. The ATP-binding cassette transporter Cbc (choline/betaine/carnitine) recruits multiple substrate-binding proteins with strong specificity for distinct quaternary ammonium compounds.

    PubMed

    Chen, Chiliang; Malek, Adel A; Wargo, Matthew J; Hogan, Deborah A; Beattie, Gwyn A

    2010-01-01

    We identified a choline, betaine and carnitine transporter, designated Cbc, from Pseudomonas syringae and Pseudomonas aeruginosa that is unusual among members of the ATP-binding cassette (ABC) transporter family in its use of multiple periplasmic substrate-binding proteins (SBPs) that are highly specific for their substrates. The SBP encoded by the cbcXWV operon, CbcX, binds choline with a high affinity (K(m), 2.6 microM) and, although it also binds betaine (K(m), 24.2 microM), CbcXWV-mediated betaine uptake did not occur in the presence of choline. The CbcX orthologue ChoX from Sinorhizobium meliloti was similar to CbcX in these binding properties. The core transporter CbcWV also interacts with the carnitine-specific SBP CaiX (K(m), 24 microM) and the betaine-specific SBP BetX (K(m), 0.6 microM). Unlike most ABC transporter loci, caiX, betX and cbcXWV are separated in the genome. CaiX-mediated carnitine uptake was reduced by CbcX and BetX only when they were bound by their individual ligands, providing the first in vivo evidence for a higher affinity for ligand-bound than ligand-free SBPs by an ABC transporter. These studies demonstrate not only that the Cbc transporter serves as a useful model for exploring ABC transporter component interactions, but also that the orphan SBP genes common to bacterial genomes can encode functional SBPs.

  17. The major facilitator superfamily transporter Knq1p modulates boron homeostasis in Kluyveromyces lactis.

    PubMed

    Svrbicka, Alexandra; Toth Hervay, Nora; Gbelska, Yvetta

    2016-03-01

    Boron is an essential micronutrient for living cells, yet its excess causes toxicity. To date, the mechanisms of boron toxicity are poorly understood. Recently, the ScATR1 gene has been identified encoding the main boron efflux pump in Saccharomyces cerevisiae. In this study, we analyzed the ScATR1 ortholog in Kluyveromyces lactis--the KNQ1 gene, to understand whether it participates in boron stress tolerance. We found that the KNQ1 gene, encoding a permease belonging to the major facilitator superfamily, is required for K. lactis boron tolerance. Deletion of the KNQ1 gene led to boron sensitivity and its overexpression increased K. lactis boron tolerance. The KNQ1 expression was induced by boron and the intracellular boron concentration was controlled by Knq1p. The KNQ1 promoter contains two putative binding motifs for the AP-1-like transcription factor KlYap1p playing a central role in oxidative stress defense. Our results indicate that the induction of the KNQ1 expression requires the presence of KlYap1p and that Knq1p like its ortholog ScAtr1p in S. cerevisiae functions as a boron efflux pump providing boron resistance in K. lactis.

  18. MDR1 and BCRP Transporter-Mediated Drug-Drug Interaction between Rilpivirine and Abacavir and Effect on Intestinal Absorption

    PubMed Central

    Reznicek, Josef; Ceckova, Martina; Ptackova, Zuzana; Martinec, Ondrej; Tupova, Lenka; Cerveny, Lukas

    2017-01-01

    ABSTRACT Rilpivirine (TMC278) is a highly potent nonnucleoside reverse transcriptase inhibitor (NNRTI) representing an effective component of combination antiretroviral therapy (cART) in the treatment of HIV-positive patients. Many antiretroviral drugs commonly used in cART are substrates of ATP-binding cassette (ABC) and/or solute carrier (SLC) drug transporters and, therefore, are prone to pharmacokinetic drug-drug interactions (DDIs). The aim of our study was to evaluate rilpivirine interactions with abacavir and lamivudine on selected ABC and SLC transporters in vitro and assess its importance for pharmacokinetics in vivo. Using accumulation assays in MDCK cells overexpressing selected ABC or SLC drug transporters, we revealed rilpivirine as a potent inhibitor of MDR1 and BCRP, but not MRP2, OCT1, OCT2, or MATE1. Subsequent transport experiments across monolayers of MDCKII-MDR1, MDCKII-BCRP, and Caco-2 cells demonstrated that rilpivirine inhibits MDR1- and BCRP-mediated efflux of abacavir and increases its transmembrane transport. In vivo experiments in male Wistar rats confirmed inhibition of MDR1/BCRP in the small intestine, leading to a significant increase in oral bioavailability of abacavir. In conclusion, rilpivirine inhibits MDR1 and BCRP transporters and may affect pharmacokinetic behavior of concomitantly administered substrates of these transporters, such as abacavir. PMID:28696229

  19. The novel ABC transporter ABCH1 is a potential target for RNAi-based insect pest control and resistance management.

    PubMed

    Guo, Zhaojiang; Kang, Shi; Zhu, Xun; Xia, Jixing; Wu, Qingjun; Wang, Shaoli; Xie, Wen; Zhang, Youjun

    2015-09-03

    Insect pests cause serious crop damage and develop high-level resistance to chemical insecticides and Bacillus thuringiensis (Bt) insecticidal Cry toxins. A new promising approach for controlling them and overcoming this resistance is RNA interference (RNAi). The RNAi-based insect control strategy depends on the selection of suitable target genes. In this study, we cloned and characterized a novel ABC transporter gene PxABCH1 in diamondback moth, Plutella xylostella (L.). Phylogenetic analysis showed that PxABCH1 is closely related to ABCA and ABCG subfamily members. Spatial-temporal expression detection revealed that PxABCH1 was expressed in all tissues and developmental stages, and highest expressed in head and male adult. Midgut sequence variation and expression analyses of PxABCH1 in all the susceptible and Bt-resistant P. xylostella strains and the functional analysis by sublethal RNAi demonstrated that Cry1Ac resistance was independent of this gene. Silencing of PxABCH1 by a relatively high dose of dsRNA dramatically reduced its expression and resulted in larval and pupal lethal phenotypes in both susceptible and Cry1Ac-resistant P. xylostella strains. To our knowledge, this study provides the first insight into ABCH1 in lepidopterans and reveals it as an excellent target for RNAi-based insect pest control and resistance management.

  20. The novel ABC transporter ABCH1 is a potential target for RNAi-based insect pest control and resistance management

    PubMed Central

    Guo, Zhaojiang; Kang, Shi; Zhu, Xun; Xia, Jixing; Wu, Qingjun; Wang, Shaoli; Xie, Wen; Zhang, Youjun

    2015-01-01

    Insect pests cause serious crop damage and develop high-level resistance to chemical insecticides and Bacillus thuringiensis (Bt) insecticidal Cry toxins. A new promising approach for controlling them and overcoming this resistance is RNA interference (RNAi). The RNAi-based insect control strategy depends on the selection of suitable target genes. In this study, we cloned and characterized a novel ABC transporter gene PxABCH1 in diamondback moth, Plutella xylostella (L.). Phylogenetic analysis showed that PxABCH1 is closely related to ABCA and ABCG subfamily members. Spatial-temporal expression detection revealed that PxABCH1 was expressed in all tissues and developmental stages, and highest expressed in head and male adult. Midgut sequence variation and expression analyses of PxABCH1 in all the susceptible and Bt-resistant P. xylostella strains and the functional analysis by sublethal RNAi demonstrated that Cry1Ac resistance was independent of this gene. Silencing of PxABCH1 by a relatively high dose of dsRNA dramatically reduced its expression and resulted in larval and pupal lethal phenotypes in both susceptible and Cry1Ac-resistant P. xylostella strains. To our knowledge, this study provides the first insight into ABCH1 in lepidopterans and reveals it as an excellent target for RNAi-based insect pest control and resistance management. PMID:26333918

  1. Multidrug ATP-binding cassette transporters are essential for hepatic development of Plasmodium sporozoites.

    PubMed

    Rijpma, Sanna R; van der Velden, Maarten; González-Pons, Maria; Annoura, Takeshi; van Schaijk, Ben C L; van Gemert, Geert-Jan; van den Heuvel, Jeroen J M W; Ramesar, Jai; Chevalley-Maurel, Severine; Ploemen, Ivo H; Khan, Shahid M; Franetich, Jean-Francois; Mazier, Dominique; de Wilt, Johannes H W; Serrano, Adelfa E; Russel, Frans G M; Janse, Chris J; Sauerwein, Robert W; Koenderink, Jan B; Franke-Fayard, Blandine M

    2016-03-01

    Multidrug resistance-associated proteins (MRPs) belong to the C-family of ATP-binding cassette (ABC) transport proteins and are known to transport a variety of physiologically important compounds and to be involved in the extrusion of pharmaceuticals. Rodent malaria parasites encode a single ABC transporter subfamily C protein, whereas human parasites encode two: MRP1 and MRP2. Although associated with drug resistance, their biological function and substrates remain unknown. To elucidate the role of MRP throughout the parasite life cycle, Plasmodium berghei and Plasmodium falciparum mutants lacking MRP expression were generated. P. berghei mutants lacking expression of the single MRP as well as P. falciparum mutants lacking MRP1, MRP2 or both proteins have similar blood stage growth kinetics and drug-sensitivity profiles as wild type parasites. We show that MRP1-deficient parasites readily invade primary human hepatocytes and develop into mature liver stages. In contrast, both P. falciparum MRP2-deficient parasites and P. berghei mutants lacking MRP protein expression abort in mid to late liver stage development, failing to produce mature liver stages. The combined P. berghei and P. falciparum data are the first demonstration of a critical role of an ABC transporter during Plasmodium liver stage development. © 2015 John Wiley & Sons Ltd.

  2. Influence of multidrug resistance and drug transport proteins on chemotherapy drug metabolism.

    PubMed

    Joyce, Helena; McCann, Andrew; Clynes, Martin; Larkin, Annemarie

    2015-05-01

    Chemotherapy involving the use of anticancer drugs remains an important strategy in the overall management of patients with metastatic cancer. Acquisition of multidrug resistance remains a major impediment to successful chemotherapy. Drug transporters in cell membranes and intracellular drug metabolizing enzymes contribute to the resistance phenotype and determine the pharmacokinetics of anticancer drugs in the body. ATP-binding cassette (ABC) transporters mediate the transport of endogenous metabolites and xenobiotics including cytotoxic drugs out of cells. Solute carrier (SLC) transporters mediate the influx of cytotoxic drugs into cells. This review focuses on the substrate interaction of these transporters, on their biology and what role they play together with drug metabolizing enzymes in eliminating therapeutic drugs from cells. The majority of anticancer drugs are substrates for the ABC transporter and SLC transporter families. Together, these proteins have the ability to control the influx and the efflux of structurally unrelated chemotherapeutic drugs, thereby modulating the intracellular drug concentration. These interactions have important clinical implications for chemotherapy because ultimately they determine therapeutic efficacy, disease progression/relapse and the success or failure of patient treatment.

  3. mRMR-ABC: A Hybrid Gene Selection Algorithm for Cancer Classification Using Microarray Gene Expression Profiling

    PubMed Central

    Alshamlan, Hala; Badr, Ghada; Alohali, Yousef

    2015-01-01

    An artificial bee colony (ABC) is a relatively recent swarm intelligence optimization approach. In this paper, we propose the first attempt at applying ABC algorithm in analyzing a microarray gene expression profile. In addition, we propose an innovative feature selection algorithm, minimum redundancy maximum relevance (mRMR), and combine it with an ABC algorithm, mRMR-ABC, to select informative genes from microarray profile. The new approach is based on a support vector machine (SVM) algorithm to measure the classification accuracy for selected genes. We evaluate the performance of the proposed mRMR-ABC algorithm by conducting extensive experiments on six binary and multiclass gene expression microarray datasets. Furthermore, we compare our proposed mRMR-ABC algorithm with previously known techniques. We reimplemented two of these techniques for the sake of a fair comparison using the same parameters. These two techniques are mRMR when combined with a genetic algorithm (mRMR-GA) and mRMR when combined with a particle swarm optimization algorithm (mRMR-PSO). The experimental results prove that the proposed mRMR-ABC algorithm achieves accurate classification performance using small number of predictive genes when tested using both datasets and compared to previously suggested methods. This shows that mRMR-ABC is a promising approach for solving gene selection and cancer classification problems. PMID:25961028

  4. mRMR-ABC: A Hybrid Gene Selection Algorithm for Cancer Classification Using Microarray Gene Expression Profiling.

    PubMed

    Alshamlan, Hala; Badr, Ghada; Alohali, Yousef

    2015-01-01

    An artificial bee colony (ABC) is a relatively recent swarm intelligence optimization approach. In this paper, we propose the first attempt at applying ABC algorithm in analyzing a microarray gene expression profile. In addition, we propose an innovative feature selection algorithm, minimum redundancy maximum relevance (mRMR), and combine it with an ABC algorithm, mRMR-ABC, to select informative genes from microarray profile. The new approach is based on a support vector machine (SVM) algorithm to measure the classification accuracy for selected genes. We evaluate the performance of the proposed mRMR-ABC algorithm by conducting extensive experiments on six binary and multiclass gene expression microarray datasets. Furthermore, we compare our proposed mRMR-ABC algorithm with previously known techniques. We reimplemented two of these techniques for the sake of a fair comparison using the same parameters. These two techniques are mRMR when combined with a genetic algorithm (mRMR-GA) and mRMR when combined with a particle swarm optimization algorithm (mRMR-PSO). The experimental results prove that the proposed mRMR-ABC algorithm achieves accurate classification performance using small number of predictive genes when tested using both datasets and compared to previously suggested methods. This shows that mRMR-ABC is a promising approach for solving gene selection and cancer classification problems.

  5. Structure of the periplasmic adaptor protein from a major facilitator superfamily (MFS) multidrug efflux pump.

    PubMed

    Hinchliffe, Philip; Greene, Nicholas P; Paterson, Neil G; Crow, Allister; Hughes, Colin; Koronakis, Vassilis

    2014-08-25

    Periplasmic adaptor proteins are key components of bacterial tripartite efflux pumps. The 2.85 Å resolution structure of an MFS (major facilitator superfamily) pump adaptor, Aquifex aeolicus EmrA, shows linearly arranged α-helical coiled-coil, lipoyl, and β-barrel domains, but lacks the fourth membrane-proximal domain shown in other pumps to interact with the inner membrane transporter. The adaptor α-hairpin, which binds outer membrane TolC, is exceptionally long at 127 Å, and the β-barrel contains a conserved disordered loop. The structure extends the view of adaptors as flexible, modular components that mediate diverse pump assembly, and suggests that in MFS tripartite pumps a hexamer of adaptors could provide a periplasmic seal. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  6. ATP and AMP Mutually Influence Their Interaction with the ATP-binding Cassette (ABC) Adenylate Kinase Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) at Separate Binding Sites*

    PubMed Central

    Randak, Christoph O.; Dong, Qian; Ver Heul, Amanda R.; Elcock, Adrian H.; Welsh, Michael J.

    2013-01-01

    Cystic fibrosis transmembrane conductance regulator (CFTR) is an anion channel in the ATP-binding cassette (ABC) transporter protein family. In the presence of ATP and physiologically relevant concentrations of AMP, CFTR exhibits adenylate kinase activity (ATP + AMP ⇆ 2 ADP). Previous studies suggested that the interaction of nucleotide triphosphate with CFTR at ATP-binding site 2 is required for this activity. Two other ABC proteins, Rad50 and a structural maintenance of chromosome protein, also have adenylate kinase activity. All three ABC adenylate kinases bind and hydrolyze ATP in the absence of other nucleotides. However, little is known about how an ABC adenylate kinase interacts with ATP and AMP when both are present. Based on data from non-ABC adenylate kinases, we hypothesized that ATP and AMP mutually influence their interaction with CFTR at separate binding sites. We further hypothesized that only one of the two CFTR ATP-binding sites is involved in the adenylate kinase reaction. We found that 8-azidoadenosine 5′-triphosphate (8-N3-ATP) and 8-azidoadenosine 5′-monophosphate (8-N3-AMP) photolabeled separate sites in CFTR. Labeling of the AMP-binding site with 8-N3-AMP required the presence of ATP. Conversely, AMP enhanced photolabeling with 8-N3-ATP at ATP-binding site 2. The adenylate kinase active center probe P1,P5-di(adenosine-5′) pentaphosphate interacted simultaneously with an AMP-binding site and ATP-binding site 2. These results show that ATP and AMP interact with separate binding sites but mutually influence their interaction with the ABC adenylate kinase CFTR. They further indicate that the active center of the adenylate kinase comprises ATP-binding site 2. PMID:23921386

  7. Exploring and Expanding the Fatty-Acid-Binding Protein Superfamily in Fasciola Species.

    PubMed

    Morphew, Russell M; Wilkinson, Toby J; Mackintosh, Neil; Jahndel, Veronika; Paterson, Steve; McVeigh, Paul; Abbas Abidi, Syed M; Saifullah, Khalid; Raman, Muthusamy; Ravikumar, Gopalakrishnan; LaCourse, James; Maule, Aaron; Brophy, Peter M

    2016-09-02

    The liver flukes Fasciola hepatica and F. gigantica infect livestock worldwide and threaten food security with climate change and problematic control measures spreading disease. Fascioliasis is also a foodborne disease with up to 17 million humans infected. In the absence of vaccines, treatment depends on triclabendazole (TCBZ), and overuse has led to widespread resistance, compromising future TCBZ control. Reductionist biology from many laboratories has predicted new therapeutic targets. To this end, the fatty-acid-binding protein (FABP) superfamily has proposed multifunctional roles, including functions intersecting vaccine and drug therapy, such as immune modulation and anthelmintic sequestration. Research is hindered by a lack of understanding of the full FABP superfamily complement. Although discovery studies predicted FABPs as promising vaccine candidates, it is unclear if uncharacterized FABPs are more relevant for vaccine formulations. We have coupled genome, transcriptome, and EST data mining with proteomics and phylogenetics to reveal a liver fluke FABP superfamily of seven clades: previously identified clades I-III and newly identified clades IV-VII. All new clade FABPs were analyzed using bioinformatics and cloned from both liver flukes. The extended FABP data set will provide new study tools to research the role of FABPs in parasite biology and as therapy targets.

  8. GPS-ABC radiated chamber testing overview and results : GPS-ABC Workshop VI : RTCA Washington, DC, March 30, 2017.

    DOT National Transportation Integrated Search

    2017-03-30

    This presentation, which was given during the GPS-ABC Workshop VI in Washington, DC on March 30, 2017 details the authors' radiated testing protocols and results. GPS receiver testing was carried out April 25-29, 2016 at the Army : Research Laborator...

  9. Hfq Influences Multiple Transport Systems and Virulence in the Plant Pathogen Agrobacterium tumefaciens

    PubMed Central

    Wilms, Ina; Möller, Philip; Stock, Anna-Maria; Gurski, Rosemarie; Lai, Erh-Min

    2012-01-01

    The Hfq protein mediates gene regulation by small RNAs (sRNAs) in about 50% of all bacteria. Depending on the species, phenotypic defects of an hfq mutant range from mild to severe. Here, we document that the purified Hfq protein of the plant pathogen and natural genetic engineer Agrobacterium tumefaciens binds to the previously described sRNA AbcR1 and its target mRNA atu2422, which codes for the substrate binding protein of an ABC transporter taking up proline and γ-aminobutyric acid (GABA). Several other ABC transporter components were overproduced in an hfq mutant compared to their levels in the parental strain, suggesting that Hfq plays a major role in controlling the uptake systems and metabolic versatility of A. tumefaciens. The hfq mutant showed delayed growth, altered cell morphology, and reduced motility. Although the DNA-transferring type IV secretion system was produced, tumor formation by the mutant strain was attenuated, demonstrating an important contribution of Hfq to plant transformation by A. tumefaciens. PMID:22821981

  10. Structural Insights into the Transport Mechanism of the Human Sodium-dependent Lysophosphatidylcholine Transporter MFSD2A*♦

    PubMed Central

    Quek, Debra Q. Y.; Nguyen, Long N.; Fan, Hao; Silver, David L.

    2016-01-01

    Major facilitator superfamily domain containing 2A (MFSD2A) was recently characterized as a sodium-dependent lysophosphatidylcholine transporter expressed at the blood-brain barrier endothelium. It is the primary route for importation of docosohexaenoic acid and other long-chain fatty acids into fetal and adult brain and is essential for mouse and human brain growth and function. Remarkably, MFSD2A is the first identified major facilitator superfamily member that uniquely transports lipids, implying that MFSD2A harbors unique structural features and transport mechanism. Here, we present three three-dimensional structural models of human MFSD2A derived by homology modeling using MelB- and LacY-based crystal structures and refined by biochemical analysis. All models revealed 12 transmembrane helices and connecting loops and represented the partially outward-open, outward-partially occluded, and inward-open states of the transport cycle. In addition to a conserved sodium-binding site, three unique structural features were identified as follows: a phosphate headgroup binding site, a hydrophobic cleft to accommodate a hydrophobic hydrocarbon tail, and three sets of ionic locks that stabilize the outward-open conformation. Ligand docking studies and biochemical assays identified Lys-436 as a key residue for transport. It is seen forming a salt bridge with the negative charge on the phosphate headgroup. Importantly, MFSD2A transported structurally related acylcarnitines but not a lysolipid without a negative charge, demonstrating the necessity of a negatively charged headgroup interaction with Lys-436 for transport. These findings support a novel transport mechanism by which lysophosphatidylcholines are “flipped” within the transporter cavity by pivoting about Lys-436 leading to net transport from the outer to the inner leaflet of the plasma membrane. PMID:26945070

  11. A test of the ABC model underlying rational emotive behavior therapy.

    PubMed

    Ziegler, Daniel J; Leslie, Yvonne M

    2003-02-01

    The ABC model underlying Ellis's Rational Emotive Behavior Therapy predicts that people who think more irrationally should respond to daily stressors or hassles differently than do people who think less irrationally. This study tested this aspect of the ABC model. 192 college students were administered the Survey of Personal Beliefs and the Hassles Scale to measure irrational thinking and daily hassles, respectively. Students who scored higher on overall irrational thinking reported a significantly higher frequency of hassles than did those who scored lower on overall irrational thinking, while students who scored higher on awfulizing and low frustration tolerance reported a significantly greater intensity of hassles than did those who scored lower on awfulizing and low frustration tolerance. This indicates support for the ABC model, especially Ellis's construct of irrational beliefs central to this model.

  12. Disruption of the ABC transporter genes PDR5, YOR1, and SNQ2, and their participation in improved fermentative activity of a sake yeast mutant showing pleiotropic drug resistance.

    PubMed

    Watanabe, M; Mizoguchi, H; Nishimura, A

    2000-01-01

    Clotrimazole-resistant mutants from sake yeasts show improved fermentative activity in sake mash and pleiotropic drug resistance (PDR). The PDR mechanism is interpreted by overexpression of ATP-binding cassette (ABC) transporters, which extrude various kinds of drugs out of a cell. In a clotrimazole-resistant mutant, CTZ21, isolated from the haploid sake yeast HL69, the levels of mRNA for three major ABC transporter genes, PDR5, SNQ2, and YOR1, markedly increased. These three genes of CTZ21 were disrupted to investigate which participated in the improved fermentative activity of CTZ21. The fermentative activities of deltapdr5 and deltasnq2 strains of CTZ21 were reduced to that of HL69 in the initial and middle stages of fermentation. In the last stage, however, the sake meter [(1/gravity - 1) x 1443] of the deltapdr5 and deltasnq2 strains rose faster than that of HL69. On the other hand, a deltayor1 strain of CTZ21 fermented sake mash in a manner nearly identical to that of CTZ21 until the last stage of fermentation. But in the last stage, fermentation of the deltayor1 slowed down compared with that of CTZ21. A deltayor1 strain of HL69 also exhibited much reduced fermentative activity in the middle and last fermentation stages. The YOR1 gene seems necessary for sake fermentation to be completed efficiently. The ATP content in sake mash brewed with CTZ21 was drastically decreased throughout the whole fermentation period. This low ATP level was restored to a medium level in the cases of both the deltapdr5 and deltasnq2 strains of CTZ21. In contrast, the deltayor1 of CTZ21 exhibited a low ATP level in sake mash in the same manner as CTZ21. These results suggest that the low ATP level of CTZ21 contributes to a certain extent its improved fermentative activity in the initial and middle stages of sake fermentation.

  13. Applying the Post-Modern Double ABC-X Model to Family Food Insecurity

    ERIC Educational Resources Information Center

    Hutson, Samantha; Anderson, Melinda; Swafford, Melinda

    2015-01-01

    This paper develops the argument that using the Double ABC-X model in family and consumer sciences (FCS) curricula is a way to educate nutrition and dietetics students regarding a family's perceptions of food insecurity. The Double ABC-X model incorporates ecological theory as a basis to explain family stress and the resulting adjustment and…

  14. Experiment Pamir-2. Fianit: A giant super-family with halo (Epsilon sub 0 at approximately 10(17) eV)

    NASA Technical Reports Server (NTRS)

    Zatsepin, G. T.

    1985-01-01

    A superfamily with halo of extremely high energy named Fianit was recorded in X-ray emulsion chamber (XEC) at the Pamirs (atmospheric depth 600 g/sq.cm.). Detailed description of the superfamily and results of its analysis are presented.

  15. Functional Diversity of Haloacid Dehalogenase Superfamily Phosphatases from Saccharomyces cerevisiae: BIOCHEMICAL, STRUCTURAL, AND EVOLUTIONARY INSIGHTS.

    PubMed

    Kuznetsova, Ekaterina; Nocek, Boguslaw; Brown, Greg; Makarova, Kira S; Flick, Robert; Wolf, Yuri I; Khusnutdinova, Anna; Evdokimova, Elena; Jin, Ke; Tan, Kemin; Hanson, Andrew D; Hasnain, Ghulam; Zallot, Rémi; de Crécy-Lagard, Valérie; Babu, Mohan; Savchenko, Alexei; Joachimiak, Andrzej; Edwards, Aled M; Koonin, Eugene V; Yakunin, Alexander F

    2015-07-24

    The haloacid dehalogenase (HAD)-like enzymes comprise a large superfamily of phosphohydrolases present in all organisms. The Saccharomyces cerevisiae genome encodes at least 19 soluble HADs, including 10 uncharacterized proteins. Here, we biochemically characterized 13 yeast phosphatases from the HAD superfamily, which includes both specific and promiscuous enzymes active against various phosphorylated metabolites and peptides with several HADs implicated in detoxification of phosphorylated compounds and pseudouridine. The crystal structures of four yeast HADs provided insight into their active sites, whereas the structure of the YKR070W dimer in complex with substrate revealed a composite substrate-binding site. Although the S. cerevisiae and Escherichia coli HADs share low sequence similarities, the comparison of their substrate profiles revealed seven phosphatases with common preferred substrates. The cluster of secondary substrates supporting significant activity of both S. cerevisiae and E. coli HADs includes 28 common metabolites that appear to represent the pool of potential activities for the evolution of novel HAD phosphatases. Evolution of novel substrate specificities of HAD phosphatases shows no strict correlation with sequence divergence. Thus, evolution of the HAD superfamily combines the conservation of the overall substrate pool and the substrate profiles of some enzymes with remarkable biochemical and structural flexibility of other superfamily members. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. The ABC's of Learning in Infancy.

    ERIC Educational Resources Information Center

    Saunders, Minta M.

    Learning in infancy is based on activity, beginnings, and curiosity, the so-called ABC's. Earliest behavior consists of mass activity, the period from birth to 24 months of sensory-motor development which provides the foundation for all future learning. Adults must provide space, toys, and affectionate care to help infants proceed through…

  17. Interaction of Food Additives with Intestinal Efflux Transporters.

    PubMed

    Sjöstedt, Noora; Deng, Feng; Rauvala, Oskari; Tepponen, Tuomas; Kidron, Heidi

    2017-11-06

    Breast cancer resistance protein (BCRP), multidrug resistance associated protein 2 (MRP2) and P-glycoprotein (P-gp) are ABC transporters that are expressed in the intestine, where they are involved in the efflux of many drugs from enterocytes back into the intestinal lumen. The inhibition of BCRP, MRP2, and P-gp can result in enhanced absorption and exposure of substrate drugs. Food additives are widely used by the food industry to improve the stability, flavor, and consistency of food products. Although they are considered safe for consumption, their interactions with intestinal transporters are poorly characterized. Therefore, in this study, selected food additives, including preservatives, colorants, and sweeteners, were studied in vitro for their inhibitory effects on intestinal ABC transporters. Among the studied compounds, several colorants were able to inhibit BCRP and MRP2, whereas P-gp was fairly insensitive to inhibition. Additionally, one sweetener was identified as a potent inhibitor of BCRP. Dose-response studies revealed that the IC 50 values of the inhibitors were lower than the estimated intestinal concentrations after the consumption of beverages containing food colorants. This suggests that there is potential for previously unrecognized transporter-mediated food additive-drug interactions.

