scholarly journals Role of the ABC transporter TruMDR2 in terbinafine, 4-nitroquinoline N-oxide and ethidium bromide susceptibility in Trichophyton rubrum

2006 ◽  
Vol 55 (8) ◽  
pp. 1093-1099 ◽  
Author(s):  
Ana Lúcia Fachin ◽  
Monica S. Ferreira-Nozawa ◽  
Walter Maccheroni ◽  
Nilce M. Martinez-Rossi
Keyword(s):  
Abc Transporter ◽  
Ethidium Bromide ◽  
Abc Transporters ◽  
Trichophyton Rubrum ◽  
Single Copy ◽  
Drug Susceptibility ◽  
Transmembrane Segments ◽  
Binding Domains ◽  
Abc Protein ◽  
Identical Nucleotide

A single-copy gene, designated TruMDR2, encoding an ATP-binding cassette (ABC) transporter was cloned and sequenced from the dermatophyte Trichophyton rubrum. The ORF of TruMDR2 was 4048 nt and the deduced amino acid sequence showed high homology with ABC transporters involved in drug efflux in other fungi. The encoded ABC protein predicted 12 transmembrane segments (TMSs) and two almost identical nucleotide-binding domains (NBDs) arranged in two halves in a (TMS6–NBD)2 configuration and could be classified as a member of the multidrug-resistance (MDR) class of ABC transporters. Northern blot analyses revealed an increased level of transcription of the TruMDR2 gene when mycelium was exposed to acriflavine, benomyl, ethidium bromide, ketoconazole, chloramphenicol, griseofulvin, fluconazole, imazalil, itraconazole, methotrexate, 4-nitroquinoline N-oxide (4NQO) or tioconazole. Disruption of the TruMDR2 gene rendered the mutant more sensitive to terbinafine, 4NQO and ethidium bromide than the control strain, suggesting that this transporter plays a role in modulating drug susceptibility in T. rubrum.

scholarly journals Insect ATP-Binding Cassette (ABC) Transporters: Roles in Xenobiotic Detoxification and Bt Insecticidal Activity

2019 ◽  
Vol 20 (11) ◽  
pp. 2829 ◽  
Author(s):  
Chao Wu ◽  
Swapan Chakrabarty ◽  
Minghui Jin ◽  
Kaiyu Liu ◽  
Yutao Xiao
Keyword(s):  
Abc Transporter ◽  
Conformational Changes ◽  
Abc Transporters ◽  
Insecticidal Activity ◽  
Atp Hydrolysis ◽  
Atp Binding ◽  
Bt Toxin ◽  
Xenobiotic Detoxification ◽  
Binding Domains ◽  
Atp Binding Cassette

ATP-binding cassette (ABC) transporters, a large class of transmembrane proteins, are widely found in organisms and play an important role in the transport of xenobiotics. Insect ABC transporters are involved in insecticide detoxification and Bacillus thuringiensis (Bt) toxin perforation. The complete ABC transporter is composed of two hydrophobic transmembrane domains (TMDs) and two nucleotide binding domains (NBDs). Conformational changes that are needed for their action are mediated by ATP hydrolysis. According to the similarity among their sequences and organization of conserved ATP-binding cassette domains, insect ABC transporters have been divided into eight subfamilies (ABCA–ABCH). This review describes the functions and mechanisms of ABC transporters in insecticide detoxification, plant toxic secondary metabolites transport and insecticidal activity of Bt toxin. With improved understanding of the role and mechanisms of ABC transporter in resistance to insecticides and Bt toxins, we can identify valuable target sites for developing new strategies to control pests and manage resistance and achieve green pest control.

scholarly journals Structure of Ycf1p reveals the transmembrane domain TMD0 and the regulatory region of ABCC transporters

2021 ◽  
Vol 118 (21) ◽  
pp. e2025853118
Author(s):  
Sarah C. Bickers ◽  
Samir Benlekbir ◽  
John L. Rubinstein ◽  
Voula Kanelis
Keyword(s):  
Abc Transporters ◽  
Posttranslational Modifications ◽  
Transmembrane Domain ◽  
Regulatory Region ◽  
Multidrug Resistance Proteins ◽  
Protein Activity ◽  
Resistance Proteins ◽  
Binding Domains ◽  
Abc Protein ◽  
R Region