  18. N-retinylidene-phosphatidylethanolamine is the preferred retinoid substrate for the photoreceptor-specific ABC transporter ABCA4 (ABCR).

    PubMed

    Beharry, Seelochan; Zhong, Ming; Molday, Robert S

    2004-12-24

    ABCA4, a member of the family of ATP binding cassette (ABC) proteins found in rod and cone photoreceptors, has been implicated in the transport of retinoid compounds across the outer segment disk membrane following the photoactivation of rhodopsin. Mutations in the ABCA4 gene are responsible for Stargardt macular dystrophy and related retinal degenerative diseases that cause a loss in vision. To identify the retinoid substrate that interacts with ABCA4, we have isolated ABCA4 from rod outer segment disk membranes on an immunoaffinity matrix and analyzed retinoid compounds that bind to ABCA4 using high performance liquid chromatography and radiolabeling methods. When all-trans-retinal was added to ABCA4 in the presence of phosphatidylethanolamine, approximately 0.9 mol of N-retinylidene-phosphatidylethanolamine and 0.3 mol of all-trans-retinal were bound per mol of ABCA4 with an apparent K(d) of 2-5 microm. ATP and GTP released these retinoids from ABCA4, whereas ADP, GDP, and nonhydrolyzable derivatives, adenosine 5'-(beta,gamma-imido)triphosphate and guanosine 5'-(beta,gamma-imido)triphosphate, were ineffective. One mole of N-retinyl-phosphatidylethanolamine, the reduced form of N-retinylidene-phosphatidylethanolamine, bound per mol of ABCA4, whereas 0.3 mol of all-trans-retinal were bound in the absence of phosphatidylethanolamine. No binding of all-trans-retinol to ABCA4 was observed. Our results indicate that ABCA4 preferentially binds N-retinylidene-phosphatidylethanolamine with high affinity in the absence of ATP. Our studies further suggest that ATP binding and hydrolysis induces a protein conformational change that causes N-retinylidene-phosphatidylethanolamine to dissociate from ABCA4.

  19. Our bodies are our own: resistance to ABC-based HIV-prevention programmes in northern Tanzanian conservation organisations.

    PubMed

    Reid-Hresko, John

    2014-01-01

    ABC-based HIV-prevention programmes have been widely employed in northern Tanzanian wildlife conservation settings in an attempt to (re)shape the sexual behaviours of conservation actors. Utilising findings from 66 semi-structured interviews conducted in 2009-2010, this paper examines ABC prevention as a form of Foucauldian governmentality--circulating technologies of power that mobilise disciplinary technologies and attempt to transform such efforts into technologies of the self--and explores how individuals understand and respond to attempts to govern their behaviour. ABC regimes attempt to rework subjectivity, positioning HIV-related behaviours within a risk-based neoliberal rationality. However, efforts to use ABC as a technology to govern populations and individual bodies are largely incommensurate with existing Tanzanian sociocultural formations, including economic and gendered inequalities, and local understandings of sexuality. The language research participants used to talk about ABC and the justifications they offered for non-compliance illuminate this discrepancy. Data reveal that the recipients of ABC campaigns are active producers of understandings that work for them in their lives, but may not produce the behavioural shifts envisioned by programme goals. These findings corroborate previous research, which questions the continued plausibility of ABC as a stand-alone HIV- prevention framework.

  20. Peroxisomal ATP-binding cassette transporters form mainly tetramers

    PubMed Central

    Geillon, Flore; Gondcaille, Catherine; Raas, Quentin; Dias, Alexandre M. M.; Pecqueur, Delphine; Truntzer, Caroline; Lucchi, Géraldine; Ducoroy, Patrick; Falson, Pierre; Savary, Stéphane; Trompier, Doriane

    2017-01-01

    ABCD1 and its homolog ABCD2 are peroxisomal ATP-binding cassette (ABC) half-transporters of fatty acyl-CoAs with both distinct and overlapping substrate specificities. Although it is established that ABC half-transporters have at least to dimerize to generate a functional unit, functional equivalents of tetramers (i.e. dimers of full-length transporters) have also been reported. However, oligomerization of peroxisomal ABCD transporters is incompletely understood but is of potential significance because more complex oligomerization might lead to differences in substrate specificity. In this work, we have characterized the quaternary structure of the ABCD1 and ABCD2 proteins in the peroxisomal membrane. Using various biochemical approaches, we clearly demonstrate that both transporters exist as both homo- and heterotetramers, with a predominance of homotetramers. In addition to tetramers, some larger molecular ABCD assemblies were also found but represented only a minor fraction. By using quantitative co-immunoprecipitation assays coupled with tandem mass spectrometry, we identified potential binding partners of ABCD2 involved in polyunsaturated fatty-acid metabolism. Interestingly, we identified calcium ATPases as ABCD2-binding partners, suggesting a role of ABCD2 in calcium signaling. In conclusion, we have shown here that ABCD1 and its homolog ABCD2 exist mainly as homotetramers in the peroxisomal membrane. PMID:28258215

  1. MDR1 and BCRP Transporter-Mediated Drug-Drug Interaction between Rilpivirine and Abacavir and Effect on Intestinal Absorption.

    PubMed

    Reznicek, Josef; Ceckova, Martina; Ptackova, Zuzana; Martinec, Ondrej; Tupova, Lenka; Cerveny, Lukas; Staud, Frantisek

    2017-09-01

    Rilpivirine (TMC278) is a highly potent nonnucleoside reverse transcriptase inhibitor (NNRTI) representing an effective component of combination antiretroviral therapy (cART) in the treatment of HIV-positive patients. Many antiretroviral drugs commonly used in cART are substrates of ATP-binding cassette (ABC) and/or solute carrier (SLC) drug transporters and, therefore, are prone to pharmacokinetic drug-drug interactions (DDIs). The aim of our study was to evaluate rilpivirine interactions with abacavir and lamivudine on selected ABC and SLC transporters in vitro and assess its importance for pharmacokinetics in vivo Using accumulation assays in MDCK cells overexpressing selected ABC or SLC drug transporters, we revealed rilpivirine as a potent inhibitor of MDR1 and BCRP, but not MRP2, OCT1, OCT2, or MATE1. Subsequent transport experiments across monolayers of MDCKII-MDR1, MDCKII-BCRP, and Caco-2 cells demonstrated that rilpivirine inhibits MDR1- and BCRP-mediated efflux of abacavir and increases its transmembrane transport. In vivo experiments in male Wistar rats confirmed inhibition of MDR1/BCRP in the small intestine, leading to a significant increase in oral bioavailability of abacavir. In conclusion, rilpivirine inhibits MDR1 and BCRP transporters and may affect pharmacokinetic behavior of concomitantly administered substrates of these transporters, such as abacavir. Copyright © 2017 American Society for Microbiology.

  2. Amino Acid Transporters and Release of Hydrophobic Amino Acids in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120.

    PubMed

    Pernil, Rafael; Picossi, Silvia; Herrero, Antonia; Flores, Enrique; Mariscal, Vicente

    2015-04-23

    Anabaena sp. strain PCC 7120 is a filamentous cyanobacterium that can use inorganic compounds such as nitrate or ammonium as nitrogen sources. In the absence of combined nitrogen, it can fix N2 in differentiated cells called heterocysts. Anabaena also shows substantial activities of amino acid uptake, and three ABC-type transporters for amino acids have been previously characterized. Seven new loci encoding predicted amino acid transporters were identified in the Anabaena genomic sequence and inactivated. Two of them were involved in amino acid uptake. Locus alr2535-alr2541 encodes the elements of a hydrophobic amino acid ABC-type transporter that is mainly involved in the uptake of glycine. ORF all0342 encodes a putative transporter from the dicarboxylate/amino acid:cation symporter (DAACS) family whose inactivation resulted in an increased uptake of a broad range of amino acids. An assay to study amino acid release from Anabaena filaments to the external medium was set up. Net release of the alanine analogue α-aminoisobutyric acid (AIB) was observed when transport system N-I (a hydrophobic amino acid ABC-type transporter) was engaged in the uptake of a specific substrate. The rate of AIB release was directly proportional to the intracellular AIB concentration, suggesting leakage from the cells by diffusion.

  3. Identification of an essential active-site residue in the α-D-phosphohexomutase enzyme superfamily.

    PubMed

    Lee, Yingying; Mehra-Chaudhary, Ritcha; Furdui, Cristina; Beamer, Lesa J

    2013-06-01

    Enzymes in the α-d-phosphohexomutase superfamily catalyze the conversion of 1-phosphosugars to their 6-phospho counterparts. Their phosphoryl transfer reaction has long been proposed to require general acid-base catalysts, but candidate residues for these key roles have not been identified. In this study, we show through mutagenesis and kinetic studies that a histidine (His329) in the active site is critical for enzyme activity in a well-studied member of the superfamily, phosphomannomutase/phosphoglucomutase from Pseudomonas aeruginosa. Crystallographic characterization of an H329A mutant protein showed no significant changes from the wild-type enzyme, excluding structural disruption as the source of its compromised activity. Mutation of the structurally analogous lysine residue in a related protein, phosphoglucomutase from Salmonella typhimurium, also results in significant catalytic impairment. Analyses of protein-ligand complexes of the P. aeruginosa enzyme show that His329 is appropriately positioned to abstract a proton from the O1/O6 hydroxyl of the phosphosugar substrates, and thus may serve as the general base in the reaction. Histidine is strongly conserved at this position in many proteins in the superfamily, and lysine is also often conserved at a structurally corresponding position, particularly in the phosphoglucomutase enzyme sub-group. These studies shed light on the mechanism of this important enzyme superfamily, and may facilitate the design of mechanism-based inhibitors. Structural data have been deposited in the Protein Data Bank with accession number 4IL8. © 2013 The Authors Journal compilation © 2013 FEBS.

  4. Purification and characterisation of the yeast plasma membrane ATP binding cassette transporter Pdr11p

    PubMed Central

    Laub, Katrine Rude; Marek, Magdalena; Stanchev, Lyubomir Dimitrov; Herrera, Sara Abad; Kanashova, Tamara; Bourmaud, Adèle; Dittmar, Gunnar

    2017-01-01

    The ATP binding cassette (ABC) transporters Pdr11p and its paralog Aus1p are expressed under anaerobic growth conditions at the plasma membrane of the yeast Saccharomyces cerevisiae and are required for sterol uptake. However, the precise mechanism by which these ABC transporters facilitate sterol movement is unknown. In this study, an overexpression and purification procedure was developed with the aim to characterise the Pdr11p transporter. Engineering of Pdr11p variants fused at the C terminus with green fluorescent protein (Pdr11p-GFP) and containing a FLAG tag at the N terminus facilitated expression analysis and one-step purification, respectively. The detergent-solubilised and purified protein displayed a stable ATPase activity with a broad pH optimum near 7.4. Mutagenesis of the conserved lysine to methionine (K788M) in the Walker A motif abolished ATP hydrolysis. Remarkably, and in contrast to Aus1p, ATPase activity of Pdr11p was insensitive to orthovanadate and not specifically stimulated by phosphatidylserine upon reconstitution into liposomes. Our results highlight distinct differences between Pdr11p and Aus1p and create an experimental basis for further biochemical studies of both ABC transporters to elucidate their function. PMID:28922409

  5. astroABC : An Approximate Bayesian Computation Sequential Monte Carlo sampler for cosmological parameter estimation

    NASA Astrophysics Data System (ADS)

    Jennings, E.; Madigan, M.

    2017-04-01

    Given the complexity of modern cosmological parameter inference where we are faced with non-Gaussian data and noise, correlated systematics and multi-probe correlated datasets,the Approximate Bayesian Computation (ABC) method is a promising alternative to traditional Markov Chain Monte Carlo approaches in the case where the Likelihood is intractable or unknown. The ABC method is called "Likelihood free" as it avoids explicit evaluation of the Likelihood by using a forward model simulation of the data which can include systematics. We introduce astroABC, an open source ABC Sequential Monte Carlo (SMC) sampler for parameter estimation. A key challenge in astrophysics is the efficient use of large multi-probe datasets to constrain high dimensional, possibly correlated parameter spaces. With this in mind astroABC allows for massive parallelization using MPI, a framework that handles spawning of processes across multiple nodes. A key new feature of astroABC is the ability to create MPI groups with different communicators, one for the sampler and several others for the forward model simulation, which speeds up sampling time considerably. For smaller jobs the Python multiprocessing option is also available. Other key features of this new sampler include: a Sequential Monte Carlo sampler; a method for iteratively adapting tolerance levels; local covariance estimate using scikit-learn's KDTree; modules for specifying optimal covariance matrix for a component-wise or multivariate normal perturbation kernel and a weighted covariance metric; restart files output frequently so an interrupted sampling run can be resumed at any iteration; output and restart files are backed up at every iteration; user defined distance metric and simulation methods; a module for specifying heterogeneous parameter priors including non-standard prior PDFs; a module for specifying a constant, linear, log or exponential tolerance level; well-documented examples and sample scripts. This code is hosted

  6. Chlorite dismutases, DyPs, and EfeB: 3 microbial heme enzyme families comprise the CDE structural superfamily

    PubMed Central

    Goblirsch, Brandon; Kurker, Richard C.; Streit, Bennett R.; Wilmot, Carrie M.; DuBois, Jennifer L.

    2011-01-01

    Heme proteins are extremely diverse, widespread, and versatile biocatalysts, sensors, and molecular transporters. The chlorite dismutase family of hemoproteins received its name due to the ability of the first-isolated members to detoxify anthropogenic ClO2−, a function believed to have evolved only in the last few decades. Family members have since been found in fifteen bacterial and archaeal genera, suggesting ancient roots. A structure- and sequence-based examination of the family is presented, in which key sequence and structural motifs are identified and possible functions for family proteins are proposed. Newly identified structural homologies moreover demonstrate clear connections to two other large, ancient, and functionally mysterious protein families. We propose calling them collectively the CDE superfamily of heme proteins. PMID:21354424

  7. Regulation of Human γδ T Cells by BTN3A1 Protein Stability and ATP-Binding Cassette Transporters

    PubMed Central

    Rhodes, David A.; Chen, Hung-Chang; Williamson, James C.; Hill, Alfred; Yuan, Jack; Smith, Sam; Rhodes, Harriet; Trowsdale, John; Lehner, Paul J.; Herrmann, Thomas; Eberl, Matthias

    2018-01-01

    Activation of human Vγ9/Vδ2 T cells by “phosphoantigens” (pAg), the microbial metabolite (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP) and the endogenous isoprenoid intermediate isopentenyl pyrophosphate, requires expression of butyrophilin BTN3A molecules by presenting cells. However, the precise mechanism of activation of Vγ9/Vδ2 T cells by BTN3A molecules remains elusive. It is not clear what conformation of the three BTN3A isoforms transmits activation signals nor how externally delivered pAg accesses the cytosolic B30.2 domain of BTN3A1. To approach these problems, we studied two HLA haplo-identical HeLa cell lines, termed HeLa-L and HeLa-M, which showed marked differences in pAg-dependent stimulation of Vγ9/Vδ2 T cells. Levels of IFN-γ secretion by Vγ9/Vδ2 T cells were profoundly increased by pAg loading, or by binding of the pan-BTN3A specific agonist antibody CD277 20.1, in HeLa-M compared to HeLa-L cells. IL-2 production from a murine hybridoma T cell line expressing human Vγ9/Vδ2 T cell receptor (TCR) transgenes confirmed that the differential responsiveness to HeLa-L and HeLa-M was TCR dependent. By tissue typing, both HeLa lines were shown to be genetically identical and full-length transcripts of the three BTN3A isoforms were detected in equal abundance with no sequence variation. Expression of BTN3A and interacting molecules, such as periplakin or RhoB, did not account for the functional variation between HeLa-L and HeLa-M cells. Instead, the data implicate a checkpoint controlling BTN3A1 stability and protein trafficking, acting at an early time point in its maturation. In addition, plasma membrane profiling was used to identify proteins upregulated in HMB-PP-treated HeLa-M. ABCG2, a member of the ATP-binding cassette (ABC) transporter family was the most significant candidate, which crucially showed reduced expression in HeLa-L. Expression of a subset of ABC transporters, including ABCA1 and ABCG1, correlated with

  8. Accelerator-based conversion (ABC) of weapons plutonium: Plant layout study and related design issues

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Cowell, B.S.; Fontana, M.H.; Krakowski, R.A.

    1995-04-01

    In preparation for and in support of a detailed R and D Plan for the Accelerator-Based Conversion (ABC) of weapons plutonium, an ABC Plant Layout Study was conducted at the level of a pre-conceptual engineering design. The plant layout is based on an adaptation of the Molten-Salt Breeder Reactor (MSBR) detailed conceptual design that was completed in the early 1070s. Although the ABC Plant Layout Study included the Accelerator Equipment as an essential element, the engineering assessment focused primarily on the Target; Primary System (blanket and all systems containing plutonium-bearing fuel salt); the Heat-Removal System (secondary-coolant-salt and supercritical-steam systems); Chemicalmore » Processing; Operation and Maintenance; Containment and Safety; and Instrumentation and Control systems. Although constrained primarily to a reflection of an accelerator-driven (subcritical) variant of MSBR system, unique features and added flexibilities of the ABC suggest improved or alternative approaches to each of the above-listed subsystems; these, along with the key technical issues in need of resolution through a detailed R&D plan for ABC are described on the bases of the ``strawman`` or ``point-of-departure`` plant layout that resulted from this study.« less

  9. Inferring population history with DIY ABC: a user-friendly approach to approximate Bayesian computation.

    PubMed

    Cornuet, Jean-Marie; Santos, Filipe; Beaumont, Mark A; Robert, Christian P; Marin, Jean-Michel; Balding, David J; Guillemaud, Thomas; Estoup, Arnaud

    2008-12-01

    Genetic data obtained on population samples convey information about their evolutionary history. Inference methods can extract part of this information but they require sophisticated statistical techniques that have been made available to the biologist community (through computer programs) only for simple and standard situations typically involving a small number of samples. We propose here a computer program (DIY ABC) for inference based on approximate Bayesian computation (ABC), in which scenarios can be customized by the user to fit many complex situations involving any number of populations and samples. Such scenarios involve any combination of population divergences, admixtures and population size changes. DIY ABC can be used to compare competing scenarios, estimate parameters for one or more scenarios and compute bias and precision measures for a given scenario and known values of parameters (the current version applies to unlinked microsatellite data). This article describes key methods used in the program and provides its main features. The analysis of one simulated and one real dataset, both with complex evolutionary scenarios, illustrates the main possibilities of DIY ABC. The software DIY ABC is freely available at http://www.montpellier.inra.fr/CBGP/diyabc.

  10. Effect of body temperature on chondroitinase ABC's ability to cleave chondroitin sulfate glycosaminoglycans.

    PubMed

    Tester, Nicole J; Plaas, Anna H; Howland, Dena R

    2007-04-01

    Chondroitinase ABC (Ch'ase ABC) is a bacterial lyase that degrades chondroitin sulfate (CS), dermatan sulfate, and hyaluronan glycosaminoglycans (GAGs). This enzyme has received significant attention as a potential therapy for promoting central nervous system and peripheral nervous system repair based on its degradation of CS GAGs. Determination of the stability of Ch'ase ABC activity at temperatures equivalent to normal (37 degrees C) and elevated (39 degrees C) body temperatures is important for optimizing its clinical usage. We report here data obtained from examining enzymatic activity at these temperatures across nine lots of commercially available protease-free Ch'ase ABC. CS GAG degrading activity was assayed by using 1) immunohistochemical detection of unsaturated disaccharide stubs generated by digestion of proteoglycans in tissue sections and 2) fluorophore-assisted carbohydrate electrophoresis (FACE) and/or high-performance liquid chromatography (HPLC) to separate and quantify unsaturated disaccharide digestion products. Our results indicate that there is a significant effect of lot and time on enzymatic thermostability. Average enzymatic activity is significantly decreased at 1 and 3 days at 39 degrees C and 37 degrees C, respectively. Furthermore, the average activity seen after 1 day was significantly different between the two temperatures. Addition of bovine serum albumin as a stabilizer significantly preserved enzymatic activity at 1 day, but not 3 days, at 39 degrees C. These results show that the CS GAG degrading activity of Ch'ase ABC is significantly decreased with incubation at body temperature over time and that all lots do not show equal thermostability. These findings are important for the design and interpretation of experimental and potential clinical studies involving Ch'ase ABC. (c) 2007 Wiley-Liss, Inc.

  11. Aerosol and Cloud Properties during the Cloud Cheju ABC Plume -Asian Monsoon Experiment (CAPMEX) 2008: Linking between Ground-based and UAV Measurements

    NASA Astrophysics Data System (ADS)

    Kim, S.; Yoon, S.; Venkata Ramana, M.; Ramanathan, V.; Nguyen, H.; Park, S.; Kim, M.

    2009-12-01

    Cheju Atmospheric Brown Cloud (ABC) Plume-Monsoon Experiment (CAPMEX), comprehsensive ground-based measurements and a series of data-gathering flights by specially equipped autonomous unmanned aerial vehicles (AUAVs) for aerosol and cloud, had conducted at Jeju (formerly, Cheju), South Korea during August-September 2008, to improve our understanding of how the reduction of anthropogenic emissions in China (so-called “great shutdown” ) during and after the Summer Beijing Olympic Games 2008 effcts on the air quliaty and radiation budgets and how atmospheric brown clouds (ABCs) influences solar radiation budget off Asian continent. Large numbers of in-situ and remote sensing instruments at the Gosan ABC observatory and miniaturized instruments on the aircraft measure a range of properties such as the quantity of soot, size-segregated aerosol particle numbers, total particle numbers, size-segregated cloud droplet numbers (only AUAV), aerosol scattering properties (only ground), aerosol vertical distribution, column-integrated aerosol properties, and meteorological variables. By integrating ground-level and high-elevation AUAV measurements with NASA-satellite observations (e.g., MODIS, CALIPSO), we investigate the long range transport of aerosols, the impact of ABCs on clouds, and the role of biogenic and anthropogenic aerosols on cloud condensation nuclei (CCN). In this talk, we will present the results from CAPMEX focusing on: (1) the characteristics of aerosol optical, physical and chemical properties at Gosan observatory, (2) aerosol solar heating calculated from the ground-based micro-pulse lidar and AERONET sun/sky radiometer synergy, and comparison with direct measurements from UAV, and (3) aerosol-cloud interactions in conjunction with measurements by satellites and Gosan observatory.

  12. Solute Carriers in the Blood-Brain Barier: Safety in Abundance.

    PubMed

    Nałęcz, Katarzyna A

    2017-03-01

    Blood-brain barrier formed by brain capillary endothelial cells, being in contact with astrocytes endfeet and pericytes, separates extracellular fluid from plasma. Supply of necessary nutrients and removal of certain metabolites takes place due to the activity of transporting proteins from ABC (ATP binding cassette) and SLC (solute carrier) superfamilies. This review is focused on the SLC families involved in transport though the blood-brain barrier of energetic substrates (glucose, monocarboxylates, creatine), amino acids, neurotransmitters and their precursors, as well as organic ions. Members of SLC1, SLC2, SLC3/SLC7, SLC5, SLC6, SLC16, SLC22, SLC38, SLC44, SLC47 and SLCO (SLC21), whose presence in the blood-brain barriers has been demonstrated are characterized with a special emphasis put on polarity of transporters localization in a luminal (blood side) versus an abluminal (brain side) membrane.

  13. Exploring Fold Space Preferences of New-born and Ancient Protein Superfamilies

    PubMed Central

    Edwards, Hannah; Abeln, Sanne; Deane, Charlotte M.

    2013-01-01

    The evolution of proteins is one of the fundamental processes that has delivered the diversity and complexity of life we see around ourselves today. While we tend to define protein evolution in terms of sequence level mutations, insertions and deletions, it is hard to translate these processes to a more complete picture incorporating a polypeptide's structure and function. By considering how protein structures change over time we can gain an entirely new appreciation of their long-term evolutionary dynamics. In this work we seek to identify how populations of proteins at different stages of evolution explore their possible structure space. We use an annotation of superfamily age to this space and explore the relationship between these ages and a diverse set of properties pertaining to a superfamily's sequence, structure and function. We note several marked differences between the populations of newly evolved and ancient structures, such as in their length distributions, secondary structure content and tertiary packing arrangements. In particular, many of these differences suggest a less elaborate structure for newly evolved superfamilies when compared with their ancient counterparts. We show that the structural preferences we report are not a residual effect of a more fundamental relationship with function. Furthermore, we demonstrate the robustness of our results, using significant variation in the algorithm used to estimate the ages. We present these age estimates as a useful tool to analyse protein populations. In particularly, we apply this in a comparison of domains containing greek key or jelly roll motifs. PMID:24244135

  14. The short version of the Activities-specific Balance Confidence (ABC) scale: its validity, reliability, and relationship to balance impairment and falls in older adults.

    PubMed

    Schepens, Stacey; Goldberg, Allon; Wallace, Melissa

    2010-01-01

    A shortened version of the ABC 16-item scale (ABC-16), the ABC-6, has been proposed as an alternative balance confidence measure. We investigated whether the ABC-6 is a valid and reliable measure of balance confidence and examined its relationship to balance impairment and falls in older adults. Thirty-five community-dwelling older adults completed the ABC-16, including the 6 questions of the ABC-6. They also completed the following clinical balance tests: unipedal stance time (UST), functional reach (FR), Timed Up and Go (TUG), and maximum step length (MSL). Participants reported 12-month falls history. Balance confidence on the ABC-6 was significantly lower than on the ABC-16, however scores were highly correlated. Fallers reported lower balance confidence than non-fallers as measured by the ABC-6 scale, but confidence did not differ between the groups with the ABC-16. The ABC-6 significantly correlated with all balance tests assessed and number of falls. The ABC-16 significantly correlated with all balance tests assessed, but not with number of falls. Test-retest reliability for the ABC-16 and ABC-6 was good to excellent. The ABC-6 is a valid and reliable measure of balance confidence in community-dwelling older adults, and shows stronger relationships to falls than does the ABC-16. The ABC-6 may be a more useful balance confidence assessment tool than the ABC-16. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

  15. Knowing the ABCs: A Comparative Effectiveness Study of Two Methods of Diabetes Education

    PubMed Central

    Naik, Aanand D.; Teal, Cayla R.; Rodriguez, Elisa; Haidet, Paul

    2011-01-01

    Objective To test an active-learning, empowerment approach to teaching patients about the “diabetes ABCs” (hemoglobin A1C, systolic blood pressure, and low density lipoprotein cholesterol). Methods 84 (97%) diabetic patients who participated in a randomized effectiveness trial of two clinic-based group educational methods and completed a post-intervention assessment. The empowerment arm participated in a group session that incorporated two educational innovations (a conceptual metaphor to foster understanding, and team-based learning methods to foster active learning). The traditional diabetes education arm received a didactic group session focused on self-management and educational materials about the diabetes ABCs. Participants in both arms received individual review of their current ABC values. Results A questionnaire evaluated knowledge, understanding, and recall of the diabetes ABCs was administered three months after enrollment in the study. At three months, participants in the empowerment group demonstrated greater understanding of the diabetes ABCs (P<.0001), greater knowledge of their own values (P<.0001), and greater knowledge of guideline-derived target goals for the ABCs compared with participants in the traditional arm (P<.0001). Conclusion An active-learning, empowerment-based approach applied to diabetes education can lead to greater understanding and knowledge retention. Practice Implications An empowerment approach to education can facilitate informed, activated patients and increase performance of self-management behaviors. PMID:21300516

  16. Inhibition of Human Drug Transporter Activities by the Pyrethroid Pesticides Allethrin and Tetramethrin

    PubMed Central

    Chedik, Lisa; Bruyere, Arnaud; Le Vee, Marc; Stieger, Bruno; Denizot, Claire; Parmentier, Yannick; Potin, Sophie; Fardel, Olivier

    2017-01-01

    Pyrethroids are widely-used chemical insecticides, to which humans are commonly exposed, and known to alter functional expression of drug metabolizing enzymes. Limited data have additionally suggested that drug transporters, that constitute key-actors of the drug detoxification system, may also be targeted by pyrethroids. The present study was therefore designed to analyze the potential regulatory effects of these pesticides towards activities of main ATP-binding cassette (ABC) and solute carrier (SLC) drug transporters, using transporter-overexpressing cells. The pyrethroids allethrin and tetramethrin were found to inhibit various ABC and SLC drug transporters, including multidrug resistance-associated protein (MRP) 2, breast cancer resistance protein (BCRP), organic anion transporter polypeptide (OATP) 1B1, organic anion transporter (OAT) 3, multidrug and toxin extrusion transporter (MATE) 1, organic cation transporter (OCT) 1 and OCT2, with IC50 values however ranging from 2.6 μM (OCT1 inhibition by allethrin) to 77.6 μM (OAT3 inhibition by tetramethrin) and thus much higher than pyrethroid concentrations (in the nM range) reached in environmentally pyrethroid-exposed humans. By contrast, allethrin and tetramethrin cis-stimulated OATP2B1 activity and failed to alter activities of OATP1B3, OAT1 and MATE2-K, whereas P-glycoprotein activity was additionally moderately inhibited. Twelve other pyrethoids used at 100 μM did not block activities of the various investigated transporters, or only moderately inhibited some of them (inhibition by less than 50%). In silico analysis of structure-activity relationships next revealed that molecular parameters, including molecular weight and lipophilicity, are associated with transporter inhibition by allethrin/tetramethrin and successfully predicted transporter inhibition by the pyrethroids imiprothrin and prallethrin. Taken together, these data fully demonstrated that two pyrethoids, i.e., allethrin and tetramethrin, can

  17. Identification and characterization of the gltK gene encoding a membrane-associated glucose transport protein of pseudomonas aeruginosa.

    PubMed

    Adewoye, L O; Worobec, E A

    2000-08-08

    The Pseudomonas aeruginosa oprB gene encodes the carbohydrate-selective OprB porin, which translocates substrate molecules across the outer membrane to the periplasmic glucose-binding protein. We identified and cloned two open reading frames (ORFs) flanking the oprB gene but are not in operonic arrangement with the oprB gene. The downstream ORF encodes a putative polypeptide homologous to members of a family of transcriptional repressors, whereas the oprB gene is preceded by an ORF encoding a putative product, which exhibits strong homology to several carbohydrate transport ATP-binding cassette (ABC) proteins. The genomic copy of the upstream ORF was mutagenized by homologous recombination. Analysis of the deletion mutant in comparison with the wild type revealed a significant reduction in [14C] glucose transport activity in the mutant strain, suggesting that this ORF likely encodes the inner membrane component of the glucose ABC transporter. It is thus designated gltK gene to reflect its homology to the Pseudomona fluorescens mtlK and its involvement in the high-affinity glucose transport system. Multiple alignment analysis revealed that the P. aeruginosa gltK gene product is a member of the MalK subfamily of ABC proteins.

  18. Transcription of putative tonoplast transporters in response to glyphosate and paraquat stress in Conyza bonariensis and Conyza canadensis and selection of reference genes for qRT-PCR.

    PubMed

    Moretti, Marcelo L; Alárcon-Reverte, Rocio; Pearce, Stephen; Morran, Sarah; Hanson, Bradley D

    2017-01-01

    Herbicide resistance is a challenge for modern agriculture further complicated by cases of resistance to multiple herbicides. Conyza bonariensis and Conyza canadensis are invasive weeds of field crops, orchards, and non-cropped areas in many parts of the world. In California, USA, Conyza populations resistant to the herbicides glyphosate and paraquat have recently been described. Although the mechanism conferring resistance to glyphosate and paraquat in these species was not elucidated, reduced translocation of these herbicides was observed under experimental conditions in both species. Glyphosate and paraquat resistance associated with reduced translocation are hypothesized to be a result of sequestration of herbicides into the vacuole, with the possible involvement of over-expression of genes encoding tonoplast transporters of ABC-transporter families in cases of glyphosate resistance or cationic amino acid transporters (CAT) in cases of paraquat resistance. However, gene expression in response to herbicide treatment has not been studied in glyphosate and paraquat resistant populations. In the current study, we evaluated the transcript levels of genes possibly involved in resistance using real-time PCR. First, we evaluated eight candidate reference genes following herbicide treatment and selected three genes that exhibited stable expression profiles; ACTIN, HEAT-SHOCK-PROTEIN-70, and CYCLOPHILIN. The reference genes identified here can be used for further studies related to plant-herbicide interactions. We used these reference genes to assay the transcript levels of EPSPS, ABC transporters, and CAT in response to herbicide treatment in susceptible and resistant Conyza spp. lines. No transcription changes were observed in EPSPS or CAT genes after glyphosate or paraquat treatment, suggesting that these genes are not involved in the resistance mechanism. Transcription of the two ABC transporter genes increased following glyphosate treatment in all Conyza spp. lines

  19. Transcription of putative tonoplast transporters in response to glyphosate and paraquat stress in Conyza bonariensis and Conyza canadensis and selection of reference genes for qRT-PCR

    PubMed Central

    Alárcon-Reverte, Rocio; Pearce, Stephen; Morran, Sarah; Hanson, Bradley D.

    2017-01-01

    Herbicide resistance is a challenge for modern agriculture further complicated by cases of resistance to multiple herbicides. Conyza bonariensis and Conyza canadensis are invasive weeds of field crops, orchards, and non-cropped areas in many parts of the world. In California, USA, Conyza populations resistant to the herbicides glyphosate and paraquat have recently been described. Although the mechanism conferring resistance to glyphosate and paraquat in these species was not elucidated, reduced translocation of these herbicides was observed under experimental conditions in both species. Glyphosate and paraquat resistance associated with reduced translocation are hypothesized to be a result of sequestration of herbicides into the vacuole, with the possible involvement of over-expression of genes encoding tonoplast transporters of ABC-transporter families in cases of glyphosate resistance or cationic amino acid transporters (CAT) in cases of paraquat resistance. However, gene expression in response to herbicide treatment has not been studied in glyphosate and paraquat resistant populations. In the current study, we evaluated the transcript levels of genes possibly involved in resistance using real-time PCR. First, we evaluated eight candidate reference genes following herbicide treatment and selected three genes that exhibited stable expression profiles; ACTIN, HEAT-SHOCK-PROTEIN-70, and CYCLOPHILIN. The reference genes identified here can be used for further studies related to plant-herbicide interactions. We used these reference genes to assay the transcript levels of EPSPS, ABC transporters, and CAT in response to herbicide treatment in susceptible and resistant Conyza spp. lines. No transcription changes were observed in EPSPS or CAT genes after glyphosate or paraquat treatment, suggesting that these genes are not involved in the resistance mechanism. Transcription of the two ABC transporter genes increased following glyphosate treatment in all Conyza spp. lines

  20. Inferring epidemiological parameters from phylogenies using regression-ABC: A comparative study

    PubMed Central

    Gascuel, Olivier

    2017-01-01

    Inferring epidemiological parameters such as the R0 from time-scaled phylogenies is a timely challenge. Most current approaches rely on likelihood functions, which raise specific issues that range from computing these functions to finding their maxima numerically. Here, we present a new regression-based Approximate Bayesian Computation (ABC) approach, which we base on a large variety of summary statistics intended to capture the information contained in the phylogeny and its corresponding lineage-through-time plot. The regression step involves the Least Absolute Shrinkage and Selection Operator (LASSO) method, which is a robust machine learning technique. It allows us to readily deal with the large number of summary statistics, while avoiding resorting to Markov Chain Monte Carlo (MCMC) techniques. To compare our approach to existing ones, we simulated target trees under a variety of epidemiological models and settings, and inferred parameters of interest using the same priors. We found that, for large phylogenies, the accuracy of our regression-ABC is comparable to that of likelihood-based approaches involving birth-death processes implemented in BEAST2. Our approach even outperformed these when inferring the host population size with a Susceptible-Infected-Removed epidemiological model. It also clearly outperformed a recent kernel-ABC approach when assuming a Susceptible-Infected epidemiological model with two host types. Lastly, by re-analyzing data from the early stages of the recent Ebola epidemic in Sierra Leone, we showed that regression-ABC provides more realistic estimates for the duration parameters (latency and infectiousness) than the likelihood-based method. Overall, ABC based on a large variety of summary statistics and a regression method able to perform variable selection and avoid overfitting is a promising approach to analyze large phylogenies. PMID:28263987

  1. IMG-ABC: A Knowledge Base To Fuel Discovery of Biosynthetic Gene Clusters and Novel Secondary Metabolites.

    PubMed

    Hadjithomas, Michalis; Chen, I-Min Amy; Chu, Ken; Ratner, Anna; Palaniappan, Krishna; Szeto, Ernest; Huang, Jinghua; Reddy, T B K; Cimermančič, Peter; Fischbach, Michael A; Ivanova, Natalia N; Markowitz, Victor M; Kyrpides, Nikos C; Pati, Amrita

    2015-07-14

    In the discovery of secondary metabolites, analysis of sequence data is a promising exploration path that remains largely underutilized due to the lack of computational platforms that enable such a systematic approach on a large scale. In this work, we present IMG-ABC (https://img.jgi.doe.gov/abc), an atlas of biosynthetic gene clusters within the Integrated Microbial Genomes (IMG) system, which is aimed at harnessing the power of "big" genomic data for discovering small molecules. IMG-ABC relies on IMG's comprehensive integrated structural and functional genomic data for the analysis of biosynthetic gene clusters (BCs) and associated secondary metabolites (SMs). SMs and BCs serve as the two main classes of objects in IMG-ABC, each with a rich collection of attributes. A unique feature of IMG-ABC is the incorporation of both experimentally validated and computationally predicted BCs in genomes as well as metagenomes, thus identifying BCs in uncultured populations and rare taxa. We demonstrate the strength of IMG-ABC's focused integrated analysis tools in enabling the exploration of microbial secondary metabolism on a global scale, through the discovery of phenazine-producing clusters for the first time in Alphaproteobacteria. IMG-ABC strives to fill the long-existent void of resources for computational exploration of the secondary metabolism universe; its underlying scalable framework enables traversal of uncovered phylogenetic and chemical structure space, serving as a doorway to a new era in the discovery of novel molecules. IMG-ABC is the largest publicly available database of predicted and experimental biosynthetic gene clusters and the secondary metabolites they produce. The system also includes powerful search and analysis tools that are integrated with IMG's extensive genomic/metagenomic data and analysis tool kits. As new research on biosynthetic gene clusters and secondary metabolites is published and more genomes are sequenced, IMG-ABC will continue to

  2. '2TM proteins': an antigenically diverse superfamily with variable functions and export pathways.

    PubMed

    Kaur, Jasweer; Hora, Rachna

    2018-01-01

    Malaria is a disease that affects millions of people annually. An intracellular habitat and lack of protein synthesizing machinery in erythrocytes pose numerous difficulties for survival of the human pathogen Plasmodium falciparum . The parasite refurbishes the infected red blood cell (iRBC) by synthesis and export of several proteins in an attempt to suffice its metabolic needs and evade the host immune response. Immune evasion is largely mediated by surface display of highly polymorphic protein families known as variable surface antigens. These include the two trans-membrane (2TM) superfamily constituted by multicopy repetitive interspersed family (RIFINs), subtelomeric variable open reading frame (STEVORs) and Plasmodium falciparum Maurer's cleft two trans-membrane proteins present only in P. falciparum and some simian infecting Plasmodium species. Their hypervariable region flanked by 2TM domains exposed on the iRBC surface is believed to generate antigenic diversity. Though historically named "2TM superfamily," several A-type RIFINs and some STEVORs assume one trans-membrane topology. RIFINs and STEVORs share varied functions in different parasite life cycle stages like rosetting, alteration of iRBC rigidity and immune evasion. Additionally, a member of the STEVOR family has been implicated in merozoite invasion. Differential expression of these families in laboratory strains and clinical isolates propose them to be important for host cell survival and defense. The role of RIFINs in modulation of host immune response and presence of protective antibodies against these surface exposed molecules in patient sera highlights them as attractive targets of antimalarial therapies and vaccines. 2TM proteins are Plasmodium export elements positive, and several of these are exported to the infected erythrocyte surface after exiting through the classical secretory pathway within parasites. Cleaved and modified proteins are trafficked after packaging in vesicles to reach

  3. Small substrate transport and mechanism of a molybdate ATP binding cassette transporter in a lipid environment.

    PubMed

    Rice, Austin J; Harrison, Alistair; Alvarez, Frances J D; Davidson, Amy L; Pinkett, Heather W

    2014-05-23

    Embedded in the plasma membrane of all bacteria, ATP binding cassette (ABC) importers facilitate the uptake of several vital nutrients and cofactors. The ABC transporter, MolBC-A, imports molybdate by passing substrate from the binding protein MolA to a membrane-spanning translocation pathway of MolB. To understand the mechanism of transport in the biological membrane as a whole, the effects of the lipid bilayer on transport needed to be addressed. Continuous wave-electron paramagnetic resonance and in vivo molybdate uptake studies were used to test the impact of the lipid environment on the mechanism and function of MolBC-A. Working with the bacterium Haemophilus influenzae, we found that MolBC-A functions as a low affinity molybdate transporter in its native environment. In periods of high extracellular molybdate concentration, H. influenzae makes use of parallel molybdate transport systems (MolBC-A and ModBC-A) to take up a greater amount of molybdate than a strain with ModBC-A alone. In addition, the movement of the translocation pathway in response to nucleotide binding and hydrolysis in a lipid environment is conserved when compared with in-detergent analysis. However, electron paramagnetic resonance spectroscopy indicates that a lipid environment restricts the flexibility of the MolBC translocation pathway. By combining continuous wave-electron paramagnetic resonance spectroscopy and substrate uptake studies, we reveal details of molybdate transport and the logistics of uptake systems that employ multiple transporters for the same substrate, offering insight into the mechanisms of nutrient uptake in bacteria. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. ATP-binding cassette transporters and sterol O-acyltransferases interact at membrane microdomains to modulate sterol uptake and esterification.

    PubMed

    Gulati, Sonia; Balderes, Dina; Kim, Christine; Guo, Zhongmin A; Wilcox, Lisa; Area-Gomez, Estela; Snider, Jamie; Wolinski, Heimo; Stagljar, Igor; Granato, Juliana T; Ruggles, Kelly V; DeGiorgis, Joseph A; Kohlwein, Sepp D; Schon, Eric A; Sturley, Stephen L

    2015-11-01

    A key component of eukaryotic lipid homeostasis is the esterification of sterols with fatty acids by sterol O-acyltransferases (SOATs). The esterification reactions are allosterically activated by their sterol substrates, the majority of which accumulate at the plasma membrane. We demonstrate that in yeast, sterol transport from the plasma membrane to the site of esterification is associated with the physical interaction of the major SOAT, acyl-coenzyme A:cholesterol acyltransferase (ACAT)-related enzyme (Are)2p, with 2 plasma membrane ATP-binding cassette (ABC) transporters: Aus1p and Pdr11p. Are2p, Aus1p, and Pdr11p, unlike the minor acyltransferase, Are1p, colocalize to sterol and sphingolipid-enriched, detergent-resistant microdomains (DRMs). Deletion of either ABC transporter results in Are2p relocalization to detergent-soluble membrane domains and a significant decrease (53-36%) in esterification of exogenous sterol. Similarly, in murine tissues, the SOAT1/Acat1 enzyme and activity localize to DRMs. This subcellular localization is diminished upon deletion of murine ABC transporters, such as Abcg1, which itself is DRM associated. We propose that the close proximity of sterol esterification and transport proteins to each other combined with their residence in lipid-enriched membrane microdomains facilitates rapid, high-capacity sterol transport and esterification, obviating any requirement for soluble intermediary proteins. © FASEB.

  5. Evolution of Enzymatic Activities in the Enolase Superfamily: D-Mannonate Dhydratase from Novosphingobium aromaticivorans

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Rakus,J.; Fedorov, A.; Fedorov, E.

    2007-01-01

    The d-mannonate dehydratase (ManD) function was assigned to a group of orthologous proteins in the mechanistically diverse enolase superfamily by screening a library of acid sugars. Structures of the wild type ManD from Novosphingobium aromaticivorans were determined at pH 7.5 in the presence of Mg2+ and also in the presence of Mg2+ and the 2-keto-3-keto-d-gluconate dehydration product; the structure of the catalytically active K271E mutant was determined at pH 5.5 in the presence of the d-mannonate substrate. As previously observed in the structures of other members of the enolase superfamily, ManD contains two domains, an N-terminal a+{beta} capping domain andmore » a ({beta}/a)7{beta}-barrel domain. The barrel domain contains the ligands for the essential Mg2+, Asp 210, Glu 236, and Glu 262, at the ends of the third, fourth, and fifth {beta}-strands of the barrel domain, respectively. However, the barrel domain lacks both the Lys acid/base catalyst at the end of the second {beta}-strand and the His-Asp dyad acid/base catalyst at the ends of the seventh and sixth {beta}-strands, respectively, that are found in many members of the superfamily. Instead, a hydrogen-bonded dyad of Tyr 159 in a loop following the second {beta}-strand and Arg 147 at the end of the second {beta}-strand are positioned to initiate the reaction by abstraction of the 2-proton. Both Tyr 159 and His 212, at the end of the third {beta}-strand, are positioned to facilitate both syn-dehydration and ketonization of the resulting enol intermediate to yield the 2-keto-3-keto-d-gluconate product with the observed retention of configuration. The identities and locations of these acid/base catalysts as well as of cationic amino acid residues that stabilize the enolate anion intermediate define a new structural strategy for catalysis (subgroup) in the mechanistically diverse enolase superfamily. With these differences, we provide additional evidence that the ligands for the essential Mg2+ are the

  6. Inferring population history with DIY ABC: a user-friendly approach to approximate Bayesian computation

    PubMed Central

    Cornuet, Jean-Marie; Santos, Filipe; Beaumont, Mark A.; Robert, Christian P.; Marin, Jean-Michel; Balding, David J.; Guillemaud, Thomas; Estoup, Arnaud

    2008-01-01

    Summary: Genetic data obtained on population samples convey information about their evolutionary history. Inference methods can extract part of this information but they require sophisticated statistical techniques that have been made available to the biologist community (through computer programs) only for simple and standard situations typically involving a small number of samples. We propose here a computer program (DIY ABC) for inference based on approximate Bayesian computation (ABC), in which scenarios can be customized by the user to fit many complex situations involving any number of populations and samples. Such scenarios involve any combination of population divergences, admixtures and population size changes. DIY ABC can be used to compare competing scenarios, estimate parameters for one or more scenarios and compute bias and precision measures for a given scenario and known values of parameters (the current version applies to unlinked microsatellite data). This article describes key methods used in the program and provides its main features. The analysis of one simulated and one real dataset, both with complex evolutionary scenarios, illustrates the main possibilities of DIY ABC. Availability: The software DIY ABC is freely available at http://www.montpellier.inra.fr/CBGP/diyabc. Contact: j.cornuet@imperial.ac.uk Supplementary information: Supplementary data are also available at http://www.montpellier.inra.fr/CBGP/diyabc PMID:18842597

  7. Role of MRP transporters in regulating antimicrobial drug inefficacy and oxidative stress-induced pathogenesis during HIV-1 and TB infections.

    PubMed

    Roy, Upal; Barber, Paul; Tse-Dinh, Yuk-Ching; Batrakova, Elena V; Mondal, Debasis; Nair, Madhavan

    2015-01-01

    Multi-Drug Resistance Proteins (MRPs) are members of the ATP binding cassette (ABC) drug-efflux transporter superfamily. MRPs are known to regulate the efficacy of a broad range of anti-retroviral drugs (ARV) used in highly active antiretroviral therapy (HAART) and antibacterial agents used in Tuberculus Bacilli (TB) therapy. Due to their role in efflux of glutathione (GSH) conjugated drugs, MRPs can also regulate cellular oxidative stress, which may contribute to both HIV and/or TB pathogenesis. This review focuses on the characteristics, functional expression, and modulation of known members of the MRP family in HIV infected cells exposed to ARV drugs and discusses their known role in drug-inefficacy in HIV/TB-induced dysfunctions. Currently, nine members of the MRP family (MRP1-MRP9) have been identified, with MRP1 and MRP2 being the most extensively studied. Details of the other members of this family have not been known until recently, but differential expression has been documented in inflammatory tissues. Researchers have found that the distribution, function, and reactivity of members of MRP family vary in different types of lymphocytes and macrophages, and are differentially expressed at the basal and apical surfaces of both endothelial and epithelial cells. Therefore, the prime objective of this review is to delineate the role of MRP transporters in HAART and TB therapy and their potential in precipitating cellular dysfunctions manifested in these chronic infectious diseases. We also provide an overview of different available options and novel experimental strategies that are being utilized to overcome the drug resistance and disease pathogenesis mediated by these membrane transporters.

  8. Structural basis of transport function in major facilitator superfamily protein from Trichoderma harzianum.

    PubMed

    Chaudhary, Nitika; Sandhu, Padmani; Ahmed, Mushtaq; Akhter, Yusuf

    2017-02-01

    Trichothecenes are the sesquiterpenes secreted by Trichoderma spp. residing in the rhizosphere. These compounds have been reported to act as plant growth promoters and bio-control agents. The structural knowledge for the transporter proteins of their efflux remained limited. In this study, three-dimensional structure of Thmfs1 protein, a trichothecene transporter from Trichoderma harzianum, was homology modelled and further Molecular Dynamics (MD) simulations were used to decipher its mechanism. Fourteen transmembrane helices of Thmfs1 protein are observed contributing to an inward-open conformation. The transport channel and ligand binding sites in Thmfs1 are identified based on heuristic, iterative algorithm and structural alignment with homologous proteins. MD simulations were performed to reveal the differential structural behaviour occurring in the ligand free and ligand bound forms. We found that two discrete trichothecene binding sites are located on either side of the central transport tunnel running from the cytoplasmic side to the extracellular side across the Thmfs1 protein. Detailed analysis of the MD trajectories showed an alternative access mechanism between N and C-terminal domains contributing to its function. These results also demonstrate that the transport of trichodermin occurs via hopping mechanism in which the substrate molecule jumps from one binding site to another lining the transport tunnel. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. A Novel Class of Modular Transporters for Vitamins in Prokaryotes ▿ †

    PubMed Central

    Rodionov, Dmitry A.; Hebbeln, Peter; Eudes, Aymerick; ter Beek, Josy; Rodionova, Irina A.; Erkens, Guus B.; Slotboom, Dirk J.; Gelfand, Mikhail S.; Osterman, Andrei L.; Hanson, Andrew D.; Eitinger, Thomas

    2009-01-01

    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. PMID:18931129

  10. An ABC Transporter System of Yersinia pestis Allows Utilization of Chelated Iron by Escherichia coli SAB11

    PubMed Central

    Bearden, Scott W.; Staggs, Teanna M.; Perry, Robert D.

    1998-01-01

    The acquisition of iron is an essential component in the pathogenesis of Yersinia pestis, the agent of bubonic and pneumonic plague. A cosmid library derived from the genomic DNA of Y. pestis KIM6+ was used for transduction of an Escherichia coli mutant (SAB11) defective in the biosynthesis of the siderophore enterobactin. Recombinant plasmids which had a common 13-kb BamHI fragment were isolated from SAB11 transductants in which growth but not enterobactin synthesis was restored on media containing the iron chelator EDDA [ethylenediamine-di(o-hydroxyphenyl acetic acid)]. Subcloning and transposon mutagenesis revealed a 5.6-kb region, designated yfe, essential for SAB11 growth stimulation. In vitro transcription-translation analysis identified polypeptides of 18, 29.5, 32, and 33 kDa encoded by the yfe locus. Sequence analysis shows this locus to be comprised of five genes in two separate operons which have potential Fur-binding sequences in both promoters. A putative polycistronic operon, yfeABCD, is Fur regulated and responds to iron and manganese. A functional Fur protein is required for the observed manganese repression of this operon. This operon encodes polypeptides which have strong similarity to the ATP-binding cassette (ABC) family of transporters and include a periplasmic binding protein (YfeA), an ATP-binding protein (YfeB), and two integral membrane proteins (YfeC and -D), which likely function in the acquisition of inorganic iron and possibly other ions. The ∼21-kDa protein encoded by the separately transcribed yfeE gene may be located in the cell envelope, since a yfeE::TnphoA fusion is PhoA+. Mutations in this gene abrogate growth of SAB11 on iron-chelated media. PMID:9495751

  11. An ABC transporter system of Yersinia pestis allows utilization of chelated iron by Escherichia coli SAB11.

    PubMed

    Bearden, S W; Staggs, T M; Perry, R D

    1998-03-01

    The acquisition of iron is an essential component in the pathogenesis of Yersinia pestis, the agent of bubonic and pneumonic plague. A cosmid library derived from the genomic DNA of Y. pestis KIM6+ was used for transduction of an Escherichia coli mutant (SAB11) defective in the biosynthesis of the siderophore enterobactin. Recombinant plasmids which had a common 13-kb BamHI fragment were isolated from SAB11 transductants in which growth but not enterobactin synthesis was restored on media containing the iron chelator EDDA [ethylenediamine-di(o-hydroxyphenyl acetic acid)]. Subcloning and transposon mutagenesis revealed a 5.6-kb region, designated yfe, essential for SAB11 growth stimulation. In vitro transcription-translation analysis identified polypeptides of 18, 29.5, 32, and 33 kDa encoded by the yfe locus. Sequence analysis shows this locus to be comprised of five genes in two separate operons which have potential Fur-binding sequences in both promoters. A putative polycistronic operon, yfeABCD, is Fur regulated and responds to iron and manganese. A functional Fur protein is required for the observed manganese repression of this operon. This operon encodes polypeptides which have strong similarity to the ATP-binding cassette (ABC) family of transporters and include a periplasmic binding protein (YfeA), an ATP-binding protein (YfeB), and two integral membrane proteins (YfeC and -D), which likely function in the acquisition of inorganic iron and possibly other ions. The approximately 21-kDa protein encoded by the separately transcribed yfeE gene may be located in the cell envelope, since a yfeE::TnphoA fusion is PhoA+. Mutations in this gene abrogate growth of SAB11 on iron-chelated media.

  12. Multidrug Resistance Proteins (MRPs/ABCCs) in Cancer Chemotherapy and Genetic Diseases

    PubMed Central

    Chen, Zhe-Sheng; Tiwari, Amit K.

    2011-01-01

    The ATP-binding cassette (ABC) transporters are a superfamily of membrane proteins that are best known for their ability to transport a wide variety of exogenous and endogenous substances across membranes against a concentration gradient via ATP hydrolysis. There are seven subfamilies of human ABC transporters, one of the largest being the ‘C’ subfamily (gene symbol ABCC). Nine ABCC subfamily members, the so-called Multidrug Resistance Proteins (MRPs) 1-9, have been implicated in mediating multidrug resistance in tumor cells to varying degrees as the efflux extrude chemotherapeutic compounds (or their metabolites) from malignant cells. Some of the MRPs are also known to either influence drug disposition in normal tissues or modulate the elimination of drugs (or their metabolites) via hepatobiliary or renal excretory pathways. In addition, the cellular efflux of physiologically important organic anions such as leukotriene C4 and cAMP is mediated by one or more of the MRPs. Finally, mutations in several MRPs are associated with human genetic disorders. In this review article, the current biochemical and physiological knowledge of MRP1-MRP9 in cancer chemotherapy and human genetic disease is summarized. The mutations in MRP2/ABCC2 leading to conjugated hyperbilirubinemia (Dubin-Johnson syndrome) and in MRP6/ABCC6 leading to the connective tissue disorder Pseudoxanthoma elasticum are also discussed. PMID:21740521

  13. Checklist of the Diptera superfamilies Tephritoidea and Sciomyzoidea of Finland (Insecta)

    PubMed Central

    Kahanpää, Jere; Winqvist, Kaj

    2014-01-01

    Abstract A revised checklist of the flies of superfamilies Tephritoidea and Sciomyzoidea of Finland is provided. The following families are covered: Eurygnathomyiidae, Lonchaeidae, Neottiophilidae, Pallopteridae, Piophilidae, Platystomatidae, Tephritidae, Ulidiidae (Tephritoidea); Coelopidae, Dryomyzidae, Heterocheilidae, Phaeomyiidae, Sciomyzidae, Sepsidae (Sciomyzoidea). PMID:25337022

  14. A global view of structure–function relationships in the tautomerase superfamily

    PubMed Central

    Davidson, Rebecca; Baas, Bert-Jan; Akiva, Eyal; Holliday, Gemma L.; Polacco, Benjamin J.; LeVieux, Jake A.; Pullara, Collin R.; Zhang, Yan Jessie; Whitman, Christian P.

    2018-01-01

    The tautomerase superfamily (TSF) consists of more than 11,000 nonredundant sequences present throughout the biosphere. Characterized members have attracted much attention because of the unusual and key catalytic role of an N-terminal proline. These few characterized members catalyze a diverse range of chemical reactions, but the full scale of their chemical capabilities and biological functions remains unknown. To gain new insight into TSF structure–function relationships, we performed a global analysis of similarities across the entire superfamily and computed a sequence similarity network to guide classification into distinct subgroups. Our results indicate that TSF members are found in all domains of life, with most being present in bacteria. The eukaryotic members of the cis-3-chloroacrylic acid dehalogenase subgroup are limited to fungal species, whereas the macrophage migration inhibitory factor subgroup has wide eukaryotic representation (including mammals). Unexpectedly, we found that 346 TSF sequences lack Pro-1, of which 85% are present in the malonate semialdehyde decarboxylase subgroup. The computed network also enabled the identification of similarity paths, namely sequences that link functionally diverse subgroups and exhibit transitional structural features that may help explain reaction divergence. A structure-guided comparison of these linker proteins identified conserved transitions between them, and kinetic analysis paralleled these observations. Phylogenetic reconstruction of the linker set was consistent with these findings. Our results also suggest that contemporary TSF members may have evolved from a short 4-oxalocrotonate tautomerase–like ancestor followed by gene duplication and fusion. Our new linker-guided strategy can be used to enrich the discovery of sequence/structure/function transitions in other enzyme superfamilies. PMID:29184004

  15. Simulation-based estimation of mean and standard deviation for meta-analysis via Approximate Bayesian Computation (ABC).

    PubMed

    Kwon, Deukwoo; Reis, Isildinha M

    2015-08-12

    When conducting a meta-analysis of a continuous outcome, estimated means and standard deviations from the selected studies are required in order to obtain an overall estimate of the mean effect and its confidence interval. If these quantities are not directly reported in the publications, they must be estimated from other reported summary statistics, such as the median, the minimum, the maximum, and quartiles. We propose a simulation-based estimation approach using the Approximate Bayesian Computation (ABC) technique for estimating mean and standard deviation based on various sets of summary statistics found in published studies. We conduct a simulation study to compare the proposed ABC method with the existing methods of Hozo et al. (2005), Bland (2015), and Wan et al. (2014). In the estimation of the standard deviation, our ABC method performs better than the other methods when data are generated from skewed or heavy-tailed distributions. The corresponding average relative error (ARE) approaches zero as sample size increases. In data generated from the normal distribution, our ABC performs well. However, the Wan et al. method is best for estimating standard deviation under normal distribution. In the estimation of the mean, our ABC method is best regardless of assumed distribution. ABC is a flexible method for estimating the study-specific mean and standard deviation for meta-analysis, especially with underlying skewed or heavy-tailed distributions. The ABC method can be applied using other reported summary statistics such as the posterior mean and 95 % credible interval when Bayesian analysis has been employed.

  16. Functional significance of the E loop, a novel motif conserved in the lantibiotic immunity ATP-binding cassette transport systems.

    PubMed

    Okuda, Ken-ichi; Yanagihara, Sae; Sugayama, Tomomichi; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji

    2010-06-01

    Lantibiotics are peptide-derived antibacterial substances produced by some Gram-positive bacteria and characterized by the presence of unusual amino acids, like lanthionines and dehydrated amino acids. Because lantibiotic producers may be attacked by self-produced lantibiotics, they express immunity proteins on the cytoplasmic membrane. An ATP-binding cassette (ABC) transport system mediated by the LanFEG protein complex is a major system in lantibiotic immunity. Multiple-sequence alignment analysis revealed that LanF proteins contain the E loop, a variant of the Q loop, which is a well-conserved motif in the nucleotide-binding domains (NBDs) of general ABC transporters. To elucidate E loop function, we introduced a mutation in the NukF protein, which is involved in the nukacin-ISK-1 immunity system. Amino acid replacement of glutamic acid in the E loop with glutamine (E85Q) resulted in slight decreases in the immunity level and transport activity. Additionally, the E85A mutation severely impaired the immunity level and transport activity. On the other hand, ATPase activities of purified E85Q and E85A mutants were almost similar to that of the wild type. These results suggested that the E loop found in ABC transporters involved in lantibiotic immunity plays a significant role in the function of these transporters, especially in the structural change of transmembrane domains.

  17. New Enhanced Artificial Bee Colony (JA-ABC5) Algorithm with Application for Reactive Power Optimization

    PubMed Central

    2015-01-01

    The standard artificial bee colony (ABC) algorithm involves exploration and exploitation processes which need to be balanced for enhanced performance. This paper proposes a new modified ABC algorithm named JA-ABC5 to enhance convergence speed and improve the ability to reach the global optimum by balancing exploration and exploitation processes. New stages have been proposed at the earlier stages of the algorithm to increase the exploitation process. Besides that, modified mutation equations have also been introduced in the employed and onlooker-bees phases to balance the two processes. The performance of JA-ABC5 has been analyzed on 27 commonly used benchmark functions and tested to optimize the reactive power optimization problem. The performance results have clearly shown that the newly proposed algorithm has outperformed other compared algorithms in terms of convergence speed and global optimum achievement. PMID:25879054

  18. New enhanced artificial bee colony (JA-ABC5) algorithm with application for reactive power optimization.

    PubMed

    Sulaiman, Noorazliza; Mohamad-Saleh, Junita; Abro, Abdul Ghani

    2015-01-01

    The standard artificial bee colony (ABC) algorithm involves exploration and exploitation processes which need to be balanced for enhanced performance. This paper proposes a new modified ABC algorithm named JA-ABC5 to enhance convergence speed and improve the ability to reach the global optimum by balancing exploration and exploitation processes. New stages have been proposed at the earlier stages of the algorithm to increase the exploitation process. Besides that, modified mutation equations have also been introduced in the employed and onlooker-bees phases to balance the two processes. The performance of JA-ABC5 has been analyzed on 27 commonly used benchmark functions and tested to optimize the reactive power optimization problem. The performance results have clearly shown that the newly proposed algorithm has outperformed other compared algorithms in terms of convergence speed and global optimum achievement.

  19. Tungsten Transport Protein A (WtpA) in Pyrococcus furiosus: the First Member of a New Class of Tungstate and Molybdate Transporters

    PubMed Central

    Bevers, Loes E.; Hagedoorn, Peter-Leon; Krijger, Gerard C.; Hagen, Wilfred R.

    2006-01-01

    A novel tungstate and molybdate binding protein has been discovered from the hyperthermophilic archaeon Pyrococcus furiosus. This tungstate transport protein A (WtpA) is part of a new ABC transporter system selective for tungstate and molybdate. WtpA has very low sequence similarity with the earlier-characterized transport proteins ModA for molybdate and TupA for tungstate. Its structural gene is present in the genome of numerous archaea and some bacteria. The identification of this new tungstate and molybdate binding protein clarifies the mechanism of tungstate and molybdate transport in organisms that lack the known uptake systems associated with the ModA and TupA proteins, like many archaea. The periplasmic protein of this ABC transporter, WtpA (PF0080), was cloned and expressed in Escherichia coli. Using isothermal titration calorimetry, WtpA was observed to bind tungstate (dissociation constant [KD] of 17 ± 7 pM) and molybdate (KD of 11 ± 5 nM) with a stoichiometry of 1.0 mol oxoanion per mole of protein. These low KD values indicate that WtpA has a higher affinity for tungstate than do ModA and TupA and an affinity for molybdate similar to that of ModA. A displacement titration of molybdate-saturated WtpA with tungstate showed that the tungstate effectively replaced the molybdate in the binding site of the protein. PMID:16952940

  20. Discovering Novel Bile Protection Systems in Bifidobacterium breve UCC2003 through Functional Genomics

    PubMed Central

    Ruiz, Lorena; Zomer, Aldert; O'Connell-Motherway, Mary; van Sinderen, Douwe

    2012-01-01

    Tolerance of gut commensals to bile salt exposure is an important feature for their survival in and colonization of the intestinal environment. A transcriptomic approach was employed to study the response of Bifidobacterium breve UCC2003 to bile, allowing the identification of a number of bile-induced genes with a range of predicted functions. The potential roles of a selection of these bile-inducible genes in bile protection were analyzed following heterologous expression in Lactococcus lactis. Genes encoding three transport systems belonging to the major facilitator superfamily (MFS), Bbr_0838, Bbr_0832, and Bbr_1756, and three ABC-type transporters, Bbr_0406-0407, Bbr_1804-1805, and Bbr_1826-1827, were thus investigated and shown to provide enhanced resistance and survival to bile exposure. This work significantly improves our understanding as to how bifidobacteria respond to and survive bile exposure. PMID:22156415

  1. Forkhead-associated (FHA) Domain Containing ABC Transporter Rv1747 Is Positively Regulated by Ser/Thr Phosphorylation in Mycobacterium tuberculosis*

    PubMed Central

    Spivey, Vicky L.; Molle, Virginie; Whalan, Rachael H.; Rodgers, Angela; Leiba, Jade; Stach, Lasse; Walker, K. Barry; Smerdon, Stephen J.; Buxton, Roger S.

    2011-01-01

    One major signaling method employed by Mycobacterium tuberculosis, the causative agent of tuberculosis, is through reversible phosphorylation of proteins mediated by protein kinases and phosphatases. This study concerns one of these enzymes, the serine/threonine protein kinase PknF, that is encoded in an operon with Rv1747, an ABC transporter that is necessary for growth of M. tuberculosis in vivo and contains two forkhead-associated (FHA) domains. FHA domains are phosphopeptide recognition motifs that specifically recognize phosphothreonine-containing epitopes. Experiments to determine how PknF regulates the function of Rv1747 demonstrated that phosphorylation occurs on two specific threonine residues, Thr-150 and Thr-208. To determine the in vivo consequences of phosphorylation, infection experiments were performed in bone marrow-derived macrophages and in mice using threonine-to-alanine mutants of Rv1747 that prevent specific phosphorylation and revealed that phosphorylation positively modulates Rv1747 function in vivo. The role of the FHA domains in this regulation was further demonstrated by isothermal titration calorimetry, using peptides containing both phosphothreonine residues. FHA-1 domain mutation resulted in attenuation in macrophages highlighting the critical role of this domain in Rv1747 function. A mutant deleted for pknF did not, however, have a growth phenotype in an infection, suggesting that other kinases can fulfill its role when it is absent. This study provides the first information on the molecular mechanism(s) regulating Rv1747 through PknF-dependent phosphorylation but also indicates that phosphorylation activates Rv1747, which may have important consequences in regulating growth of M. tuberculosis. PMID:21622570

  2. Comparison between the Movement ABC-2 and the Zurich Neuromotor Assessment in Preschool Children.

    PubMed

    Kakebeeke, Tanja H; Knaier, Elisa; Köchli, Sabrina; Chaouch, Aziz; Rousson, Valentin; Kriemler, Susi; Jenni, Oskar G

    2016-12-01

    An established test instrument for the assessment of motor performance in children between 3 and 16 years is the Movement Assessment Battery for Children - Second Edition (M-ABC-2). The Zurich Neuromotor Assessment (ZNA) is also widely used for the evaluation of children's motor performance but has not been compared with the M-ABC-2 for children below five years for the purpose of convergent validity. Forty-seven children (26 boys, 21 girls) between three and five years of age were assessed using the M-ABC-2 and the ZNA3-5. Rank correlations between scores of different test components were calculated. Only low-to-moderate correlations were observed when separate components of these tests were compared (.31 to .68, p < .05), especially when involving the associated movements from the ZNA3-5 (-.05 to -.13, p > .05). However, the correlation between summary scores of the two tests was .77 (p < .001), and it increased to .84 when associated movements were excluded, which was comparable in magnitude to the test-retest reliability of the M-ABC-2, supporting convergent validity between the two tests. Although the ZNA3-5 and M-ABC-2 measure different aspects of motor behavior, the two instruments may thus measure essentially the same construct. © The Author(s) 2016.

  3. Neuroprotective effect of chondroitinase ABC on primary and secondary brain injury after stroke in hypertensive rats.

    PubMed

    Chen, Xin-ran; Liao, Song-jie; Ye, Lan-xiang; Gong, Qiong; Ding, Qiao; Zeng, Jin-sheng; Yu, Jian

    2014-01-16

    Focal cerebral infarction causes secondary damage in the ipsilateral ventroposterior thalamic nucleus (VPN). Chondroitin sulfate proteoglycans (CSPGs) are a family of putative inhibitory components, and its degradation by chondroitinase ABC (ChABC) promotes post-injury neurogenesis. This study investigated the role of ChABC in the primary and secondary injury post stroke in hypertension. Renovascular hypertensive Sprague-Dawley rats underwent middle cerebral artery occlusion (MCAO), and were subjected to continuous intra-infarct infusion of ChABC (0.12 U/d for 7 days) 24 h later. Neurological function was evaluated by a modified neurologic severity score. Neurons were counted in the peri-infarct region and the ipsilateral VPN 8 and 14 days after MCAO by Nissl staining and NeuN labeling. The expressions of CSPGs, growth-associated protein-43 (GAP-43) and synaptophysin (SYN) were detected with immunofluorescence or Western blotting. The intra-infarct infusion of ChABC, by degrading accumulated CSPGs, rescued neuronal loss and increased the levels of GAP-43 and SYN in both the ipsilateral cortex and VPN, indicating enhancd neuron survival as well as augmented axonal growth and synaptic plasticity, eventually improving overall neurological function. The study demonstrated that intra-infarct ChABC infusion could salvage the brain from both primary and secondary injury by the intervention on the neuroinhibitory environment post focal cerebral infarction. © 2013 Published by Elsevier B.V.

  4. Homologues of the human multidrug resistance genes MRP and MDR contribute to heavy metal resistance in the soil nematode Caenorhabditis elegans.

    PubMed Central

    Broeks, A; Gerrard, B; Allikmets, R; Dean, M; Plasterk, R H

    1996-01-01

    Acquired resistance of mammalian cells to multiple chemotherapeutic drugs can result from enhanced expression of the multidrug resistance-associated protein (MRP), which belongs to the ABC transporter superfamily. ABC transporters play a role in the protection of organisms against exogenous toxins by cellular detoxification processes. We have identified four MRP homologues in the soil nematode Caenorhabditis elegans, and we have studied one member, mrp-1, in detail. Using an mrp::lacZ gene fusion, mrp-l expression was found in cells of the pharynx, the pharynx-intestinal valve and the anterior intestinal cells, the rectum-intestinal valve and the epithelial cells of the vulva. Targeted inactivation of mrp-l resulted in increased sensitivity to the heavy metal ions cadmium and arsenite, to which wild-type worms are highly tolerant. The most pronounced effect of the mrp-1 mutation is on the ability of animals to recover from temporary exposure to high concentrations of heavy metals. Nematodes were found to be hypersensitive to heavy metals when both the MRP homologue, mrp-1, and a member of the P-glycoprotein (Pgp) gene family, pgp-1, were deleted. We conclude that nematodes have multiple proteins, homologues of mammalian proteins involved in the cellular resistance to chemotherapeutic drugs, that protect them against heavy metals. Images PMID:8947035

  5. The Role of Activity Based Costing (ABC) in Educational Support Services: A White Paper.

    ERIC Educational Resources Information Center

    Edds, Daniel B.

    Many front-line managers who are assuming more financial responsibility for their organizations find traditional cost accounting inadequate for their needs and are turning to Activity Based Costing (ABC). ABC is not a financial reporting system to serve the needs of regulatory agencies, but a tool that tracks costs from the general ledger…

  6. Effect of SOS-induced levels of imuABC on spontaneous and damage-induced mutagenesis in Caulobacter crescentus.

    PubMed

    Alves, Ingrid R; Lima-Noronha, Marco A; Silva, Larissa G; Fernández-Silva, Frank S; Freitas, Aline Luiza D; Marques, Marilis V; Galhardo, Rodrigo S

    2017-11-01

    imuABC (imuAB dnaE2) genes are responsible for SOS-mutagenesis in Caulobacter crescentus and other bacterial species devoid of umuDC. In this work, we have constructed operator-constitutive mutants of the imuABC operon. We used this genetic tool to investigate the effect of SOS-induced levels of these genes upon both spontaneous and damage-induced mutagenesis. We showed that constitutive expression of imuABC does not increase spontaneous or damage-induced mutagenesis, nor increases cellular resistance to DNA-damaging agents. Nevertheless, the presence of the operator-constitutive mutation rescues mutagenesis in a recA background, indicating that imuABC are the only genes required at SOS-induced levels for translesion synthesis (TLS) in C. crescentus. Furthermore, these data also show that TLS mediated by ImuABC does not require RecA, unlike umuDC-dependent mutagenesis in E. coli. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. IMG-ABC. A knowledge base to fuel discovery of biosynthetic gene clusters and novel secondary metabolites

    DOE PAGES

    Hadjithomas, Michalis; Chen, I-Min Amy; Chu, Ken; ...

    2015-07-14

    In the discovery of secondary metabolites, analysis of sequence data is a promising exploration path that remains largely underutilized due to the lack of computational platforms that enable such a systematic approach on a large scale. In this work, we present IMG-ABC (https://img.jgi.doe.gov/abc), an atlas of biosynthetic gene clusters within the Integrated Microbial Genomes (IMG) system, which is aimed at harnessing the power of “big” genomic data for discovering small molecules. IMG-ABC relies on IMG’s comprehensive integrated structural and functional genomic data for the analysis of biosynthetic gene clusters (BCs) and associated secondary metabolites (SMs). SMs and BCs serve asmore » the two main classes of objects in IMG-ABC, each with a rich collection of attributes. A unique feature of IMG-ABC is the incorporation of both experimentally validated and computationally predicted BCs in genomes as well as metagenomes, thus identifying BCs in uncultured populations and rare taxa. We demonstrate the strength of IMG-ABC’s focused integrated analysis tools in enabling the exploration of microbial secondary metabolism on a global scale, through the discovery of phenazine-producing clusters for the first time in lphaproteobacteria. IMG-ABC strives to fill the long-existent void of resources for computational exploration of the secondary metabolism universe; its underlying scalable framework enables traversal of uncovered phylogenetic and chemical structure space, serving as a doorway to a new era in the discovery of novel molecules. IMG-ABC is the largest publicly available database of predicted and experimental biosynthetic gene clusters and the secondary metabolites they produce. The system also includes powerful search and analysis tools that are integrated with IMG’s extensive genomic/metagenomic data and analysis tool kits. As new research on biosynthetic gene clusters and secondary metabolites is published and more genomes are sequenced, IMG-ABC

  8. ABCA Transporter Gene Expression and Poor Outcome in Epithelial Ovarian Cancer

    PubMed Central

    Hedditch, Ellen L.; Gao, Bo; Russell, Amanda J.; Lu, Yi; Emmanuel, Catherine; Beesley, Jonathan; Johnatty, Sharon E.; Chen, Xiaoqing; Harnett, Paul; George, Joshy; Williams, Rebekka T.; Flemming, Claudia; Lambrechts, Diether; Despierre, Evelyn; Lambrechts, Sandrina; Vergote, Ignace; Karlan, Beth; Lester, Jenny; Orsulic, Sandra; Walsh, Christine; Fasching, Peter; Beckmann, Matthias W.; Ekici, Arif B.; Hein, Alexander; Matsuo, Keitaro; Hosono, Satoyo; Nakanishi, Toru; Yatabe, Yasushi; Pejovic, Tanja; Bean, Yukie; Heitz, Florian; Harter, Philipp; du Bois, Andreas; Schwaab, Ira; Hogdall, Estrid; Kjaer, Susan K.; Jensen, Allan; Hogdall, Claus; Lundvall, Lene; Engelholm, Svend Aage; Brown, Bob; Flanagan, James; Metcalf, Michelle D; Siddiqui, Nadeem; Sellers, Thomas; Fridley, Brooke; Cunningham, Julie; Schildkraut, Joellen; Iversen, Ed; Weber, Rachel P.; Berchuck, Andrew; Goode, Ellen; Bowtell, David D.; Chenevix-Trench, Georgia; deFazio, Anna; Norris, Murray D.; MacGregor, Stuart; Haber, Michelle; Henderson, Michelle J.

    2014-01-01

    Background ATP-binding cassette (ABC) transporters play various roles in cancer biology and drug resistance, but their association with outcomes in serous epithelial ovarian cancer (EOC) is unknown. Methods The relationship between clinical outcomes and ABC transporter gene expression in two independent cohorts of high-grade serous EOC tumors was assessed with real-time quantitative polymerase chain reaction, analysis of expression microarray data, and immunohistochemistry. Associations between clinical outcomes and ABCA transporter gene single nucleotide polymorphisms were tested in a genome-wide association study. Impact of short interfering RNA–mediated gene suppression was determined by colony forming and migration assays. Association with survival was assessed with Kaplan–Meier analysis and log-rank tests. All statistical tests were two-sided. Results Associations with outcome were observed with ABC transporters of the “A” subfamily, but not with multidrug transporters. High-level expression of ABCA1, ABCA6, ABCA8, and ABCA9 in primary tumors was statistically significantly associated with reduced survival in serous ovarian cancer patients. Low levels of ABCA5 and the C-allele of rs536009 were associated with shorter overall survival (hazard ratio for death = 1.50; 95% confidence interval [CI] =1.26 to 1.79; P = 6.5e−6). The combined expression pattern of ABCA1, ABCA5, and either ABCA8 or ABCA9 was associated with particularly poor outcome (mean overall survival in group with adverse ABCA1, ABCA5 and ABCA9 gene expression = 33.2 months, 95% CI = 26.4 to 40.1; vs 55.3 months in the group with favorable ABCA gene expression, 95% CI = 49.8 to 60.8; P = .001), independently of tumor stage or surgical debulking status. Suppression of cholesterol transporter ABCA1 inhibited ovarian cancer cell growth and migration in vitro, and statin treatment reduced ovarian cancer cell migration. Conclusions Expression of ABCA transporters was associated with poor

  9. ABCA transporter gene expression and poor outcome in epithelial ovarian cancer.

    PubMed

    Hedditch, Ellen L; Gao, Bo; Russell, Amanda J; Lu, Yi; Emmanuel, Catherine; Beesley, Jonathan; Johnatty, Sharon E; Chen, Xiaoqing; Harnett, Paul; George, Joshy; Williams, Rebekka T; Flemming, Claudia; Lambrechts, Diether; Despierre, Evelyn; Lambrechts, Sandrina; Vergote, Ignace; Karlan, Beth; Lester, Jenny; Orsulic, Sandra; Walsh, Christine; Fasching, Peter; Beckmann, Matthias W; Ekici, Arif B; Hein, Alexander; Matsuo, Keitaro; Hosono, Satoyo; Nakanishi, Toru; Yatabe, Yasushi; Pejovic, Tanja; Bean, Yukie; Heitz, Florian; Harter, Philipp; du Bois, Andreas; Schwaab, Ira; Hogdall, Estrid; Kjaer, Susan K; Jensen, Allan; Hogdall, Claus; Lundvall, Lene; Engelholm, Svend Aage; Brown, Bob; Flanagan, James; Metcalf, Michelle D; Siddiqui, Nadeem; Sellers, Thomas; Fridley, Brooke; Cunningham, Julie; Schildkraut, Joellen; Iversen, Ed; Weber, Rachel P; Berchuck, Andrew; Goode, Ellen; Bowtell, David D; Chenevix-Trench, Georgia; deFazio, Anna; Norris, Murray D; MacGregor, Stuart; Haber, Michelle; Henderson, Michelle J

    2014-07-01

    ATP-binding cassette (ABC) transporters play various roles in cancer biology and drug resistance, but their association with outcomes in serous epithelial ovarian cancer (EOC) is unknown. The relationship between clinical outcomes and ABC transporter gene expression in two independent cohorts of high-grade serous EOC tumors was assessed with real-time quantitative polymerase chain reaction, analysis of expression microarray data, and immunohistochemistry. Associations between clinical outcomes and ABCA transporter gene single nucleotide polymorphisms were tested in a genome-wide association study. Impact of short interfering RNA-mediated gene suppression was determined by colony forming and migration assays. Association with survival was assessed with Kaplan-Meier analysis and log-rank tests. All statistical tests were two-sided. Associations with outcome were observed with ABC transporters of the "A" subfamily, but not with multidrug transporters. High-level expression of ABCA1, ABCA6, ABCA8, and ABCA9 in primary tumors was statistically significantly associated with reduced survival in serous ovarian cancer patients. Low levels of ABCA5 and the C-allele of rs536009 were associated with shorter overall survival (hazard ratio for death = 1.50; 95% confidence interval [CI] =1.26 to 1.79; P = 6.5e-6). The combined expression pattern of ABCA1, ABCA5, and either ABCA8 or ABCA9 was associated with particularly poor outcome (mean overall survival in group with adverse ABCA1, ABCA5 and ABCA9 gene expression = 33.2 months, 95% CI = 26.4 to 40.1; vs 55.3 months in the group with favorable ABCA gene expression, 95% CI = 49.8 to 60.8; P = .001), independently of tumor stage or surgical debulking status. Suppression of cholesterol transporter ABCA1 inhibited ovarian cancer cell growth and migration in vitro, and statin treatment reduced ovarian cancer cell migration. Expression of ABCA transporters was associated with poor outcome in serous ovarian cancer, implicating lipid

  10. Calculus ABCs: A Gateway for Freshman Calculus

    ERIC Educational Resources Information Center

    Fulton, Scott R.

    2003-01-01

    This paper describes a gateway testing program designed to ensure that students acquire basic skills in freshman calculus. Students must demonstrate they have mastered standards for "Absolutely Basic Competency"--the Calculus ABCs--in order to pass the course with a grade of C or better. We describe the background, standards, and testing program.…

  11. Sustainable urban stormwater management in the tropics: An evaluation of Singapore's ABC Waters Program

    NASA Astrophysics Data System (ADS)

    Lim, H. S.; Lu, X. X.

    2016-07-01

    The Active Beautiful Clean (ABC) Waters Program was implemented in 2006 as part of Singapore's stormwater management strategy and reflects the country's move towards Water Sensitive Urbanism through the adoption of Low-Impact Development (LID) ideology and practices. It is the first holistic and comprehensive LID program in the tropics and holds promise for extension to other tropical cities. This paper presents a comprehensive summary of the goals, LID practices (ABC design features) and design considerations as well as results of several monitored sites, including a constructed wetland, two rain gardens, green roofs and three canal restoration projects. We evaluate the ABC Waters Program based on these initial results and consider the challenges, issues and the research needs for it to meet its hydrological and water quality remediation goals. So far, the ABC design features evaluated perform well in removing particulates. Performance in nutrient removal is poor. With over 60 projects completed within 10 years, post-project monitoring and evaluation is necessary and complements on-going laboratory and modelling research projects conducted by local academic institutions.

  12. Ensembler: Enabling High-Throughput Molecular Simulations at the Superfamily Scale.

    PubMed

    Parton, Daniel L; Grinaway, Patrick B; Hanson, Sonya M; Beauchamp, Kyle A; Chodera, John D

    2016-06-01

    The rapidly expanding body of available genomic and protein structural data provides a rich resource for understanding protein dynamics with biomolecular simulation. While computational infrastructure has grown rapidly, simulations on an omics scale are not yet widespread, primarily because software infrastructure to enable simulations at this scale has not kept pace. It should now be possible to study protein dynamics across entire (super)families, exploiting both available structural biology data and conformational similarities across homologous proteins. Here, we present a new tool for enabling high-throughput simulation in the genomics era. Ensembler takes any set of sequences-from a single sequence to an entire superfamily-and shepherds them through various stages of modeling and refinement to produce simulation-ready structures. This includes comparative modeling to all relevant PDB structures (which may span multiple conformational states of interest), reconstruction of missing loops, addition of missing atoms, culling of nearly identical structures, assignment of appropriate protonation states, solvation in explicit solvent, and refinement and filtering with molecular simulation to ensure stable simulation. The output of this pipeline is an ensemble of structures ready for subsequent molecular simulations using computer clusters, supercomputers, or distributed computing projects like Folding@home. Ensembler thus automates much of the time-consuming process of preparing protein models suitable for simulation, while allowing scalability up to entire superfamilies. A particular advantage of this approach can be found in the construction of kinetic models of conformational dynamics-such as Markov state models (MSMs)-which benefit from a diverse array of initial configurations that span the accessible conformational states to aid sampling. We demonstrate the power of this approach by constructing models for all catalytic domains in the human tyrosine kinase

  13. RCN1/OsABCG5, an ATP-binding cassette (ABC) transporter, is required for hypodermal suberization of roots in rice (Oryza sativa).

    PubMed

    Shiono, Katsuhiro; Ando, Miho; Nishiuchi, Shunsaku; Takahashi, Hirokazu; Watanabe, Kohtaro; Nakamura, Motoaki; Matsuo, Yuichi; Yasuno, Naoko; Yamanouchi, Utako; Fujimoto, Masaru; Takanashi, Hideki; Ranathunge, Kosala; Franke, Rochus B; Shitan, Nobukazu; Nishizawa, Naoko K; Takamure, Itsuro; Yano, Masahiro; Tsutsumi, Nobuhiro; Schreiber, Lukas; Yazaki, Kazufumi; Nakazono, Mikio; Kato, Kiyoaki

    2014-10-01

    Suberin is a complex polymer composed of aliphatic and phenolic compounds. It is a constituent of apoplastic plant interfaces. In many plant species, including rice (Oryza sativa), the hypodermis in the outer part of roots forms a suberized cell wall (the Casparian strip and/or suberin lamellae), which inhibits the flow of water and ions and protects against pathogens. To date, there is no genetic evidence that suberin forms an apoplastic transport barrier in the hypodermis. We discovered that a rice reduced culm number1 (rcn1) mutant could not develop roots longer than 100 mm in waterlogged soil. The mutated gene encoded an ATP-binding cassette (ABC) transporter named RCN1/OsABCG5. RCN1/OsABCG5 gene expression in the wild type was increased in most hypodermal and some endodermal roots cells under stagnant deoxygenated conditions. A GFP-RCN1/OsABCG5 fusion protein localized at the plasma membrane of the wild type. Under stagnant deoxygenated conditions, well suberized hypodermis developed in wild types but not in rcn1 mutants. Under stagnant deoxygenated conditions, apoplastic tracers (periodic acid and berberine) were blocked at the hypodermis in the wild type but not in rcn1, indicating that the apoplastic barrier in the mutant was impaired. The amount of the major aliphatic suberin monomers originating from C(28) and C(30) fatty acids or ω-OH fatty acids was much lower in rcn1 than in the wild type. These findings suggest that RCN1/OsABCG5 has a role in the suberization of the hypodermis of rice roots, which contributes to formation of the apoplastic barrier. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  14. ABC's of Being Smart: I Can "C" Clearly Now

    ERIC Educational Resources Information Center

    Foster, Joanne

    2011-01-01

    In this paper, the author focuses on C of the ABC's of being smart. She continues to categorize the points for readers. These categories include the following: (1) being; (2) doing; and (3) stretching.

  15. Compilation of accelerated bridge construction (ABC) bridges : final report.

    DOT National Transportation Integrated Search

    2016-01-01

    Development of accelerated bridge construction (ABC) technologies has been occurring across the country, many times in : isolation. Although FHWA and others have worked to facilitate communication between these efforts, there was not a : comprehensiv...

  16. The intriguing biology of the tumour necrosis factor/tumour necrosis factor receptor superfamily: players, rules and the games.

    PubMed

    Hehlgans, Thomas; Pfeffer, Klaus

    2005-05-01

    The members of the tumour necrosis factor (TNF)/tumour necrosis factor receptor (TNFR) superfamily are critically involved in the maintenance of homeostasis of the immune system. The biological functions of this system encompass beneficial and protective effects in inflammation and host defence as well as a crucial role in organogenesis. At the same time, members of this superfamily are responsible for host damaging effects in sepsis, cachexia, and autoimmune diseases. This review summarizes recent progress in the immunobiology of the TNF/TNFR superfamily focusing on results obtained from animal studies using gene targeted mice. The different modes of signalling pathways affecting cell proliferation, survival, differentiation, apoptosis, and immune organ development as well as host defence are reviewed. Molecular and cellular mechanisms that demonstrate a therapeutic potential by targeting individual receptors or ligands for the treatment of chronic inflammatory or autoimmune diseases are discussed.

  17. The ABC model of recombination reinterpreted: Impact on understanding carrier transport and efficiency droop in InGaN/GaN light emitting diodes

    NASA Astrophysics Data System (ADS)

    Hopkins, M. A.; Allsopp, D. W. E.; Kappers, M. J.; Oliver, R. A.; Humphreys, C. J.

    2017-12-01

    The efficiency of light emitting diodes (LEDs) remains a topic of great contemporary interest due to their potential to reduce the amount of energy consumed in lighting. The current consensus is that electrons and holes distribute themselves through the emissive region by a drift-diffusion process which results in a highly non-uniform distribution of the light emission and can reduce efficiency. In this paper, the measured variations in the external quantum efficiency of a range of InGaN/GaN LEDs with different numbers of quantum wells (QWs) are shown to compare closely with the predictions of a revised ABC model, in which it is assumed that the electrically injected electrons and holes are uniformly distributed through the multi-quantum well (MQW) region, or nearly so, and hence carrier recombination occurs equally in all the quantum wells. The implications of the reported results are that drift-diffusion plays a far lesser role in cross-well carrier transport than previously thought; that the dominant cause of efficiency droop is intrinsic to the quantum wells and that reductions in the density of non-radiative recombination centers in the MQW would enable the use of more QWs and thereby reduce Auger losses by spreading carriers more evenly across a wider emissive region.

  18. Protein complexes formed during the incision reaction catalyzed by the Escherichia coli UvrABC endonuclease.

    PubMed Central

    Yeung, A T; Mattes, W B; Grossman, L

    1986-01-01

    An examination has been made into the nature of the nucleoprotein complexes formed during the incision reaction catalyzed by the Escherichia coli UvrABC endonuclease when acting on a pyrimidine dimer-containing fd RF-I DNA species. The complexes of proteins and DNA form in unique stages. The first stage of binding involves an ATP-stimulated interaction of the UvrA protein with duplex DNA containing pyrimidine dimer sites. The UvrB protein significantly stabilizes the UvrA-pyrimidine dimer containing DNA complex which, in turn, provides a foundation for the binding of UvrC to activate the UvrABC endonuclease. The binding of one molecule of UvrC to each UvrAB-damaged DNA complex is needed to catalyze incision in the vicinity of pyrimidine dimer sites. The UvrABC-DNA complex persists after the incision event suggesting that the lack of UvrABC turnover may be linked to other activities in the excision-repair pathway beyond the initial incision reaction. PMID:3960727

  19. Classification of E-Nose Aroma Data of Four Fruit Types by ABC-Based Neural Network

    PubMed Central

    Adak, M. Fatih; Yumusak, Nejat

    2016-01-01

    Electronic nose technology is used in many areas, and frequently in the beverage industry for classification and quality-control purposes. In this study, four different aroma data (strawberry, lemon, cherry, and melon) were obtained using a MOSES II electronic nose for the purpose of fruit classification. To improve the performance of the classification, the training phase of the neural network with two hidden layers was optimized using artificial bee colony algorithm (ABC), which is known to be successful in exploration. Test data were given to two different neural networks, each of which were trained separately with backpropagation (BP) and ABC, and average test performances were measured as 60% for the artificial neural network trained with BP and 76.39% for the artificial neural network trained with ABC. Training and test phases were repeated 30 times to obtain these average performance measurements. This level of performance shows that the artificial neural network trained with ABC is successful in classifying aroma data. PMID:26927124

  20. Classification of E-Nose Aroma Data of Four Fruit Types by ABC-Based Neural Network.

    PubMed

    Adak, M Fatih; Yumusak, Nejat

    2016-02-27

    Electronic nose technology is used in many areas, and frequently in the beverage industry for classification and quality-control purposes. In this study, four different aroma data (strawberry, lemon, cherry, and melon) were obtained using a MOSES II electronic nose for the purpose of fruit classification. To improve the performance of the classification, the training phase of the neural network with two hidden layers was optimized using artificial bee colony algorithm (ABC), which is known to be successful in exploration. Test data were given to two different neural networks, each of which were trained separately with backpropagation (BP) and ABC, and average test performances were measured as 60% for the artificial neural network trained with BP and 76.39% for the artificial neural network trained with ABC. Training and test phases were repeated 30 times to obtain these average performance measurements. This level of performance shows that the artificial neural network trained with ABC is successful in classifying aroma data.

  1. Volume Quantification of Acute Infratentorial Hemorrhage with Computed Tomography: Validation of the Formula 1/2ABC and 2/3SH

    PubMed Central

    Zhang, Yunyun; Yan, Jing; Fu, Yi; Chen, Shengdi

    2013-01-01

    Objective To compare the accuracy of formula 1/2ABC with 2/3SH on volume estimation for hypertensive infratentorial hematoma. Methods One hundred and forty-seven CT scans diagnosed as hypertensive infratentorial hemorrhage were reviewed. Based on the shape, hematomas were categorized as regular or irregular. Multilobular was defined as a special shape of irregular. Hematoma volume was calculated employing computer-assisted volumetric analysis (CAVA), 1/2ABC and 2/3SH, respectively. Results The correlation coefficients between 1/2ABC (or 2/3SH) and CAVA were greater than 0.900 in all subgroups. There were neither significant differences in absolute values of volume deviation nor percentage deviation between 1/2ABC and 2/3SH for regular hemorrhage (P>0.05). While for cerebellar, brainstem and irregular hemorrhages, the absolute values of volume deviation and percentage deviation by formula 1/2ABC were greater than 2/3SH (P<0.05). 1/2ABC and 2/3SH underestimated hematoma volume each by 10% and 5% for cerebellar hemorrhage, 14% and 9% for brainstem hemorrhage, 19% and 16% for regular hemorrhage, 9% and 3% for irregular hemorrhage, respectively. In addition, for the multilobular hemorrhage, 1/2ABC underestimated the volume by 9% while 2/3SH overestimated it by 2%. Conclusions For regular hemorrhage volume calculation, the accuracy of 2/3SH is similar to 1/2ABC. While for cerebellar, brainstem or irregular hemorrhages (including multilobular), 2/3SH is more accurate than 1/2ABC. PMID:23638025

  2. So, You Need To Justify Your Existing ABC Program (or Lobby for a New One).

    ERIC Educational Resources Information Center

    Walsh, Jean Terry; Gillis, Lee

    1998-01-01

    Advice for adventure-based counseling (ABC) programs seeking funding includes setting realistic goals, designing an evaluation that matches program resources, and keeping it simple. Low recidivism is most important to grantors. Published research on ABC is scarce, but on-site process research generates useful data, and local schools and agencies…

  3. Membrane and Protein Interactions of the Pleckstrin Homology Domain Superfamily

    PubMed Central

    Lenoir, Marc; Kufareva, Irina; Abagyan, Ruben; Overduin, Michael

    2015-01-01

    The human genome encodes about 285 proteins that contain at least one annotated pleckstrin homology (PH) domain. As the first phosphoinositide binding module domain to be discovered, the PH domain recruits diverse protein architectures to cellular membranes. PH domains constitute one of the largest protein superfamilies, and have diverged to regulate many different signaling proteins and modules such as Dbl homology (DH) and Tec homology (TH) domains. The ligands of approximately 70 PH domains have been validated by binding assays and complexed structures, allowing meaningful extrapolation across the entire superfamily. Here the Membrane Optimal Docking Area (MODA) program is used at a genome-wide level to identify all membrane docking PH structures and map their lipid-binding determinants. In addition to the linear sequence motifs which are employed for phosphoinositide recognition, the three dimensional structural features that allow peripheral membrane domains to approach and insert into the bilayer are pinpointed and can be predicted ab initio. The analysis shows that conserved structural surfaces distinguish which PH domains associate with membrane from those that do not. Moreover, the results indicate that lipid-binding PH domains can be classified into different functional subgroups based on the type of membrane insertion elements they project towards the bilayer. PMID:26512702

  4. Diversity, classification and function of the plant protein kinase superfamily

    PubMed Central

    Lehti-Shiu, Melissa D.; Shiu, Shin-Han

    2012-01-01

    Eukaryotic protein kinases belong to a large superfamily with hundreds to thousands of copies and are components of essentially all cellular functions. The goals of this study are to classify protein kinases from 25 plant species and to assess their evolutionary history in conjunction with consideration of their molecular functions. The protein kinase superfamily has expanded in the flowering plant lineage, in part through recent duplications. As a result, the flowering plant protein kinase repertoire, or kinome, is in general significantly larger than other eukaryotes, ranging in size from 600 to 2500 members. This large variation in kinome size is mainly due to the expansion and contraction of a few families, particularly the receptor-like kinase/Pelle family. A number of protein kinases reside in highly conserved, low copy number families and often play broadly conserved regulatory roles in metabolism and cell division, although functions of plant homologues have often diverged from their metazoan counterparts. Members of expanded plant kinase families often have roles in plant-specific processes and some may have contributed to adaptive evolution. Nonetheless, non-adaptive explanations, such as kinase duplicate subfunctionalization and insufficient time for pseudogenization, may also contribute to the large number of seemingly functional protein kinases in plants. PMID:22889912

  5. Taxonomic distribution and origins of the extended LHC (light-harvesting complex) antenna protein superfamily

    PubMed Central

    2010-01-01

    Background The extended light-harvesting complex (LHC) protein superfamily is a centerpiece of eukaryotic photosynthesis, comprising the LHC family and several families involved in photoprotection, like the LHC-like and the photosystem II subunit S (PSBS). The evolution of this complex superfamily has long remained elusive, partially due to previously missing families. Results In this study we present a meticulous search for LHC-like sequences in public genome and expressed sequence tag databases covering twelve representative photosynthetic eukaryotes from the three primary lineages of plants (Plantae): glaucophytes, red algae and green plants (Viridiplantae). By introducing a coherent classification of the different protein families based on both, hidden Markov model analyses and structural predictions, numerous new LHC-like sequences were identified and several new families were described, including the red lineage chlorophyll a/b-binding-like protein (RedCAP) family from red algae and diatoms. The test of alternative topologies of sequences of the highly conserved chlorophyll-binding core structure of LHC and PSBS proteins significantly supports the independent origins of LHC and PSBS families via two unrelated internal gene duplication events. This result was confirmed by the application of cluster likelihood mapping. Conclusions The independent evolution of LHC and PSBS families is supported by strong phylogenetic evidence. In addition, a possible origin of LHC and PSBS families from different homologous members of the stress-enhanced protein subfamily, a diverse and anciently paralogous group of two-helix proteins, seems likely. The new hypothesis for the evolution of the extended LHC protein superfamily proposed here is in agreement with the character evolution analysis that incorporates the distribution of families and subfamilies across taxonomic lineages. Intriguingly, stress-enhanced proteins, which are universally found in the genomes of green plants

  6. An Efficient Strategy for Small-Scale Screening and Production of Archaeal Membrane Transport Proteins in Escherichia coli

    PubMed Central

    Ma, Pikyee; Varela, Filipa; Magoch, Malgorzata; Silva, Ana Rita; Rosário, Ana Lúcia; Brito, José; Oliveira, Tânia Filipa; Nogly, Przemyslaw; Pessanha, Miguel; Stelter, Meike; Kletzin, Arnulf; Henderson, Peter J. F.; Archer, Margarida

    2013-01-01

    Background Membrane proteins play a key role in many fundamental cellular processes such as transport of nutrients, sensing of environmental signals and energy transduction, and account for over 50% of all known drug targets. Despite their importance, structural and functional characterisation of membrane proteins still remains a challenge, partially due to the difficulties in recombinant expression and purification. Therefore the need for development of efficient methods for heterologous production is essential. Methodology/Principal Findings Fifteen integral membrane transport proteins from Archaea were selected as test targets, chosen to represent two superfamilies widespread in all organisms known as the Major Facilitator Superfamily (MFS) and the 5-Helix Inverted Repeat Transporter superfamily (5HIRT). These proteins typically have eleven to twelve predicted transmembrane helices and are putative transporters for sugar, metabolite, nucleobase, vitamin or neurotransmitter. They include a wide range of examples from the following families: Metabolite-H+-symporter; Sugar Porter; Nucleobase-Cation-Symporter-1; Nucleobase-Cation-Symporter-2; and neurotransmitter-sodium-symporter. Overproduction of transporters was evaluated with three vectors (pTTQ18, pET52b, pWarf) and two Escherichia coli strains (BL21 Star and C43 (DE3)). Thirteen transporter genes were successfully expressed; only two did not express in any of the tested vector-strain combinations. Initial trials showed that seven transporters could be purified and six of these yielded quantities of ≥ 0.4 mg per litre suitable for functional and structural studies. Size-exclusion chromatography confirmed that two purified transporters were almost homogeneous while four others were shown to be non-aggregating, indicating that they are ready for up-scale production and crystallisation trials. Conclusions/Significance Here, we describe an efficient strategy for heterologous production of membrane transport

  7. Unveiling the functional diversity of the alpha/beta hydrolase superfamily in the plant kingdom.

    PubMed

    Mindrebo, Jeffrey T; Nartey, Charisse M; Seto, Yoshiya; Burkart, Michael D; Noel, Joseph P

    2016-12-01

    The alpha/beta hydrolase (ABH) superfamily is a widespread and functionally malleable protein fold recognized for its diverse biochemical activities across all three domains of life. ABH enzymes possess unexpected catalytic activity in the green plant lineage through selective alterations in active site architecture and chemistry. Furthermore, the ABH fold serves as the core structure for phytohormone and ligand receptors in the gibberellin, strigolactone, and karrikin signaling pathways in plants. Despite recent discoveries, the ABH family is sparsely characterized in plants, a sessile kingdom known to evolve complex and specialized chemical adaptations as survival responses to widely varying biotic and abiotic ecologies. This review calls attention to the ABH superfamily in the plant kingdom to highlight the functional adaptability of the ABH fold. Copyright © 2016. Published by Elsevier Ltd.

  8. K-ABC Mental Processing Profiles for Gifted Referrals.

    ERIC Educational Resources Information Center

    Harrison, Patti L.; And Others

    This study sought to extend previous research by investigating performance of intellectucally gifted children on the Mental Processing Composite of the Kaufman Assessment Battery for Children (K-ABC). A sample of 54 children (aged 6-12) referred for possible gifted placement were administered the Sequential and Simultaneous scales. Average scores…

  9. ATP-Binding Cassette Efflux Transporters in Human Placenta

    PubMed Central

    Ni, Zhanglin; Mao, Qingcheng

    2010-01-01

    Pregnant women are often complicated with diseases including viral or bacterial infections, epilepsy, hypertension, or pregnancy-induced conditions such as depression and gestational diabetes that require treatment with medication. In addition, substance abuse during pregnancy remains a major public health problem. Many drugs used by pregnant women are off label without the necessary dose, efficacy, and safety data required for rational dosing regimens of these drugs. Thus, a major concern arising from the widespread use of drugs by pregnant women is the transfer of drugs across the placental barrier, leading to potential toxicity to the developing fetus. Knowledge regarding the ATP-binding cassette (ABC) efflux transporters, which play an important role in drug transfer across the placental barrier, is absolutely critical for optimizing the therapeutic strategy to treat the mother while protecting the fetus during pregnancy. Such transporters include P-glycoprotein (P-gp, gene symbol ABCB1), the breast cancer resistance protein (BCRP, gene symbol ABCG2), and the multidrug resistance proteins (MRPs, gene symbol ABCCs). In this review, we summarize the current knowledge with respect to developmental expression and regulation, membrane localization, functional significance, and genetic polymorphisms of these ABC transporters in the placenta and their relevance to fetal drug exposure and toxicity. PMID:21118087

  10. The First Mitochondrial Genome for the Superfamily Hagloidea and Implications for Its Systematic Status in Ensifera

    PubMed Central

    Zhou, Zhijun; Shi, Fuming; Zhao, Ling

    2014-01-01

    Hagloidea Handlirsch, 1906 was an ancient group of Ensifera, that was much more diverse in the past extending at least into the Triassic, apparently diminishing in diversity through the Cretaceous, and now only represented by a few extant species. In this paper, we report the complete mitochondrial genome (mitogenome) of Tarragoilus diuturnus Gorochov, 2001, representing the first mitogenome of the superfamily Hagloidea. The size of the entire mitogenome of T. diuturnus is 16144 bp, containing 13 protein-coding genes (PCGs), 2 ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes and one control region. The order and orientation of the gene arrangement pattern is identical to that of D. yakuba and most ensiferans species. A phylogenomic analysis was carried out based on the concatenated dataset of 13 PCGs and 2 rRNA genes from mitogenome sequences of 15 ensiferan species, comprising four superfamilies Grylloidea, Tettigonioidae, Rhaphidophoroidea and Hagloidea. Both maximum likelihood and Bayesian inference analyses strongly support Hagloidea T. diuturnus and Rhaphidophoroidea Troglophilus neglectus as forming a monophyletic group, sister to the Tettigonioidea. The relationships among four superfamilies of Ensifera were (Grylloidea, (Tettigonioidea, (Hagloidea, Rhaphidophoroidea))). PMID:24465850

  11. Caenorhabditis elegans ABCRNAi Transporters Interact Genetically With rde-2 and mut-7

    PubMed Central

    Sundaram, Prema; Han, Wang; Cohen, Nancy; Echalier, Benjamin; Albin, John; Timmons, Lisa

    2008-01-01

    RNA interference (RNAi) mechanisms are conserved and consist of an interrelated network of activities that not only respond to exogenous dsRNA, but also perform endogenous functions required in the fine tuning of gene expression and in maintaining genome integrity. Not surprisingly, RNAi functions have widespread influences on cellular function and organismal development. Previously, we observed a reduced capacity to mount an RNAi response in nine Caenorhabditis elegans mutants that are defective in ABC transporter genes (ABCRNAi mutants). Here, we report an exhaustive study of mutants, collectively defective in 49 different ABC transporter genes, that allowed for the categorization of one additional transporter into the ABCRNAi gene class. Genetic complementation tests reveal functions for ABCRNAi transporters in the mut-7/rde-2 branch of the RNAi pathway. These second-site noncomplementation interactions suggest that ABCRNAi proteins and MUT-7/RDE-2 function together in parallel pathways and/or as multiprotein complexes. Like mut-7 and rde-2, some ABCRNAi mutants display transposon silencing defects. Finally, our analyses reveal a genetic interaction network of ABCRNAi gene function with respect to this part of the RNAi pathway. From our results, we speculate that the coordinated activities of ABCRNAi transporters, through their effects on endogenous RNAi-related mechanisms, ultimately affect chromosome function and integrity. PMID:18245353

  12. A Sensory Complex Consisting of an ATP-binding Cassette Transporter and a Two-component Regulatory System Controls Bacitracin Resistance in Bacillus subtilis*

    PubMed Central

    Dintner, Sebastian; Heermann, Ralf; Fang, Chong; Jung, Kirsten; Gebhard, Susanne

    2014-01-01

    Resistance against antimicrobial peptides in many Firmicutes bacteria is mediated by detoxification systems that are composed of a two-component regulatory system (TCS) and an ATP-binding cassette (ABC) transporter. The histidine kinases of these systems depend entirely on the transporter for sensing of antimicrobial peptides, suggesting a novel mode of signal transduction where the transporter constitutes the actual sensor. The aim of this study was to investigate the molecular mechanisms of this unusual signaling pathway in more detail, using the bacitracin resistance system BceRS-BceAB of Bacillus subtilis as an example. To analyze the proposed communication between TCS and the ABC transporter, we characterized their interactions by bacterial two-hybrid analyses and could show that the permease BceB and the histidine kinase BceS interact directly. In vitro pulldown assays confirmed this interaction, which was found to be independent of bacitracin. Because it was unknown whether BceAB-type transporters could detect their substrate peptides directly or instead recognized the peptide-target complex in the cell envelope, we next analyzed substrate binding by the transport permease, BceB. Direct and specific binding of bacitracin by BceB was demonstrated by surface plasmon resonance spectroscopy. Finally, in vitro signal transduction assays indicated that complex formation with the transporter influenced the autophosphorylation activity of the histidine kinase. Taken together, our findings clearly show the existence of a sensory complex composed of TCS and ABC transporters and provide the first functional insights into the mechanisms of stimulus perception, signal transduction, and antimicrobial resistance employed by Bce-like detoxification systems. PMID:25118291

  13. Transepithelial transport of rosuvastatin and effect of ursolic acid on its transport in Caco-2 monolayers.

    PubMed

    Hua, Wen Jin; Fang, Hu Jin; Hua, Wei Xiao

    2012-09-01

    The aim of this study was to determine transepithelial transport characteristics of rosuvastatin and effect of ursolic acid (P-gp potential inhibitor) and ko143 (ABC transporters selective inhibitor) on its transport in Caco-2 monolayers. A reliable Caco-2 cell monolayers model was established. The TEER value was used to inspect integrity of cell model. Apparent permeability coefficients (Papp(BL-AP) and Papp(AP-BL)) were used to analyze transepithelial transport of rosuvastatin. Uptake of rosuvastatin was time- and concentration-dependent in Caco-2 cell. The ko143 but not ursolic acid had effect on the uptake of rosuvastatin in Caco-2 cell monolayer model and affected apparent permeability coefficient and apparent permeability of rosuvastatin. Active transport and passive diffusion absorption existed in transepithelial transport of rosuvastatin in Caco-2 cell model. Ursolic acid had no effect on transport of rosuvastatin in Caco-2 cell monolayer. The result indicated that ursolic acid may not cause effect on intestinal absorption of rosuvastatin.

  14. The Glutathione-S-Transferase, Cytochrome P450 and Carboxyl/Cholinesterase Gene Superfamilies in Predatory Mite Metaseiulus occidentalis

    PubMed Central

    Hoy, Marjorie A.

    2016-01-01

    Pesticide-resistant populations of the predatory mite Metaseiulus (= Typhlodromus or Galendromus) occidentalis (Arthropoda: Chelicerata: Acari: Phytoseiidae) have been used in the biological control of pest mites such as phytophagous Tetranychus urticae. However, the pesticide resistance mechanisms in M. occidentalis remain largely unknown. In other arthropods, members of the glutathione-S-transferase (GST), cytochrome P450 (CYP) and carboxyl/cholinesterase (CCE) gene superfamilies are involved in the diverse biological pathways such as the metabolism of xenobiotics (e.g. pesticides) in addition to hormonal and chemosensory processes. In the current study, we report the identification and initial characterization of 123 genes in the GST, CYP and CCE superfamilies in the recently sequenced M. occidentalis genome. The gene count represents a reduction of 35% compared to T. urticae. The distribution of genes in the GST and CCE superfamilies in M. occidentalis differs significantly from those of insects and resembles that of T. urticae. Specifically, we report the presence of the Mu class GSTs, and the J’ and J” clade CCEs that, within the Arthropoda, appear unique to Acari. Interestingly, the majority of CCEs in the J’ and J” clades contain a catalytic triad, suggesting that they are catalytically active. They likely represent two Acari-specific CCE clades that may participate in detoxification of xenobiotics. The current study of genes in these superfamilies provides preliminary insights into the potential molecular components that may be involved in pesticide metabolism as well as hormonal/chemosensory processes in the agriculturally important M. occidentalis. PMID:27467523

  15. Arnold Diffusion of Charged Particles in ABC Magnetic Fields

    NASA Astrophysics Data System (ADS)

    Luque, Alejandro; Peralta-Salas, Daniel

    2017-06-01

    We prove the existence of diffusing solutions in the motion of a charged particle in the presence of ABC magnetic fields. The equations of motion are modeled by a 3DOF Hamiltonian system depending on two parameters. For small values of these parameters, we obtain a normally hyperbolic invariant manifold and we apply the so-called geometric methods for a priori unstable systems developed by A. Delshams, R. de la Llave and T.M. Seara. We characterize explicitly sufficient conditions for the existence of a transition chain of invariant tori having heteroclinic connections, thus obtaining global instability (Arnold diffusion). We also check the obtained conditions in a computer-assisted proof. ABC magnetic fields are the simplest force-free-type solutions of the magnetohydrodynamics equations with periodic boundary conditions, and can be considered as an elementary model for the motion of plasma-charged particles in a tokamak.

  16. Know a Baby Who Needs Help? Call ABC.

    ERIC Educational Resources Information Center

    Kromer, Megan E.

    The booklet describes Project ABC (Any Baby Can), a model networking effort to promote coordinated services for disabled and high-risk infants in San Antonio, Texas. The model features a volunteer, grass-roots emphasis in an aggressive community awareness campaign with a long-term goal of improving the effectiveness of social services and health…

  17. Effect of drug efflux transporters on placental transport of antiretroviral agent abacavir.

    PubMed

    Neumanova, Zuzana; Cerveny, Lukas; Greenwood, Susan L; Ceckova, Martina; Staud, Frantisek

    2015-11-01

    Abacavir is as a frequent part of combination antiretroviral therapy used in pregnant women. The aim of this study was to investigate, using in vitro, in situ and ex vivo experimental approaches, whether the transplacental pharmacokinetics of abacavir is affected by ATP-binding cassette (ABC) efflux transporters functionally expressed in the placenta: P-glycoprotein (ABCB1), breast cancer resistance protein (ABCG2), multidrug resistance-associated protein 2 (ABCC2) and multidrug resistance-associated protein 5 (ABCC5). In vitro transport assays revealed that abacavir is a substrate of human ABCB1 and ABCG2 transporters but not of ABCC2 or ABCC5. In addition, in situ experiments using dually perfused rat term placenta confirmed interactions of abacavir with placental Abcb1/Abcg2. In contrast, uptake studies in human placental villous fragments did not reveal any interaction of abacavir with efflux transporters suggesting a large contribution of passive diffusion and/or influx mechanisms to net transplacental abacavir transfer. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Structural Validity of the Movement ABC-2 Test: Factor Structure Comparisons across Three Age Groups

    ERIC Educational Resources Information Center

    Schulz, Joerg; Henderson, Sheila E.; Sugden, David A.; Barnett, Anna L.

    2011-01-01

    Background: The Movement ABC test is one of the most widely used assessments in the field of Developmental Coordination Disorder (DCD). Improvements to the 2nd edition of the test (M-ABC-2) include an extension of the age range and reduction in the number of age bands as well as revision of tasks. The total test score provides a measure of motor…

  19. The ABC of non-inferiority margin setting from indirect comparisons.

    PubMed

    Julious, Steven A

    2011-01-01

    In a non-inferiority trial to assess a new investigative treatment, there may need to be consideration of an indirect comparison with placebo using the active control in the current trial. We can, therefore, use the fact that there is a common active control in the comparisons of the investigative treatment and placebo. In analysing a non-inferiority trial, the ABC of: Assay sensitivity, Bias minimisation and Constancy assumption needs to be considered. It is highlighted how the ABC assumptions can potentially fail when there is placebo creep or a patient population shift. In this situation, the belief about the placebo response expressed in terms of a prior probability in Bayesian formulation could be used with the observed treatment effects to set the non-inferiority limit. Copyright © 2011 John Wiley & Sons, Ltd.

  20. Preclinical properties and human in vivo assessment of 123 I-ABC577 as a novel SPECT agent for imaging amyloid-β

    PubMed Central

    Okumura, Yuki; Kobayashi, Ryohei; Onishi, Takako; Shoyama, Yoshinari; Barret, Olivier; Alagille, David; Jennings, Danna; Marek, Kenneth; Seibyl, John; Tamagnan, Gilles; Tanaka, Akihiro; Shirakami, Yoshifumi

    2016-01-01

    Abstract Non-invasive imaging of amyloid-β in the brain, a hallmark of Alzheimer’s disease, may support earlier and more accurate diagnosis of the disease. In this study, we assessed the novel single photon emission computed tomography tracer 123 I-ABC577 as a potential imaging biomarker for amyloid-β in the brain. The radio-iodinated imidazopyridine derivative 123 I-ABC577 was designed as a candidate for a novel amyloid-β imaging agent. The binding affinity of 123 I-ABC577 for amyloid-β was evaluated by saturation binding assay and in vitro autoradiography using post-mortem Alzheimer’s disease brain tissue. Biodistribution experiments using normal rats were performed to evaluate the biokinetics of 123 I-ABC577. Furthermore, to validate 123 I-ABC577 as a biomarker for Alzheimer’s disease, we performed a clinical study to compare the brain uptake of 123 I-ABC577 in three patients with Alzheimer’s disease and three healthy control subjects. 123 I-ABC577 binding was quantified by use of the standardized uptake value ratio, which was calculated for the cortex using the cerebellum as a reference region. Standardized uptake value ratio images were visually scored as positive or negative. As a result, 123 I-ABC577 showed high binding affinity for amyloid-β and desirable pharmacokinetics in the preclinical studies. In the clinical study, 123 I-ABC577 was an effective marker for discriminating patients with Alzheimer’s disease from healthy control subjects based on visual images or the ratio of cortical-to-cerebellar binding. In patients with Alzheimer’s disease, 123 I-ABC577 demonstrated clear retention in cortical regions known to accumulate amyloid, such as the frontal cortex, temporal cortex, and posterior cingulate. In contrast, less, more diffuse, and non-specific uptake without localization to these key regions was observed in healthy controls. At 150 min after injection, the cortical standardized uptake value ratio increased by ∼60% in patients

  1. Mechanistic and Evolutionary Insights from Comparative Enzymology of Phosphomonoesterases and Phosphodiesterases across the Alkaline Phosphatase Superfamily

    PubMed Central

    2016-01-01

    Naively one might have expected an early division between phosphate monoesterases and diesterases of the alkaline phosphatase (AP) superfamily. On the contrary, prior results and our structural and biochemical analyses of phosphate monoesterase PafA, from Chryseobacterium meningosepticum, indicate similarities to a superfamily phosphate diesterase [Xanthomonas citri nucleotide pyrophosphatase/phosphodiesterase (NPP)] and distinct differences from the three metal ion AP superfamily monoesterase, from Escherichia coli AP (EcAP). We carried out a series of experiments to map out and learn from the differences and similarities between these enzymes. First, we asked why there would be independent instances of monoesterases in the AP superfamily? PafA has a much weaker product inhibition and slightly higher activity relative to EcAP, suggesting that different metabolic evolutionary pressures favored distinct active-site architectures. Next, we addressed the preferential phosphate monoester and diester catalysis of PafA and NPP, respectively. We asked whether the >80% sequence differences throughout these scaffolds provide functional specialization for each enzyme’s cognate reaction. In contrast to expectations from this model, PafA and NPP mutants with the common subset of active-site groups embedded in each native scaffold had the same monoesterase:diesterase specificities; thus, the >107-fold difference in native specificities appears to arise from distinct interactions at a single phosphoryl substituent. We also uncovered striking mechanistic similarities between the PafA and EcAP monoesterases, including evidence for ground-state destabilization and functional active-site networks that involve different active-site groups but may play analogous catalytic roles. Discovering common network functions may reveal active-site architectural connections that are critical for function, and identifying regions of functional modularity may facilitate the design of new enzymes

  2. Water-mediated protein-fluorophore interactions modulate the affinity of an ABC-ATPase/TNP-ADP complex.

    PubMed

    Oswald, Christine; Jenewein, Stefan; Smits, Sander H J; Holland, I Barry; Schmitt, Lutz

    2008-04-01

    TNP-modified nucleotides have been used extensively to study protein-nucleotide interactions. In the case of ABC-ATPases, application of these powerful tools has been greatly restricted due to the significantly higher affinity of the TNP-nucleotide for the corresponding ABC-ATPase in comparison to the non-modified nucleotides. To understand the molecular changes occurring upon binding of the TNP-nucleotide to an ABC-ATPase, we have determined the crystal structure of the TNP-ADP/HlyB-NBD complex at 1.6A resolution. Despite the higher affinity of TNP-ADP, no direct fluorophore-protein interactions were observed. Unexpectedly, only water-mediated interactions were detected between the TNP moiety and Tyr(477), that is engaged in pi-pi stacking with the adenine ring, as well as with two serine residues (Ser(504) and Ser(509)) of the Walker A motif. Interestingly, the side chains of these two serine residues adopt novel conformations that are not observed in the corresponding ADP structure. However, in the crystal structure of the S504A mutant, which binds TNP-ADP with similar affinity to the wild type enzyme, a novel TNP-water interaction compensates for the missing serine side chain. Since this water molecule is not present in the wild type enzyme, these results suggest that only water-mediated interactions provide a structural explanation for the increased affinity of TNP-nucleotides towards ABC-ATPases. However, our results also imply that in silico approaches such as docking or modeling cannot directly be applied to generate 'affinity-adopted' ADP- or ATP-analogs for ABC-ATPases.

  3. Selections from the ABC 2011 Annual Convention, Montreal, Canada

    ERIC Educational Resources Information Center

    Whalen, D. Joel; Andersen, Ken; Campbell, Gloria; Crenshaw, Cheri; Cross, Geoffrey A.; Grinols, Anne Bradstreet; Hildebrand, John; Newman, Amy; Ortiz, Lorelei A.; Paulson, Edward; Phillabaum, Melinda; Powell, Elizabeth A.; Sloan, Ryan

    2012-01-01

    The 12 Favorite Assignments featured in this article were presented at the 2011 Annual Convention of the Association for Business Communication (ABC), Montreal, Canada. A variety of learning objectives are featured: delivering bad news, handling difficult people, persuasion, reporting financial analysis, electronic media, face-to-face…

  4. Selections from the ABC 2012 Annual Convention, Honolulu, Hawaii

    ERIC Educational Resources Information Center

    Whalen, D. Joel

    2013-01-01

    The 13 Favorite Assignments featured here were presented at the 2012 Association for Business Communication (ABC) Annual Convention, Honolulu, Hawaii. A variety of learning objectives are featured, including the following: enhancing resume's visual impact, interpersonal skills, social media, team building, web design, community service projects,…

  5. In vitro methanol production from methyl coenzyme M using the Methanosarcina barkeri MtaABC protein complex.

    PubMed

    Dong, Ming; Gonzalez, Tara D; Klems, Meghan M; Steinberg, Lisa M; Chen, Wilfred; Papoutsakis, Eleftherios T; Bahnson, Brian J

    2017-09-01

    Methanol:coenzyme M methyltransferase is an enzyme complex composed of three subunits, MtaA, MtaB, and MtaC, found in methanogenic archaea and is needed for their growth on methanol ultimately producing methane. MtaABC catalyzes the energetically favorable methyl transfer from methanol to coenzyme M to form methyl coenzyme M. Here we demonstrate that this important reaction for possible production of methanol from the anaerobic oxidation of methane can be reversed in vitro. To this effect, we have expressed and purified the Methanosarcina barkeri MtaABC enzyme, and developed an in vitro functional assay that demonstrates MtaABC can catalyze the energetically unfavorable (ΔG° = 27 kJ/mol) reverse reaction starting from methyl coenzyme M and generating methanol as a product. Demonstration of an in vitro ability of MtaABC to produce methanol may ultimately enable the anaerobic oxidation of methane to produce methanol and from methanol alternative fuel or fuel-precursor molecules. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1243-1249, 2017. © 2017 American Institute of Chemical Engineers.

  6. SVM-Fold: a tool for discriminative multi-class protein fold and superfamily recognition

    PubMed Central

    Melvin, Iain; Ie, Eugene; Kuang, Rui; Weston, Jason; Stafford, William Noble; Leslie, Christina

    2007-01-01

    Background Predicting a protein's structural class from its amino acid sequence is a fundamental problem in computational biology. Much recent work has focused on developing new representations for protein sequences, called string kernels, for use with support vector machine (SVM) classifiers. However, while some of these approaches exhibit state-of-the-art performance at the binary protein classification problem, i.e. discriminating between a particular protein class and all other classes, few of these studies have addressed the real problem of multi-class superfamily or fold recognition. Moreover, there are only limited software tools and systems for SVM-based protein classification available to the bioinformatics community. Results We present a new multi-class SVM-based protein fold and superfamily recognition system and web server called SVM-Fold, which can be found at . Our system uses an efficient implementation of a state-of-the-art string kernel for sequence profiles, called the profile kernel, where the underlying feature representation is a histogram of inexact matching k-mer frequencies. We also employ a novel machine learning approach to solve the difficult multi-class problem of classifying a sequence of amino acids into one of many known protein structural classes. Binary one-vs-the-rest SVM classifiers that are trained to recognize individual structural classes yield prediction scores that are not comparable, so that standard "one-vs-all" classification fails to perform well. Moreover, SVMs for classes at different levels of the protein structural hierarchy may make useful predictions, but one-vs-all does not try to combine these multiple predictions. To deal with these problems, our method learns relative weights between one-vs-the-rest classifiers and encodes information about the protein structural hierarchy for multi-class prediction. In large-scale benchmark results based on the SCOP database, our code weighting approach significantly improves

  7. [German adaptation of the "Activities-Specific Balance Confidence (ABC) scale" for the assessment of falls-related self-efficacy].

    PubMed

    Schott, Nadja

    2008-12-01

    The present study investigates the psychometric properties and the factorial structure of the German adaptation of the Activities-Specific Balance Confidence (ABC) scale (Powell & Myers, 1995) for the evaluation of falls-related self-efficacy in community-dwelling older adults. The German adaptation of the ABC using a forward-backward procedure was administered to 113 older adults (age 68.9+/-8.5 years). The following internationally accepted instruments were used for validation: The Short Form Health Survey SF 36, the Geriatric Depression Scale (GDS), the Trail Making Test and the Letter Number Sequencing Test, and motor tests (balance, strength, mobility). The internal consistency (0.91-0.95) as well as the test-retest reliability of the subscales was excellent (0.94-0.98). The correlation coefficients with the validation instruments ranged between 0.33 and 0.58. Significant differences in the ABC-D scores were found in older adults with and without falls. Older adults with a recent fall history scored lower on the ABC-D than older adults without a recent fall history. To conclude, the German version of the ABC has properties analogous to the original English version and is apparently useful in assessing falls-related self-efficacy.

  8. Specific stabilization of CFTR by phosphatidylserine.

    PubMed

    Hildebrandt, Ellen; Khazanov, Netaly; Kappes, John C; Dai, Qun; Senderowitz, Hanoch; Urbatsch, Ina L

    2017-02-01

    The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR, ABCC7) is a plasma membrane chloride ion channel in the ABC transporter superfamily. CFTR is a key target for cystic fibrosis drug development, and its structural elucidation would advance those efforts. However, the limited in vivo and in vitro stability of the protein, particularly its nucleotide binding domains, has made structural studies challenging. Here we demonstrate that phosphatidylserine uniquely stimulates and thermally stabilizes the ATP hydrolysis function of purified human CFTR. Among several lipids tested, the greatest stabilization was observed with brain phosphatidylserine, which shifted the Tm for ATPase activity from 22.7±0.8°C to 35.0±0.2°C in wild-type CFTR, and from 26.6±0.7°C to 42.1±0.2°C in a more stable mutant CFTR having deleted regulatory insertion and S492P/A534P/I539T mutations. When ATPase activity was measured at 37°C in the presence of brain phosphatidylserine, Vmax for wild-type CFTR was 240±60nmol/min/mg, a rate higher than previously reported and consistent with rates for other purified ABC transporters. The significant thermal stabilization of CFTR by phosphatidylserine may be advantageous in future structural and biophysical studies of CFTR. Copyright © 2016. Published by Elsevier B.V.

  9. Glaucarubinone sensitizes KB cells to paclitaxel by inhibiting ABC transporters via ROS-dependent and p53-mediated activation of apoptotic signaling pathways

    PubMed Central

    Karthikeyan, Subburayan; Hoti, Sugeerappa Laxmanappa; Nazeer, Yasin; Hegde, Harsha Vasudev

    2016-01-01

    Multidrug resistance (MDR) is considered to be the major contributor to failure of chemotherapy in oral squamous cell carcinoma (SCC). This study was aimed to explore the effects and mechanisms of glaucarubinone (GLU), one of the major quassinoids from Simarouba glauca DC, in potentiating cytotoxicity of paclitaxel (PTX), an anticancer drug in KB cells. Our data showed that the administration of GLU pre-treatment significantly enhanced PTX anti-proliferative effect in ABCB1 over-expressing KB cells. The Rh 123 drug efflux studies revealed that there was a significant transport function inhibition by GLU-PTX treatment. Interestingly, it was also found that this enhanced anticancer efficacy of GLU was associated with PTX-induced cell arrest in the G2/M phase of cell cycle. Further, the combined treatment of GLU-PTX had significant decrease in the expression levels of P-gp, MRPs, and BCRP in resistant KB cells at both mRNA and protein levels. Furthermore, the combination treatments showed significant reactive oxygen species (ROS) production, chromatin condensation and reduced mitochondrial membrane potential in resistant KB cells. The results from DNA fragmentation analysis also demonstrated the GLU induced apoptosis in KB cells and its synergy with PTX. Importantly, GLU and/or PTX triggered apoptosis through the activation of pro-apoptotic proteins such as p53, Bax, and caspase-9. Our findings demonstrated for the first time that GLU causes cell death in human oral cancer cells via the ROS-dependent suppression of MDR transporters and p53-mediated activation of the intrinsic mitochondrial pathway of apoptosis. Additionally, the present study also focussed on investigation of the protective effect of GLU and combination drugs in human normal blood lymphocytes. Normal blood lymphocytes assay indicated that GLU is able to induce selective toxicity in cancer cells and in silico molecular docking studies support the choice of GLU as ABC inhibitor to enhance PTX efficacy

  10. Comparing the results of DAADD and ABC of children included in autism spectrum disorders.

    PubMed

    Barbosa, Milene Rossi Pereira; Fernandes, Fernanda Dreux Miranda

    2014-01-01

    To verify if there are characteristic behaviors of the different diagnosis included in the autism spectrum according to the Differential Assessment of Autism and Other Developmental Disorders (DAADD) and to the Autism Behavior Checklist (ABC). Participants were 45 individuals and their respective speech-language therapists. All therapists are graduate students working with the children for at least 1 year. This time was considered sufficient to the therapists to have the information required by the DAADD questionnaire. It is comprised by 3 protocols specifically designed to children with 2 to 4 years, 4 to 6 years and 6 to 8 years, the same criteria used to separate the research groups, G1, G2 and G3, respectively. Data referring to the ABC were retrieved from the subject's files at the Laboratório de Investigação Fonoaudiológica nos Distúrbios do Espectro do Autismo (Research Laboratory on Language Disorders in the Autism Spectrum) of the School of Medicine, Universidade de São Paulo, where it is routinely applied during the annual assessment. Answers to the different areas of DAADD are similar to the different areas of ABC. These data show data the diagnosis by DAADD is easier in older children. Although there is no significant difference, the large occurrence of Rett's syndrome diagnosis according to the DAADD was associated to higher risk for autism according to the ABC in G1. With increasing age this tendency decreases and either in G2 and G3 Autism is the most frequent diagnosis. Although the results of both questionnaires tend to agree more with increasing age, the DAADD is more sensitive in the different ages while the ABC if more specific only to older children.

  11. ATP-binding cassette transporters and sterol O-acyltransferases interact at membrane microdomains to modulate sterol uptake and esterification

    PubMed Central

    Gulati, Sonia; Balderes, Dina; Kim, Christine; Guo, Zhongmin A.; Wilcox, Lisa; Area-Gomez, Estela; Snider, Jamie; Wolinski, Heimo; Stagljar, Igor; Granato, Juliana T.; Ruggles, Kelly V.; DeGiorgis, Joseph A.; Kohlwein, Sepp D.; Schon, Eric A.; Sturley, Stephen L.

    2015-01-01

    A key component of eukaryotic lipid homeostasis is the esterification of sterols with fatty acids by sterol O-acyltransferases (SOATs). The esterification reactions are allosterically activated by their sterol substrates, the majority of which accumulate at the plasma membrane. We demonstrate that in yeast, sterol transport from the plasma membrane to the site of esterification is associated with the physical interaction of the major SOAT, acyl-coenzyme A:cholesterol acyltransferase (ACAT)-related enzyme (Are)2p, with 2 plasma membrane ATP-binding cassette (ABC) transporters: Aus1p and Pdr11p. Are2p, Aus1p, and Pdr11p, unlike the minor acyltransferase, Are1p, colocalize to sterol and sphingolipid-enriched, detergent-resistant microdomains (DRMs). Deletion of either ABC transporter results in Are2p relocalization to detergent-soluble membrane domains and a significant decrease (53–36%) in esterification of exogenous sterol. Similarly, in murine tissues, the SOAT1/Acat1 enzyme and activity localize to DRMs. This subcellular localization is diminished upon deletion of murine ABC transporters, such as Abcg1, which itself is DRM associated. We propose that the close proximity of sterol esterification and transport proteins to each other combined with their residence in lipid-enriched membrane microdomains facilitates rapid, high-capacity sterol transport and esterification, obviating any requirement for soluble intermediary proteins.—Gulati, S., Balderes, D., Kim, C., Guo, Z. A., Wilcox, L., Area-Gomez, E., Snider, J., Wolinski, H., Stagljar, I., Granato, J. T., Ruggles, K. V., DeGiorgis, J. A., Kohlwein, S. D., Schon, E. A., Sturley, S. L. ATP-binding cassette transporters and sterol O-acyltransferases interact at membrane microdomains to modulate sterol uptake and esterification. PMID:26220175

  12. A comprehensive analysis of the Omp85/TpsB protein superfamily structural diversity, taxonomic occurrence, and evolution

    PubMed Central

    Heinz, Eva; Lithgow, Trevor

    2014-01-01

    Members of the Omp85/TpsB protein superfamily are ubiquitously distributed in Gram-negative bacteria, and function in protein translocation (e.g., FhaC) or the assembly of outer membrane proteins (e.g., BamA). Several recent findings are suggestive of a further level of variation in the superfamily, including the identification of the novel membrane protein assembly factor TamA and protein translocase PlpD. To investigate the diversity and the causal evolutionary events, we undertook a comprehensive comparative sequence analysis of the Omp85/TpsB proteins. A total of 10 protein subfamilies were apparent, distinguished in their domain structure and sequence signatures. In addition to the proteins FhaC, BamA, and TamA, for which structural and functional information is available, are families of proteins with so far undescribed domain architectures linked to the Omp85 β-barrel domain. This study brings a classification structure to a dynamic protein superfamily of high interest given its essential function for Gram-negative bacteria as well as its diverse domain architecture, and we discuss several scenarios of putative functions of these so far undescribed proteins. PMID:25101071

  13. Drosophila ABC Transporter DmHMT-1 Confers Tolerance to Cadmium.

    USDA-ARS?s Scientific Manuscript database

    Half molecule ATP-binding cassette transporters of the HMT1(heavy metal tolerance factor 1)subfamily are required for Cd2+ tolerance in Schizosaccharomyces pombe, Caenorhabditis elegans and Chlamydomonas reinhardtii, and have homologs in other species, including plants and humans. Based on studies i...

  14. Discovery and characterization of two Nimrod superfamily members in Anopheles gambiae.

    PubMed

    Midega, Janet; Blight, Joshua; Lombardo, Fabrizio; Povelones, Michael; Kafatos, Fotis; Christophides, George K

    2013-12-01

    Anti-bacterial proteins in mosquitoes are known to play an important modulatory role on immune responses to infections with human pathogens including malaria parasites. In this study we characterized two members of the Anopheles gambiae Nimrod superfamily, namely AgNimB2 and AgEater. We confirm that current annotation of the An. gambiae genome incorrectly identifies AgNimB2 and AgEater as a single gene, AGAP009762. Through in silico and experimental approaches, it has been shown that AgNimB2 is a secreted protein that mediates phagocytosis of Staphylococcus aureus but not of Escherichia coli bacteria. We also reveal that this function does not involve a direct interaction of AgNimB2 with S. aureus. Therefore, AgNimB2 may act downstream of complement-like pathway activation, first requiring bacterial opsonization. In addition, it has been shown that AgNimB2 has an anti-Plasmodium effect. Conversely, AgEater is a membrane-bound protein that either functions redundantly or is dispensable for phagocytosis of E. coli or S. aureus. Our study provides insights into the role of members of the complex Nimrod superfamily in An. gambiae, the most important African vector of human malaria.

  15. Discovery and characterization of two Nimrod superfamily members in Anopheles gambiae

    PubMed Central

    Midega, Janet; Blight, Joshua; Lombardo, Fabrizio; Povelones, Michael; Kafatos, Fotis; Christophides, George K

    2013-01-01

    Anti-bacterial proteins in mosquitoes are known to play an important modulatory role on immune responses to infections with human pathogens including malaria parasites. In this study we characterized two members of the Anopheles gambiae Nimrod superfamily, namely AgNimB2 and AgEater. We confirm that current annotation of the An. gambiae genome incorrectly identifies AgNimB2 and AgEater as a single gene, AGAP009762. Through in silico and experimental approaches, it has been shown that AgNimB2 is a secreted protein that mediates phagocytosis of Staphylococcus aureus but not of Escherichia coli bacteria. We also reveal that this function does not involve a direct interaction of AgNimB2 with S. aureus. Therefore, AgNimB2 may act downstream of complement-like pathway activation, first requiring bacterial opsonization. In addition, it has been shown that AgNimB2 has an anti-Plasmodium effect. Conversely, AgEater is a membrane-bound protein that either functions redundantly or is dispensable for phagocytosis of E. coli or S. aureus. Our study provides insights into the role of members of the complex Nimrod superfamily in An. gambiae, the most important African vector of human malaria. PMID:24428830

  16. Selections from the ABC 2009 Annual Convention, Portsmouth, Virginia

    ERIC Educational Resources Information Center

    Whalen, D. Joel

    2010-01-01

    The "My Favorite Assignment" Session at the 2009 Association for Business Communication (ABC) annual convention in Portsmouth, Virginia, featured over a dozen teachers sharing pedagogical innovations in a fast-paced, 4-minute format designed by Dan Dietrich. The wide variety of ideas and techniques presented makes these sessions popular…

  17. Strategies for neurotrophin-3 and chondroitinase ABC release from freeze-cast chitosan-alginate nerve-guidance scaffolds.

    PubMed

    Francis, Nicola L; Hunger, Philipp M; Donius, Amalie E; Wegst, Ulrike G K; Wheatley, Margaret A

    2017-01-01

    Freeze casting, or controlled unidirectional solidification, can be used to fabricate chitosan-alginate (C-A) scaffolds with highly aligned porosity that are suitable for use as nerve-guidance channels. To augment the guidance of growth across a spinal cord injury lesion, these scaffolds are now evaluated in vitro to assess their ability to release neurotrophin-3 (NT-3) and chondroitinase ABC (chABC) in a controlled manner. Protein-loaded microcapsules were incorporated into C-A scaffolds prior to freeze casting without affecting the original scaffold architecture. In vitro protein release was not significantly different when comparing protein loaded directly into the scaffolds with release from scaffolds containing incorporated microcapsules. NT-3 was released from the C-A scaffolds for 8 weeks in vitro, while chABC was released for up to 7 weeks. Low total percentages of protein released from the scaffolds over this time period were attributed to limitation of diffusion by the interpenetrating polymer network matrix of the scaffold walls. NT-3 and chABC released from the scaffolds retained bioactivity, as determined by a neurite outgrowth assay, and the promotion of neurite growth across an inhibitory barrier of chondroitin sulphate proteoglycans. This demonstrates the potential of these multifunctional scaffolds for enhancing axonal regeneration through growth-inhibiting glial scars via the sustained release of chABC and NT-3. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

  18. The Components of the Unique Zur Regulon of Cupriavidus metallidurans Mediate Cytoplasmic Zinc Handling

    PubMed Central

    Bütof, Lucy; Schmidt-Vogler, Christopher; Herzberg, Martin; Große, Cornelia

    2017-01-01

    ABSTRACT Zinc is an essential trace element, yet it is toxic at high concentrations. In the betaproteobacterium Cupriavidus metallidurans, the highly efficient removal of surplus zinc from the periplasm is responsible for the outstanding metal resistance of the organism. Rather than having a typical Zur-dependent, high-affinity ATP-binding cassette transporter of the ABC protein superfamily for zinc uptake at low concentrations, C. metallidurans has the secondary zinc importer ZupT of the zinc-regulated transporter, iron-regulated transporter (ZRT/IRT)-like protein (ZIP) family. It is important to understand, therefore, how this zinc-resistant bacterium copes with exposure to low zinc concentrations. Members of the Zur regulon in C. metallidurans were identified by comparing the transcriptomes of a Δzur mutant and its parent strain. The consensus sequence of the Zur-binding box was derived for the zupTp promoter-regulatory region by use of a truncation assay. The motif was used to predict possible Zur boxes upstream of Zur regulon members. The binding of Zur to these boxes was confirmed. Two Zur boxes upstream of the cobW1 gene, encoding a putative zinc chaperone, proved to be required for complete repression of cobW1 and its downstream genes in cells cultivated in mineral salts medium. A Zur box upstream of each of zur-cobW2, cobW3, and zupT permitted both low expression levels of these genes and their upregulation under conditions of zinc starvation. This demonstrates a compartmentalization of zinc homeostasis in C. metallidurans, where the periplasm is responsible for the removal of surplus zinc, cytoplasmic components are responsible for the management of zinc as an essential cofactor, and the two compartments are connected by ZupT. IMPORTANCE Elucidating zinc homeostasis is necessary for understanding both host-pathogen interactions and the performance of free-living bacteria in their natural environments. Escherichia coli acquires zinc under conditions of

  19. A sensory complex consisting of an ATP-binding cassette transporter and a two-component regulatory system controls bacitracin resistance in Bacillus subtilis.

    PubMed

    Dintner, Sebastian; Heermann, Ralf; Fang, Chong; Jung, Kirsten; Gebhard, Susanne

    2014-10-03

    Resistance against antimicrobial peptides in many Firmicutes bacteria is mediated by detoxification systems that are composed of a two-component regulatory system (TCS) and an ATP-binding cassette (ABC) transporter. The histidine kinases of these systems depend entirely on the transporter for sensing of antimicrobial peptides, suggesting a novel mode of signal transduction where the transporter constitutes the actual sensor. The aim of this study was to investigate the molecular mechanisms of this unusual signaling pathway in more detail, using the bacitracin resistance system BceRS-BceAB of Bacillus subtilis as an example. To analyze the proposed communication between TCS and the ABC transporter, we characterized their interactions by bacterial two-hybrid analyses and could show that the permease BceB and the histidine kinase BceS interact directly. In vitro pulldown assays confirmed this interaction, which was found to be independent of bacitracin. Because it was unknown whether BceAB-type transporters could detect their substrate peptides directly or instead recognized the peptide-target complex in the cell envelope, we next analyzed substrate binding by the transport permease, BceB. Direct and specific binding of bacitracin by BceB was demonstrated by surface plasmon resonance spectroscopy. Finally, in vitro signal transduction assays indicated that complex formation with the transporter influenced the autophosphorylation activity of the histidine kinase. Taken together, our findings clearly show the existence of a sensory complex composed of TCS and ABC transporters and provide the first functional insights into the mechanisms of stimulus perception, signal transduction, and antimicrobial resistance employed by Bce-like detoxification systems. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. The jABC Approach to Rigorous Collaborative Development of SCM Applications

    NASA Astrophysics Data System (ADS)

    Hörmann, Martina; Margaria, Tiziana; Mender, Thomas; Nagel, Ralf; Steffen, Bernhard; Trinh, Hong

    Our approach to the model-driven collaborative design of IKEA's P3 Delivery Management Process uses the jABC [9] for model driven mediation and choreography to complement a RUP-based (Rational Unified Process) development process. jABC is a framework for service development based on Lightweight Process Coordination. Users (product developers and system/software designers) easily develop services and applications by composing reusable building-blocks into (flow-) graph structures that can be animated, analyzed, simulated, verified, executed, and compiled. This way of handling the collaborative design of complex embedded systems has proven to be effective and adequate for the cooperation of non-programmers and non-technical people, which is the focus of this contribution, and it is now being rolled out in the operative practice.

  1. HMMER Cut-off Threshold Tool (HMMERCTTER): Supervised classification of superfamily protein sequences with a reliable cut-off threshold.

    PubMed

    Pagnuco, Inti Anabela; Revuelta, María Victoria; Bondino, Hernán Gabriel; Brun, Marcel; Ten Have, Arjen

    2018-01-01

    Protein superfamilies can be divided into subfamilies of proteins with different functional characteristics. Their sequences can be classified hierarchically, which is part of sequence function assignation. Typically, there are no clear subfamily hallmarks that would allow pattern-based function assignation by which this task is mostly achieved based on the similarity principle. This is hampered by the lack of a score cut-off that is both sensitive and specific. HMMER Cut-off Threshold Tool (HMMERCTTER) adds a reliable cut-off threshold to the popular HMMER. Using a high quality superfamily phylogeny, it clusters a set of training sequences such that the cluster-specific HMMER profiles show cluster or subfamily member detection with 100% precision and recall (P&R), thereby generating a specific threshold as inclusion cut-off. Profiles and thresholds are then used as classifiers to screen a target dataset. Iterative inclusion of novel sequences to groups and the corresponding HMMER profiles results in high sensitivity while specificity is maintained by imposing 100% P&R self detection. In three presented case studies of protein superfamilies, classification of large datasets with 100% precision was achieved with over 95% recall. Limits and caveats are presented and explained. HMMERCTTER is a promising protein superfamily sequence classifier provided high quality training datasets are used. It provides a decision support system that aids in the difficult task of sequence function assignation in the twilight zone of sequence similarity. All relevant data and source codes are available from the Github repository at the following URL: https://github.com/BBCMdP/HMMERCTTER.

  2. HMMER Cut-off Threshold Tool (HMMERCTTER): Supervised classification of superfamily protein sequences with a reliable cut-off threshold

    PubMed Central

    Pagnuco, Inti Anabela; Revuelta, María Victoria; Bondino, Hernán Gabriel; Brun, Marcel

    2018-01-01

    Background Protein superfamilies can be divided into subfamilies of proteins with different functional characteristics. Their sequences can be classified hierarchically, which is part of sequence function assignation. Typically, there are no clear subfamily hallmarks that would allow pattern-based function assignation by which this task is mostly achieved based on the similarity principle. This is hampered by the lack of a score cut-off that is both sensitive and specific. Results HMMER Cut-off Threshold Tool (HMMERCTTER) adds a reliable cut-off threshold to the popular HMMER. Using a high quality superfamily phylogeny, it clusters a set of training sequences such that the cluster-specific HMMER profiles show cluster or subfamily member detection with 100% precision and recall (P&R), thereby generating a specific threshold as inclusion cut-off. Profiles and thresholds are then used as classifiers to screen a target dataset. Iterative inclusion of novel sequences to groups and the corresponding HMMER profiles results in high sensitivity while specificity is maintained by imposing 100% P&R self detection. In three presented case studies of protein superfamilies, classification of large datasets with 100% precision was achieved with over 95% recall. Limits and caveats are presented and explained. Conclusions HMMERCTTER is a promising protein superfamily sequence classifier provided high quality training datasets are used. It provides a decision support system that aids in the difficult task of sequence function assignation in the twilight zone of sequence similarity. All relevant data and source codes are available from the Github repository at the following URL: https://github.com/BBCMdP/HMMERCTTER. PMID:29579071

  3. A new fluorescent dye accumulation assay for parallel measurements of the ABCG2, ABCB1 and ABCC1 multidrug transporter functions.

    PubMed

    Szabó, Edit; Türk, Dóra; Telbisz, Ágnes; Kucsma, Nóra; Horváth, Tamás; Szakács, Gergely; Homolya, László; Sarkadi, Balázs; Várady, György

    2018-01-01

    ABC multidrug transporters are key players in cancer multidrug resistance and in general xenobiotic elimination, thus their functional assays provide important tools for research and diagnostic applications. In this study we have examined the potential interactions of three key human ABC multidrug transporters with PhenGreen diacetate (PGD), a cell permeable fluorescent metal ion indicator. The non-fluorescent, hydrophobic PGD rapidly enters the cells and, after cleavage by cellular esterases, in the absence of quenching metal ions, PhenGreen (PG) becomes highly fluorescent. We found that in cells expressing functional ABCG2, ABCB1, or ABCC1 transporters, cellular PG fluorescence is strongly reduced. This fluorescence signal in the presence of specific transporter inhibitors is increased to the fluorescence levels in the control cells. Thus the PG accumulation assay is a new, unique tool for the parallel determination of the function of the ABCG2, ABCB1, and ABCC1 multidrug transporters. Since PG has very low cellular toxicity, the PG accumulation assay also allows the selection, separation and culturing of selected cell populations expressing either of these transporters.

  4. LhnR and upstream operon LhnABC in Agrobacterium vitis regulate the induction of tobacco hypersensitive responses, grape necrosis and swarming motility.

    PubMed

    Zheng, Desen; Hao, Guixia; Cursino, Luciana; Zhang, Hongsheng; Burr, Thomas J

    2012-09-01

    The characterization of Tn5 transposon insertional mutants of Agrobacterium vitis strain F2/5 revealed a gene encoding a predicted LysR-type transcriptional regulator, lhnR (for 'LysR-type regulator associated with HR and necrosis'), and an immediate upstream operon consisting of three open reading frames (lhnABC) required for swarming motility, surfactant production and the induction of a hypersensitive response (HR) on tobacco and necrosis on grape. The operon lhnABC is unique to A. vitis among the sequenced members in Rhizobiaceae. Mutagenesis of lhnR and lhnABC by gene disruption and complementation of ΔlhnR and ΔlhnABC confirmed their roles in the expression of these phenotypes. Mutation of lhnR resulted in complete loss of HR, swarming motility, surfactant production and reduced necrosis, whereas mutation of lhnABC resulted in loss of swarming motility, delayed and reduced HR development and reduced surfactant production and necrosis. The data from promoter-green fluorescent protein (gfp) fusions showed that lhnR suppresses the expression of lhnABC and negatively autoregulates its own expression. It was also shown that lhnABC negatively affects its own expression and positively affects the transcription of lhnR. lhnR and lhnABC constitute a regulatory circuit that coordinates the transcription level of lhnR, resulting in the expression of swarming, surfactant, HR and necrosis phenotypes. © 2012 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

  5. Transport capabilities of environmental Pseudomonads for sulfur compounds

    DOE PAGES

    Zerbs, Sarah; Korajczyk, Peter J.; Noirot, Philippe H.; ...

    2017-01-27

    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 ligandmore » 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. As a result, 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.« less

  6. The ABCs for Pre-Service Teacher Cultural Competency Development

    ERIC Educational Resources Information Center

    He, Ye; Cooper, Jewell E.

    2009-01-01

    In an effort to combine pre-service teachers' self-reflection with their field experiences to enhance their cultural competency, this study adopted Schmidt's ABC's (Autobiography, Biography, and Cross-cultural Comparison) Model in two courses in a pre-service teacher education program. Through group comparisons, this study measured the impact that…

  7. The Value of Green Technology at ABC Community College

    ERIC Educational Resources Information Center

    McAllister, Bernadette

    2012-01-01

    A challenge facing community colleges nationwide is to reduce the carbon footprint of campuses by initiating green technology initiatives. This case study assessed the effect of switching from paper assignments to a learning management system at ABC Community College. The topic is important because federal and state funding, as well as…

  8. Increasing the cytotoxicity of doxorubicin in breast cancer MCF-7 cells with multidrug resistance using a mesoporous silica nanoparticle drug delivery system.

    PubMed

    Wang, Xin; Teng, Zhaogang; Wang, Haiyan; Wang, Chunyan; Liu, Ying; Tang, Yuxia; Wu, Jiang; Sun, Jin; Wang, Hai; Wang, Jiandong; Lu, Guangming

    2014-01-01

    Resistance to cytotoxic chemotherapy is the main cause of therapeutic failure and death in women with breast cancer. Overexpression of various members of the superfamily of adenosine triphosphate binding cassette (ABC)-transporters has been shown to be associated with multidrug resistance (MDR) phenotype in breast cancer cells. MDR1 protein promotes the intracellular efflux of drugs. A novel approach to address cancer drug resistance is to take advantage of the ability of nanocarriers to sidestep drug resistance mechanisms by endosomal delivery of chemotherapeutic agents. Doxorubicin (DOX) is an anthracycline antibiotic commonly used in breast cancer chemotherapy and a substrate for ABC-mediated drug efflux. In the present study, we developed breast cancer MCF-7 cells with overexpression of MDR1 and designed mesoporous silica nanoparticles (MSNs) which were used as a drug delivery system. We tested the efficacy of DOX in the breast cancer cell line MCF-7/MDR1 and in a MCF-7/MDR1 xenograft nude mouse model using the MSNs drug delivery system. Our data show that drug resistance in the human breast cancer cell line MCF-7/MDR1 can be overcome by treatment with DOX encapsulated within mesoporous silica nanoparticles.

  9. Structures of bacterial homologues of SWEET transporters in two distinct conformations.

    PubMed

    Xu, Yan; Tao, Yuyong; Cheung, Lily S; Fan, Chao; Chen, Li-Qing; Xu, Sophia; Perry, Kay; Frommer, Wolf B; Feng, Liang

    2014-11-20

    SWEETs and their prokaryotic homologues are monosaccharide and disaccharide transporters that are present from Archaea to plants and humans. SWEETs play crucial roles in cellular sugar efflux processes: that is, in phloem loading, pollen nutrition and nectar secretion. Their bacterial homologues, which are called SemiSWEETs, are among the smallest known transporters. Here we show that SemiSWEET molecules, which consist of a triple-helix bundle, form symmetrical, parallel dimers, thereby generating the translocation pathway. Two SemiSWEET isoforms were crystallized, one in an apparently open state and one in an occluded state, indicating that SemiSWEETs and SWEETs are transporters that undergo rocking-type movements during the transport cycle. The topology of the triple-helix bundle is similar yet distinct to that of the basic building block of animal and plant major facilitator superfamily (MFS) transporters (for example, GLUTs and SUTs). This finding indicates two possibilities: that SWEETs and MFS transporters evolved from an ancestral triple-helix bundle or that the triple-helix bundle represents convergent evolution. In SemiSWEETs and SWEETs, two triple-helix bundles are arranged in a parallel configuration to produce the 6- and 6 + 1-transmembrane-helix pores, respectively. In the 12-transmembrane-helix MFS transporters, four triple-helix bundles are arranged into an alternating antiparallel configuration, resulting in a much larger 2 × 2 triple-helix bundle forming the pore. Given the similarity of SemiSWEETs and SWEETs to PQ-loop amino acid transporters and to mitochondrial pyruvate carriers (MPCs), the structures characterized here may also be relevant to other transporters in the MtN3 clan. The insight gained from the structures of these transporters and from the analysis of mutations of conserved residues will improve the understanding of the transport mechanism, as well as allow comparative studies of the different superfamilies involved in sugar

  10. IMG-ABC: An Atlas of Biosynthetic Gene Clusters to Fuel the Discovery of Novel Secondary Metabolites

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chen, I-Min; Chu, Ken; Ratner, Anna

    2014-10-28

    In the discovery of secondary metabolites (SMs), large-scale analysis of sequence data is a promising exploration path that remains largely underutilized due to the lack of relevant computational resources. We present IMG-ABC (https://img.jgi.doe.gov/abc/) -- An Atlas of Biosynthetic gene Clusters within the Integrated Microbial Genomes (IMG) system1. IMG-ABC is a rich repository of both validated and predicted biosynthetic clusters (BCs) in cultured isolates, single-cells and metagenomes linked with the SM chemicals they produce and enhanced with focused analysis tools within IMG. The underlying scalable framework enables traversal of phylogenetic dark matter and chemical structure space -- serving as a doorwaymore » to a new era in the discovery of novel molecules.« less

  11. Antitumor effect of the novel sphingosine kinase 2 inhibitor ABC294640 is enhanced by inhibition of autophagy and by sorafenib in human cholangiocarcinoma cells.

    PubMed

    Ding, Xiwei; Chaiteerakij, Roongruedee; Moser, Catherine D; Shaleh, Hassan; Boakye, Jeffrey; Chen, Gang; Ndzengue, Albert; Li, Ying; Zhou, Yanling; Huang, Shengbing; Sinicrope, Frank A; Zou, Xiaoping; Thomas, Melanie B; Smith, Charles D; Roberts, Lewis R

    2016-04-12

    Sphingosine kinase 2 (Sphk2) has an oncogenic role in cancer. A recently developed first-in-class Sphk2 specific inhibitor ABC294640 displays antitumor activity in many cancer models. However, the role of Sphk2 and the antitumor activity of its inhibitor ABC294640 are not known in cholangiocarcinoma. We investigated the potential of targeting Sphk2 for the treatment of cholangiocarcinoma. We found that Sphk2 is overexpressed in five established human cholangiocarcinoma cell lines (WITT, HuCCT1, EGI-1, OZ and HuH28) and a new patient-derived cholangiocarcinoma cell line (LIV27) compared to H69 normal cholangiocytes. Inhibition of Sphk2 by ABC294640 inhibited proliferation and induced caspase-dependent apoptosis. Furthermore, we found that ABC294640 inhibited STAT3 phosphorylation, one of the key signaling pathways regulating cholangiocarcinoma cell proliferation and survival. ABC294640 also induced autophagy. Inhibition of autophagy by bafilomycin A1 or chloroquine potentiated ABC294640-induced cytotoxicity and apoptosis. In addition, ABC294640 in combination with sorafenib synergistically inhibited cell proliferation of cholangiocarcinoma cells. Strong decreases in STAT3 phosphorylation were observed in WITT and HuCCT1 cells exposed to the ABC294640 and sorafenib combination. These findings provide novel evidence that Sphk2 may be a rational therapeutic target in cholangiocarcinoma. Combinations of ABC294640 with sorafenib and/or autophagy inhibitors may provide novel strategies for the treatment of cholangiocarcinoma.

  12. Evolution of Enzymatic Activities in the Enolase Superfamily: L-Rhamnonate Dehydratase

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Rakus,J.; Fedorov, A.; Fedorov, E.

    2008-01-01

    The l-rhamnonate dehydratase (RhamD) function was assigned to a previously uncharacterized family in the mechanistically diverse enolase superfamily that is encoded by the genome of Escherichia coli K-12. We screened a library of acid sugars to discover that the enzyme displays a promiscuous substrate specificity: l-rhamnonate (6-deoxy-l-mannonate) has the 'best' kinetic constants, with l-mannonate, l-lyxonate, and d-gulonate dehydrated less efficiently. Crystal structures of the RhamDs from both E. coli K-12 and Salmonella typhimurium LT2 (95% sequence identity) were obtained in the presence of Mg2+; the structure of the RhamD from S. typhimurium was also obtained in the presence of 3-deoxy-l-rhamnonatemore » (obtained by reduction of the product with NaBH4). Like other members of the enolase superfamily, RhamD contains an N-terminal a + {beta} capping domain and a C-terminal ({beta}/a)7{beta}-barrel (modified TIM-barrel) catalytic domain with the active site located at the interface between the two domains. In contrast to other members, the specificity-determining '20s loop' in the capping domain is extended in length and the '50s loop' is truncated. The ligands for the Mg2+ are Asp 226, Glu 252 and Glu 280 located at the ends of the third, fourth and fifth {beta}-strands, respectively. The active site of RhamD contains a His 329-Asp 302 dyad at the ends of the seventh and sixth {beta}-strands, respectively, with His 329 positioned to function as the general base responsible for abstraction of the C2 proton of l-rhamnonate to form a Mg2+-stabilized enediolate intermediate. However, the active site does not contain other acid/base catalysts that have been implicated in the reactions catalyzed by other members of the MR subgroup of the enolase superfamily. Based on the structure of the liganded complex, His 329 also is expected to function as the general acid that both facilitates departure of the 3-OH group in a syn-dehydration reaction and delivers a proton to

  13. Starting with ABC and Finishing with XYZ: What Financial Reporting Model Best Fits a Faculty and Why?

    ERIC Educational Resources Information Center

    Berry, Prudence Jane

    2014-01-01

    This article looks at the range of financial reporting models available for use in the Australian higher education sector, the possible application of activity-based costing (ABC) in faculties and the eventual rejection of ABC in favour of a more qualitative model designed specifically for use in one institution, in a particular Faculty. The…

  14. Identification of residues in ABCG2 affecting protein trafficking and drug transport, using co-evolutionary analysis of ABCG sequences.

    PubMed

    Haider, Ameena J; Cox, Megan H; Jones, Natalie; Goode, Alice J; Bridge, Katherine S; Wong, Kelvin; Briggs, Deborah; Kerr, Ian D

    2015-07-17

    ABCG2 is an ABC (ATP-binding cassette) transporter with a physiological role in urate transport in the kidney and is also implicated in multi-drug efflux from a number of organs in the body. The trafficking of the protein and the mechanism by which it recognizes and transports diverse drugs are important areas of research. In the current study, we have made a series of single amino acid mutations in ABCG2 on the basis of sequence analysis. Mutant isoforms were characterized for cell surface expression and function. One mutant (I573A) showed disrupted glycosylation and reduced trafficking kinetics. In contrast with many ABC transporter folding mutations which appear to be 'rescued' by chemical chaperones or low temperature incubation, the I573A mutation was not enriched at the cell surface by either treatment, with the majority of the protein being retained in the endoplasmic reticulum (ER). Two other mutations (P485A and M549A) showed distinct effects on transport of ABCG2 substrates reinforcing the role of TM helix 3 in drug recognition and transport and indicating the presence of intracellular coupling regions in ABCG2. © 2015 Authors.

  15. Oxidative Stress in HIV Infection and Alcohol Use: Role of Redox Signals in Modulation of Lipid Rafts and ATP-Binding Cassette Transporters.

    PubMed

    Thangavel, Samikkannu; Mulet, Carmen T; Atluri, Venkata S R; Agudelo, Marisela; Rosenberg, Rhonda; Devieux, Jessy G; Nair, Madhavan P N

    2018-02-01

    Human immunodeficiency virus (HIV) infection induces oxidative stress and alcohol use accelerates disease progression, subsequently causing immune dysfunction. However, HIV and alcohol impact on lipid rafts-mediated immune dysfunction remains unknown. In this study, we investigate the modulation by which oxidative stress induces reactive oxygen species (ROS) affecting redox expression, lipid rafts caveiloin-1, ATP-binding cassette (ABC) transporters, and transcriptional sterol regulatory element-binding protein (SREBP) gene and protein modification and how these mechanisms are associated with arachidonic acid (AA) metabolites in HIV positive alcohol users, and how they escalate immune dysfunction. In both alcohol using HIV-positive human subjects and in vitro studies of alcohol with HIV-1 gp120 protein in peripheral blood mononuclear cells, increased ROS production significantly affected redox expression in glutathione synthetase (GSS), super oxide dismutase (SOD), and glutathione peroxidase (GPx), and subsequently impacted lipid rafts Cav-1, ABC transporters ABCA1, ABCG1, ABCB1, and ABCG4, and SREBP transcription. The increased level of rate-limiting enzyme 3-hydroxy-3-methylglutaryl HMG-CoA reductase (HMGCR), subsequently, inhibited 7-dehydrocholesterol reductase (DHCR-7). Moreover, the expression of cyclooxygenase-2 (COX-2) and lipoxygenase-5 (5-LOX) mRNA and protein modification tentatively increased the levels of prostaglandin E2 synthases (PGE 2 ) in plasma when compared with either HIV or alcohol alone. This article suggests for the first time that the redox inhibition affects lipid rafts, ABC-transporter, and SREBP transcription and modulates AA metabolites, serving as an important intermediate signaling network during immune cell dysfunction in HIV-positive alcohol users. These findings indicate that HIV infection induces oxidative stress and redox inhibition, affecting lipid rafts and ABC transports, subsequently upregulating AA metabolites and leading to

  16. Short interspersed elements (SINEs) of the Geomyoidea superfamily rodents.

    PubMed

    Gogolevsky, Konstantin P; Kramerov, Dmitri A

    2006-05-24

    A new short interspersed element (SINE) was isolated from the genome of desert kangaroo rat (Dipodomys deserti) using single-primer PCR. This SINE consists of two monomers: the left monomer (IDL) resembles rodent ID element and other tRNAAla(CGC)-derived SINEs, whereas the right one (Geo) shows no similarity with known SINE sequences. PCR and hybridization analyses demonstrated that IDL-Geo SINE is restricted to the rodent superfamily Geomyoidea (families Geomyidea and Heteromyidea). Isolation and analysis of IDL-Geo from California pocket mouse (Chaetodipus californicus) and Botta's pocket gopher (Thomomys bottae) revealed some species-specific features of this SINE family. The structure and evolution of known dimeric SINEs are discussed.

  17. Aldehyde Dehydrogenase Gene Superfamily in Populus: Organization and Expression Divergence between Paralogous Gene Pairs.

    PubMed

    Tian, Feng-Xia; Zang, Jian-Lei; Wang, Tan; Xie, Yu-Li; Zhang, Jin; Hu, Jian-Jun

    2015-01-01

    Aldehyde dehydrogenases (ALDHs) constitute a superfamily of NAD(P)+-dependent enzymes that catalyze the irreversible oxidation of a wide range of reactive aldehydes to their corresponding nontoxic carboxylic acids. ALDHs have been studied in many organisms from bacteria to mammals; however, no systematic analyses incorporating genome organization, gene structure, expression profiles, and cis-acting elements have been conducted in the model tree species Populus trichocarpa thus far. In this study, a comprehensive analysis of the Populus ALDH gene superfamily was performed. A total of 26 Populus ALDH genes were found to be distributed across 12 chromosomes. Genomic organization analysis indicated that purifying selection may have played a pivotal role in the retention and maintenance of PtALDH gene families. The exon-intron organizations of PtALDHs were highly conserved within the same family, suggesting that the members of the same family also may have conserved functionalities. Microarray data and qRT-PCR analysis indicated that most PtALDHs had distinct tissue-specific expression patterns. The specificity of cis-acting elements in the promoter regions of the PtALDHs and the divergence of expression patterns between nine paralogous PtALDH gene pairs suggested that gene duplications may have freed the duplicate genes from the functional constraints. The expression levels of some ALDHs were up- or down-regulated by various abiotic stresses, implying that the products of these genes may be involved in the adaptation of Populus to abiotic stresses. Overall, the data obtained from our investigation contribute to a better understanding of the complexity of the Populus ALDH gene superfamily and provide insights into the function and evolution of ALDH gene families in vascular plants.

  18. Solitary BioY Proteins Mediate Biotin Transport into Recombinant Escherichia coli

    PubMed Central

    Finkenwirth, Friedrich; Kirsch, Franziska

    2013-01-01

    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. PMID:23836870

  19. Crystal structure of the vitamin B3 transporter PnuC, a full-length SWEET homolog.

    PubMed

    Jaehme, Michael; Guskov, Albert; Slotboom, Dirk Jan

    2014-11-01

    PnuC transporters catalyze cellular uptake of the NAD+ precursor nicotinamide riboside (NR) and belong to a large superfamily that includes the SWEET sugar transporters. We present a crystal structure of Neisseria mucosa PnuC, which adopts a highly symmetrical fold with 3+1+3 membrane topology not previously observed in any protein. The high symmetry of PnuC with a single NR bound in the center suggests a simple alternating-access translocation mechanism.

  20. Downregulation of miR-29a/b/c in placenta accreta inhibits apoptosis of implantation site intermediate trophoblast cells by targeting MCL1.

    PubMed

    Gu, Yongzhong; Bian, Yuehong; Xu, Xiaofei; Wang, Xietong; Zuo, Changting; Meng, Jinlai; Li, Hongyan; Zhao, Shigang; Ning, Yunnan; Cao, Yongzhi; Huang, Tao; Yan, Junhao; Chen, Zi-Jiang

    2016-12-01

    Placenta accreta is defined as abnormal adhesion of placental villi to the uterine myometrium. Although this condition has become more common as a result of the increasing rate of cesarean sections, the underlying causative mechanism(s) remain elusive. Because microRNA-29a/b/c (miR-29a/b/c) have been shown to play important roles in placental development, this study evaluated the roles of these microRNAs in placenta accreta. Expression of miR-29a/b/c and myeloid cell leukemia-1 (MCL1) were quantified in patient tissues and HTR8/SVneo trophoblast cells using the real-time quantitative polymerase chain reaction. Western blotting was used to analyze expression of the MCL1 protein in HTR8/SVneo trophoblast cells with altered expression of miR-29a/b/c. To determine their role in apoptosis, miR-29a/b/c were overexpressed in HTR-8/SVneo cells, and levels of apoptosis were analyzed by flow cytometry. Luciferase activity assays were used to determine whether MCL1 is a target gene of miR-29a/b/c. Expression of miR-29a/b/c was significantly lower in creta sites compared to noncreta sites (p = 0.018, 0.041, and 0.022, respectively), but expression of MCL1 was upregulated in creta sites (p = 0.039). MCL1 expression was significantly downregulated in HTR-8/SVneo cells overexpressing miR-29a/b/c (p = 0.002, 0.008, and 0.013, respectively). Luciferase activity assays revealed that miR-29a/b/c directly target the 3' untranslated region of MCL1 in 293T cells. Over-expression of miR-29a/b/c induced apoptosis in the HTR-8/SVneo trophoblast cell line. Moreover, histopathological evaluation revealed that the number of implantation site intermediate trophoblast (ISIT) cells was increased in creta sites and that these cells were positive for MCL1. Our results demonstrate that in placenta accreta, miR-29a/b/c inhibits apoptosis of ISIT cells by targeting MCL1. These findings provide new insights into the pathogenesis of placenta accreta. Copyright © 2016 Elsevier Ltd. All rights

  1. Reproducibility of lung tumor position and reduction of lung mass within the planning target volume using active breathing control (ABC).

    PubMed

    Cheung, Patrick C F; Sixel, Katharina E; Tirona, Romeo; Ung, Yee C

    2003-12-01

    The active breathing control (ABC) device allows for temporary immobilization of respiratory motion by implementing a breath hold at a predefined relative lung volume and air flow direction. The purpose of this study was to quantitatively evaluate the ability of the ABC device to immobilize peripheral lung tumors at a reproducible position, increase total lung volume, and thereby reduce lung mass within the planning target volume (PTV). Ten patients with peripheral non-small-cell lung cancer tumors undergoing radiotherapy had CT scans of their thorax with and without ABC inspiration breath hold during the first 5 days of treatment. Total lung volumes were determined from the CT data sets. Each peripheral lung tumor was contoured by one physician on all CT scans to generate gross tumor volumes (GTVs). The lung density and mass contained within a 1.5-cm PTV margin around each peripheral tumor was calculated using CT numbers. Using the center of the GTV from the Day 1 ABC scan as the reference, the displacement of subsequent GTV centers on Days 2 to 5 for each patient with ABC applied was calculated in three dimensions. With the use of ABC inspiration breath hold, total lung volumes increased by an average of 42%. This resulted in an average decrease in lung mass of 18% within a standard 1.5-cm PTV margin around the GTV. The average (+/- standard deviation) displacement of GTV centers with ABC breath hold applied was 0.3 mm (+/- 1.8 mm), 1.2 mm (+/- 2.3 mm), and 1.1 mm (+/- 3.5 mm) in the lateral direction, anterior-posterior direction, and superior-inferior direction, respectively. Results from this study indicate that there remains some inter-breath hold variability in peripheral lung tumor position with the use of ABC inspiration breath hold, which prevents significant PTV margin reduction. However, lung volumes can significantly increase, thereby decreasing the mass of lung within a standard PTV.

  2. A Trk/HKT-type K+ transporter from Trypanosoma brucei.

    PubMed

    Mosimann, Marc; Goshima, Shinobu; Wenzler, Tanja; Lüscher, Alexandra; Uozumi, Nobuyuki; Mäser, Pascal

    2010-04-01

    The molecular mechanisms of K(+) homeostasis are only poorly understood for protozoan parasites. Trypanosoma brucei subsp. parasites, the causative agents of human sleeping sickness and nagana, are strictly extracellular and need to actively concentrate K(+) from their hosts' body fluids. The T. brucei genome contains two putative K(+) channel genes, yet the trypanosomes are insensitive to K(+) antagonists and K(+) channel-blocking agents, and they do not spontaneously depolarize in response to high extracellular K(+) concentrations. However, the trypanosomes are extremely sensitive to K(+) ionophores such as valinomycin. Surprisingly, T. brucei possesses a member of the Trk/HKT superfamily of monovalent cation permeases which so far had only been known from bacteria, archaea, fungi, and plants. The protein was named TbHKT1 and functions as a Na(+)-independent K(+) transporter when expressed in Escherichia coli, Saccharomyces cerevisiae, or Xenopus laevis oocytes. In trypanosomes, TbHKT1 is expressed in both the mammalian bloodstream stage and the Tsetse fly midgut stage; however, RNA interference (RNAi)-mediated silencing of TbHKT1 expression did not produce a growth phenotype in either stage. The presence of HKT genes in trypanosomatids adds a further piece to the enigmatic phylogeny of the Trk/HKT superfamily of K(+) transporters. Parsimonial analysis suggests that the transporters were present in the first eukaryotes but subsequently lost in several of the major eukaryotic lineages, in at least four independent events.

  3. The ABCs of School Choice: The Comprehensive Guide to Every Private School Choice Program in America. 2013 Edition

    ERIC Educational Resources Information Center

    Friedman Foundation for Educational Choice, 2013

    2013-01-01

    "The ABCs of School Choice" is the most comprehensive guide to every private school choice program in America, showcasing the voucher, tax-credit scholarship, education savings accounts, and individual tax credit/deduction programs currently operating in 21 states and Washington, D.C. "The ABCs of School Choice" provides policymakers, advocates,…

  4. As Easy as ABC: Re-engineering the Cost Accounting System.

    ERIC Educational Resources Information Center

    Trussel, John M.; Bitner, Larry N.

    1996-01-01

    To be useful for management decision making, the college or university's cost accounting system must capture and measure improvements. Activity-based costing (ABC), which determines more accurately the full costs of services and products, tracks improvements and should proceed alongside reengineering of institutional accounting. Guidelines are…

  5. Historical perspectives on tumor necrosis factor and its superfamily: 25 years later, a golden journey

    PubMed Central

    Gupta, Subash C.; Kim, Ji Hye

    2012-01-01

    Although activity that induced tumor regression was observed and termed tumor necrosis factor (TNF) as early as the 1960s, the true identity of TNF was not clear until 1984, when Aggarwal and coworkers reported, for the first time, the isolation of 2 cytotoxic factors: one, derived from macrophages (molecular mass 17 kDa), was named TNF, and the second, derived from lymphocytes (20 kDa), was named lymphotoxin. Because the 2 cytotoxic factors exhibited 50% amino acid sequence homology and bound to the same receptor, they came to be called TNF-α and TNF-β. Identification of the protein sequences led to cloning of their cDNA. Based on sequence homology to TNF-α, now a total of 19 members of the TNF superfamily have been identified, along with 29 interacting receptors, and several molecules that interact with the cytoplasmic domain of these receptors. The roles of the TNF superfamily in inflammation, apoptosis, proliferation, invasion, angiogenesis, metastasis, and morphogenesis have been documented. Their roles in immunologic, cardiovascular, neurologic, pulmonary, and metabolic diseases are becoming apparent. TNF superfamily members are active targets for drug development, as indicated by the recent approval and expanding market of TNF blockers used to treat rheumatoid arthritis, psoriasis, Crohns disease, and osteoporosis, with a total market of more than US $20 billion. As we learn more about this family, more therapeutics will probably emerge. In this review, we summarize the initial discovery of TNF-α, and the insights gained regarding the roles of this molecule and its related family members in normal physiology and disease. PMID:22053109

  6. Plasma membrane dynamics and tetrameric organisation of ABCG2 transporters in mammalian cells revealed by single particle imaging techniques.

    PubMed

    Wong, Kelvin; Briddon, Stephen J; Holliday, Nicholas D; Kerr, Ian D

    2016-01-01

    ABCG2 is one of three human ATP binding cassette (ABC) transporters involved in the export from cells of a chemically and structurally diverse range of compounds. This multidrug efflux capability, together with a broad tissue distribution in the body, means that ABCG2 exerts a range of effects on normal physiology such as kidney urate transport, as well as contributing towards the pharmacokinetic profiles of many exogenous drugs. The primary sequence of ABCG2 contains only half the number of domains required for a functioning ABC transporter and so it must oligomerise in order to function, yet its oligomeric state in intact cell membranes remains uncharacterized. We have analysed ABCG2 in living cell membranes using a combination of fluorescence correlation spectroscopy, photon counting histogram analysis, and stepwise photobleaching to demonstrate a predominantly tetrameric structure for ABCG2 in the presence or absence of transport substrates. These results provide the essential basis for exploring pharmacological manipulation of oligomeric state as a strategy to modulate ABCG2 activity in future selective therapeutics. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. ATP Hydrolysis Induced Conformational Changes in the Vitamin B12 Transporter BtuCD Revealed by MD Simulations

    PubMed Central

    Pan, Chao; Weng, Jingwei; Wang, Wenning

    2016-01-01

    ATP binding cassette (ABC) transporters utilize the energy of ATP hydrolysis to uni-directionally transport substrates across cell membrane. ATP hydrolysis occurs at the nucleotide-binding domain (NBD) dimer interface of ABC transporters, whereas substrate translocation takes place at the translocation pathway between the transmembrane domains (TMDs), which is more than 30 angstroms away from the NBD dimer interface. This raises the question of how the hydrolysis energy released at NBDs is “transmitted” to trigger the conformational changes at TMDs. Using molecular dynamics (MD) simulations, we studied the post-hydrolysis state of the vitamin B12 importer BtuCD. Totally 3-μs MD trajectories demonstrate a predominantly asymmetric arrangement of the NBD dimer interface, with the ADP-bound site disrupted and the ATP-bound site preserved in most of the trajectories. TMDs response to ATP hydrolysis by separation of the L-loops and opening of the cytoplasmic gate II, indicating that hydrolysis of one ATP could facilitate substrate translocation by opening the cytoplasmic end of translocation pathway. It was also found that motions of the L-loops and the cytoplasmic gate II are coupled with each other through a contiguous interaction network involving a conserved Asn83 on the extended stretch preceding TM3 helix plus the cytoplasmic end of TM2/6/7 helix bundle. These findings entail a TMD-NBD communication mechanism for type II ABC importers. PMID:27870912

  8. Methanogenic heterodisulfide reductase (HdrABC-MvhAGD) uses two noncubane [4Fe-4S] clusters for reduction.

    PubMed

    Wagner, Tristan; Koch, Jürgen; Ermler, Ulrich; Shima, Seigo

    2017-08-18

    In methanogenic archaea, the carbon dioxide (CO 2 ) fixation and methane-forming steps are linked through the heterodisulfide reductase (HdrABC)-[NiFe]-hydrogenase (MvhAGD) complex that uses flavin-based electron bifurcation to reduce ferredoxin and the heterodisulfide of coenzymes M and B. Here, we present the structure of the native heterododecameric HdrABC-MvhAGD complex at 2.15-angstrom resolution. HdrB contains two noncubane [4Fe-4S] clusters composed of fused [3Fe-4S]-[2Fe-2S] units sharing 1 iron (Fe) and 1 sulfur (S), which were coordinated at the CCG motifs. Soaking experiments showed that the heterodisulfide is clamped between the two noncubane [4Fe-4S] clusters and homolytically cleaved, forming coenzyme M and B bound to each iron. Coenzymes are consecutively released upon one-by-one electron transfer. The HdrABC-MvhAGD atomic model serves as a structural template for numerous HdrABC homologs involved in diverse microbial metabolic pathways. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  9. Diversification of a single ancestral gene into a successful toxin superfamily in highly venomous Australian funnel-web spiders.

    PubMed

    Pineda, Sandy S; Sollod, Brianna L; Wilson, David; Darling, Aaron; Sunagar, Kartik; Undheim, Eivind A B; Kely, Laurence; Antunes, Agostinho; Fry, Bryan G; King, Glenn F

    2014-03-05

    Spiders have evolved pharmacologically complex venoms that serve to rapidly subdue prey and deter predators. The major toxic factors in most spider venoms are small, disulfide-rich peptides. While there is abundant evidence that snake venoms evolved by recruitment of genes encoding normal body proteins followed by extensive gene duplication accompanied by explosive structural and functional diversification, the evolutionary trajectory of spider-venom peptides is less clear. Here we present evidence of a spider-toxin superfamily encoding a high degree of sequence and functional diversity that has evolved via accelerated duplication and diversification of a single ancestral gene. The peptides within this toxin superfamily are translated as prepropeptides that are posttranslationally processed to yield the mature toxin. The N-terminal signal sequence, as well as the protease recognition site at the junction of the propeptide and mature toxin are conserved, whereas the remainder of the propeptide and mature toxin sequences are variable. All toxin transcripts within this superfamily exhibit a striking cysteine codon bias. We show that different pharmacological classes of toxins within this peptide superfamily evolved under different evolutionary selection pressures. Overall, this study reinforces the hypothesis that spiders use a combinatorial peptide library strategy to evolve a complex cocktail of peptide toxins that target neuronal receptors and ion channels in prey and predators. We show that the ω-hexatoxins that target insect voltage-gated calcium channels evolved under the influence of positive Darwinian selection in an episodic fashion, whereas the κ-hexatoxins that target insect calcium-activated potassium channels appear to be under negative selection. A majority of the diversifying sites in the ω-hexatoxins are concentrated on the molecular surface of the toxins, thereby facilitating neofunctionalisation leading to new toxin pharmacology.

  10. Beyond TNF: TNF superfamily cytokines as targets for the treatment of rheumatic diseases.

    PubMed

    Croft, Michael; Siegel, Richard M

    2017-04-01

    TNF blockers are highly efficacious at dampening inflammation and reducing symptoms in rheumatic diseases such as rheumatoid arthritis, psoriatic arthritis and ankylosing spondylitis, and also in nonrheumatic syndromes such as inflammatory bowel disease. As TNF belongs to a superfamily of 19 structurally related proteins that have both proinflammatory and anti-inflammatory activity, reagents that disrupt the interaction between proinflammatory TNF family cytokines and their receptors, or agonize the anti-inflammatory receptors, are being considered for the treatment of rheumatic diseases. Biologic agents that block B cell activating factor (BAFF) and receptor activator of nuclear factor-κB ligand (RANKL) have been approved for the treatment of systemic lupus erythematosus and osteoporosis, respectively. In this Review, we focus on additional members of the TNF superfamily that could be relevant for the pathogenesis of rheumatic disease, including those that can strongly promote activity of immune cells or increase activity of tissue cells, as well as those that promote death pathways and might limit inflammation. We examine preclinical mouse and human data linking these molecules to the control of damage in the joints, muscle, bone or other tissues, and discuss their potential as targets for future therapy of rheumatic diseases.

  11. ConoDictor: a tool for prediction of conopeptide superfamilies.

    PubMed

    Koua, Dominique; Brauer, Age; Laht, Silja; Kaplinski, Lauris; Favreau, Philippe; Remm, Maido; Lisacek, Frédérique; Stöcklin, Reto

    2012-07-01

    ConoDictor is a tool that enables fast and accurate classification of conopeptides into superfamilies based on their amino acid sequence. ConoDictor combines predictions from two complementary approaches-profile hidden Markov models and generalized profiles. Results appear in a browser as tables that can be downloaded in various formats. This application is particularly valuable in view of the exponentially increasing number of conopeptides that are being identified. ConoDictor was written in Perl using the common gateway interface module with a php submission page. Sequence matching is performed with hmmsearch from HMMER 3 and ps_scan.pl from the pftools 2.3 package. ConoDictor is freely accessible at http://conco.ebc.ee.

  12. The TNF receptor and Ig superfamily members form an integrated signaling circuit controlling dendritic cell homeostasis

    PubMed Central

    De Trez, Carl; Ware, Carl F.

    2008-01-01

    Dendritic cells (DC) constitute the most potent antigen presenting cells of the immune system, playing a key role bridging innate and adaptive immune responses. Specialized DC subsets differ depending on their origin, tissue location and the influence of trophic factors, the latter remain to be fully understood. Stromal cell and myeloid-associated Lymphotoxin-β receptor (LTβR) signaling is required for the local proliferation of lymphoid tissue DC. This review focuses the LTβR signaling cascade as a crucial positive trophic signal in the homeostasis of DC subsets. The noncanonical coreceptor pathway comprised of the Immunoglobulin (Ig) superfamily member, B and T lymphocyte attenuator (BTLA) and TNFR superfamily member, Herpesvirus entry mediator (HVEM) counter regulates the trophic signaling by LTβR. Together both pathways form an integrated signaling circuit achieving homeostasis of DC subsets. PMID:18511331

  13. Identification of Transport-critical Residues in a Folate Transporter from the Folate-Biopterin Transporter (FBT) Family*

    PubMed Central

    Eudes, Aymerick; Kunji, Edmund R. S.; Noiriel, Alexandre; Klaus, Sebastian M. J.; Vickers, Tim J.; Beverley, Stephen M.; Gregory, Jesse F.; Hanson, Andrew D.

    2010-01-01

    The Synechocystis Slr0642 protein and its plastidial Arabidopsis (Arabidopsis thaliana) ortholog At2g32040 belong to the folate-biopterin transporter (FBT) family within the major facilitator superfamily. Both proteins transport folates when expressed in Escherichia coli. Because the structural requirements for transport activity are not known for any FBT protein, we applied mutational analysis to identify residues that are critical to transport and interpreted the results using a comparative structural model based on E. coli lactose permease. Folate transport was assessed via the growth of an E. coli pabA abgT strain, which cannot synthesize or take up folates or p-aminobenzoylglutamate. In total, 47 residues were replaced with Cys or Ala. Mutations at 22 positions abolished folate uptake without affecting Slr0642 expression in membranes, whereas other mutations had no effect. Residues important for function mostly line the predicted central cavity and are concentrated in the core α-helices H1, H4, H7, and H10. The essential residue locations are consistent with a folate-binding site lying roughly equidistant from both faces of the transporter. Arabidopsis has eight FBT proteins besides At2g32040, often lacking conserved critical residues. When six of these proteins were expressed in E. coli or in Leishmania folate or pterin transporter mutants, none showed evidence of folate or pterin transport activity, and only At2g32040 was isolated by functional screening of Arabidopsis cDNA libraries in E. coli. Such negative data could reflect roles in transport of other substrates. These studies provide the first insights into the native structure and catalytic mechanism of FBT family carriers. PMID:19923217

  14. Response to Bile Salts in Clinical Strains of Acinetobacter baumannii Lacking the AdeABC Efflux Pump: Virulence Associated with Quorum Sensing

    PubMed Central

    López, Maria; Blasco, Lucia; Gato, Eva; Perez, Astrid; Fernández-Garcia, Laura; Martínez-Martinez, Luis; Fernández-Cuenca, Felipe; Rodríguez-Baño, Jesús; Pascual, Alvaro; Bou, German; Tomás, Maria

    2017-01-01

    Introduction: Acinetobacter baumannii is an opportunistic nosocomial pathogen associated with multiple infections. This pathogen usually colonizes (first stage of microbial infection) host tissues that are in contact with the external environment. As one of the sites of entry in human hosts is the gastrointestinal tract, the pathogen must be capable of tolerating bile salts. However, studies analyzing the molecular characteristics involved in the response to bile salts in clinical strains of A. baumannii are scarce. Material and Methods: Microbiological and transcriptional studies (arrays and RT-PCR) in the response to bile salts were carried out in isogenic (A. baumanni ΔadeB ATCC 17978 and A. baumannii ΔadeL ATCC 17978) and clinical strains from clone ST79/PFGE-HUI-1 which is characterized by lacking the AdeABC efflux pump and by overexpression the AdeFGH efflux pump. Results and Discussion: In presence of bile salts, in addition to the glutamate/aspartate transporter were found overexpressed in A. baumannii ΔadeB ATCC 17978, the virulence factors (surface motility, biofilm, and Type VI Secretion System) which are associated with activation of the Quorum Sensing system. Overexpression of these factors was confirmed in clinical strains of clone ST79/PFGE-HUI-1. Conclusions: This the first study about the adaptive response to bile salts investigating the molecular and microbiological characteristics in response to bile salts of an isogenic model of A. baumannii ATCC 17978 and clinical isolates of A. baumannii (clinical strains of ST79/PFGE-HUI-1) lacking the main RND efflux pump (AdeABC). Clinical isolates of A. baumannii lacking the AdeABC efflux pump (clone ST79/PFGE-HUI-1) displayed a new clinical profile (increased invasiveness) possibly associated with the response to stress conditions (such as the presence of bile salts). PMID:28536672

  15. The Reign of Confusion: ABC and the "Crisis in Iran."

    ERIC Educational Resources Information Center

    Palmerton, Patricia R.

    A study examined reports broadcast by ABC News between November 8, 1979 and December 7, 1979 in its series entitled "Crisis in Iran: America Held Hostage." Transcripts of approximately 50% of actual broadcasts were subjected to rhetorical critical analysis, from which the finding emerged that confusion was the predominant characteristic…

  16. The ABCs of School Choice: The Comprehensive Guide to Every Private School Choice Program in America. 2016 Edition

    ERIC Educational Resources Information Center

    Friedman Foundation for Educational Choice, 2016

    2016-01-01

    "The ABCs of School Choice" is a comprehensive, data-rich guide to every private school choice program in America. The 2016 edition of "The ABCs of School Choice" is the best yet, not just because of the new look and the fantastic growth in the number of school choice programs, but also because it is now paired with a…

  17. Nonlinear resonances in the ABC-flow

    NASA Astrophysics Data System (ADS)

    Didov, A. A.; Uleysky, M. Yu.

    2018-01-01

    In this paper, we study resonances of the ABC-flow in the near integrable case ( C ≪1 ). This is an interesting example of a Hamiltonian system with 3/2 degrees of freedom in which simultaneous existence of two resonances of the same order is possible. Analytical conditions of the resonance existence are received. It is shown numerically that the largest n :1 (n = 1, 2, 3) resonances exist, and their energies are equal to theoretical energies in the near integrable case. We provide analytical and numerical evidences for existence of two branches of the two largest n :1 (n = 1, 2) resonances in the region of finite motion.

  18. Role of the Fur Regulon in Iron Transport in Bacillus subtilis

    PubMed Central

    Ollinger, Juliane; Song, Kyung-Bok; Antelmann, Haike; Hecker, Michael; Helmann, John D.

    2006-01-01

    The Bacillus subtilis ferric uptake regulator (Fur) protein mediates the iron-dependent repression of at least 20 operons encoding ∼40 genes. We investigated the physiological roles of Fur-regulated genes by the construction of null mutations in 14 transcription units known or predicted to function in siderophore biosynthesis or iron uptake. We demonstrate that ywbLMN, encoding an elemental iron uptake system orthologous to the copper oxidase-dependent Fe(III) uptake system of Saccharomyces cerevisiae, is essential for growth in low iron minimal medium lacking citric acid. 2,3-Dihydroxybenzoyl-glycine (Itoic acid), the siderophore precursor produced by laboratory strains of B. subtilis, is of secondary importance. In the presence of citrate, the YfmCDEF ABC transporter is required for optimal growth. B. subtilis is unable to grow in minimal medium containing the iron chelator EDDHA unless the ability to synthesize the intact bacillibactin siderophore is restored (by the introduction of a functional sfp gene) or exogenous siderophores are provided. Utilization of the catecholate siderophores bacillibactin and enterobactin requires the FeuABC importer and the YusV ATPase. Utilization of hydroxamate siderophores requires the FhuBGC ABC transporter together with the FhuD (ferrichrome) or YxeB (ferrioxamine) substrate-binding proteins. Growth with schizokinen or arthrobactin is at least partially dependent on the YfhA YfiYZ importer and the YusV ATPase. We have also investigated the effects of a fur mutation on the proteome and documented the derepression of 11 Fur-regulated proteins, including a newly identified thioredoxin reductase homolog, YcgT. PMID:16672620

  19. Role of the Fur regulon in iron transport in Bacillus subtilis.

    PubMed

    Ollinger, Juliane; Song, Kyung-Bok; Antelmann, Haike; Hecker, Michael; Helmann, John D

    2006-05-01

    The Bacillus subtilis ferric uptake regulator (Fur) protein mediates the iron-dependent repression of at least 20 operons encoding approximately 40 genes. We investigated the physiological roles of Fur-regulated genes by the construction of null mutations in 14 transcription units known or predicted to function in siderophore biosynthesis or iron uptake. We demonstrate that ywbLMN, encoding an elemental iron uptake system orthologous to the copper oxidase-dependent Fe(III) uptake system of Saccharomyces cerevisiae, is essential for growth in low iron minimal medium lacking citric acid. 2,3-Dihydroxybenzoyl-glycine (Itoic acid), the siderophore precursor produced by laboratory strains of B. subtilis, is of secondary importance. In the presence of citrate, the YfmCDEF ABC transporter is required for optimal growth. B. subtilis is unable to grow in minimal medium containing the iron chelator EDDHA unless the ability to synthesize the intact bacillibactin siderophore is restored (by the introduction of a functional sfp gene) or exogenous siderophores are provided. Utilization of the catecholate siderophores bacillibactin and enterobactin requires the FeuABC importer and the YusV ATPase. Utilization of hydroxamate siderophores requires the FhuBGC ABC transporter together with the FhuD (ferrichrome) or YxeB (ferrioxamine) substrate-binding proteins. Growth with schizokinen or arthrobactin is at least partially dependent on the YfhA YfiYZ importer and the YusV ATPase. We have also investigated the effects of a fur mutation on the proteome and documented the derepression of 11 Fur-regulated proteins, including a newly identified thioredoxin reductase homolog, YcgT.

  20. Arsenic and Antimony Transporters in Eukaryotes

    PubMed Central

    Maciaszczyk-Dziubinska, Ewa; Wawrzycka, Donata; Wysocki, Robert

    2012-01-01

    Arsenic and antimony are toxic metalloids, naturally present in the environment and all organisms have developed pathways for their detoxification. The most effective metalloid tolerance systems in eukaryotes include downregulation of metalloid uptake, efflux out of the cell, and complexation with phytochelatin or glutathione followed by sequestration into the vacuole. Understanding of arsenic and antimony transport system is of high importance due to the increasing usage of arsenic-based drugs in the treatment of certain types of cancer and diseases caused by protozoan parasites as well as for the development of bio- and phytoremediation strategies for metalloid polluted areas. However, in contrast to prokaryotes, the knowledge about specific transporters of arsenic and antimony and the mechanisms of metalloid transport in eukaryotes has been very limited for a long time. Here, we review the recent advances in understanding of arsenic and antimony transport pathways in eukaryotes, including a dual role of aquaglyceroporins in uptake and efflux of metalloids, elucidation of arsenic transport mechanism by the yeast Acr3 transporter and its role in arsenic hyperaccumulation in ferns, identification of vacuolar transporters of arsenic-phytochelatin complexes in plants and forms of arsenic substrates recognized by mammalian ABC transporters. PMID:22489166