ATP binding cassette (ABC) proteins typically function in active transport of solutes across membranes. The ABC core structure is composed of two transmembrane domains (TMD1 and TMD2) and two cytosolic nucleotide binding domains (NBD1 and NBD2). Some members of the C-subfamily of ABC (ABCC) proteins, including human multidrug resistance proteins (MRPs), also possess an N-terminal transmembrane domain (TMD0) that contains five transmembrane α-helices and is connected to the ABC core by the L0 linker. While TMD0 was resolved in SUR1, the atypical ABCC protein that is part of the hetero-octameric ATP-sensitive K+ channel, little is known about the structure of TMD0 in monomeric ABC transporters. Here, we present the structure of yeast cadmium factor 1 protein (Ycf1p), a homolog of human MRP1, determined by electron cryo-microscopy (cryo-EM). A comparison of Ycf1p, SUR1, and a structure of MRP1 that showed TMD0 at low resolution demonstrates that TMD0 can adopt different orientations relative to the ABC core, including a ∼145° rotation between Ycf1p and SUR1. The cryo-EM map also reveals that segments of the regulatory (R) region, which links NBD1 to TMD2 and was poorly resolved in earlier ABCC structures, interacts with the L0 linker, NBD1, and TMD2. These interactions, combined with fluorescence quenching experiments of isolated NBD1 with and without the R region, suggest how posttranslational modifications of the R region modulate ABC protein activity. Mapping known mutations from MRP2 and MRP6 onto the Ycf1p structure explains how mutations involving TMD0 and the R region of these proteins lead to disease.

scholarly journals Curcumin Modulates Efflux Mediated by Yeast ABC Multidrug Transporters and Is Synergistic with Antifungals

2009 ◽  
Vol 53 (8) ◽  
pp. 3256-3265 ◽  
Author(s):  
Monika Sharma ◽  
Raman Manoharlal ◽  
Suneet Shukla ◽  
Nidhi Puri ◽  
Tulika Prasad ◽  
...  
Keyword(s):  
Abc Transporters ◽  
Curcuma Longa ◽  
Drug Susceptibility ◽  
Drug Susceptibility Testing ◽  
Antifungal Drugs ◽  
Major Facilitator Superfamily ◽  
Multidrug Transporters ◽  
Abc Protein ◽  
Major Facilitator ◽  
Mfs Transporters

ABSTRACT Curcumin (CUR), a natural product of turmeric, from rhizomes of Curcuma longa, is a known agent of reversal of drug resistance phenotypes in cancer cells overexpressing ATP-binding cassette (ABC) transporters, viz., ABCB1, ABCG2, and ABCC1. In the present study, we evaluated whether CUR could also modulate multidrug transporters of yeasts that belong either to the ABC family or to the major facilitator superfamily (MFS). The effect of CUR on multidrug transporter proteins was demonstrated by examining rhodamine 6G (R6G) efflux in Saccharomyces cerevisiae cells overexpressing the Candida albicans ABC transporters Cdr1p and Cdr2p (CaCdr1p and CaCdr2p, respectively) and the MFS transporters CaMdr1p and S. cerevisiae Pdr5p. CUR decreased the extracellular concentration of R6G in ABC transporter-expressing cells but had no effect on methotrexate efflux mediated through the MFS transporter CaMdr1p. CUR competitively inhibited R6G efflux and the photolabeling of CaCdr1p by [125I]iodoarylazidoprazosin, a drug analogue of the substrate prazosin (50% inhibitory concentration, 14.2 μM). Notably, the mutant variants of CaCdr1p that displayed abrogated efflux of R6G also showed reduced modulation by CUR. Drug susceptibility testing of ABC protein-expressing cells by spot assays and checkerboard tests revealed that CUR was selectively synergistic with drug substrates such as R6G, ketoconazole, itraconazole, and miconazole but not with fluconazole, voriconazole, anisomycin, cycloheximide, or FK520. Taken together, our results provide the first evidence that CUR modulates only ABC multidrug transporters and could be exploited in combination with certain conventional antifungal drugs to reverse multidrug resistance in Candida cells.

scholarly journals Distinct allosteric mechanisms of first-generation MsbA inhibitors

2021 ◽  
Author(s):  
Francois A Thelot ◽  
Wenyi Zhang ◽  
KangKang Song ◽  
Chen Xu ◽  
Jing Huang ◽  
...  
Keyword(s):  
Abc Transporter ◽  
Abc Transporters ◽  
Drug Targets ◽  
Rational Design ◽  
First Generation ◽  
Atp Hydrolysis ◽  
Conformational Transition ◽  
Structural Data ◽  
Atpase Inhibitor ◽  
Binding Domains

Present in all kingdoms of life, ATP-binding cassette (ABC) transporters couple ATP hydrolysis to mechanical force and facilitate trafficking of diverse substrates across biological membranes. Although many ABC transporters are promising drug targets, their mechanisms of regulation by small molecule inhibitors remain largely unknown. Herein, we used the lipopolysaccharide (LPS) flippase MsbA, a prototypical ABC exporter, as a model system to probe mechanisms of allosteric modulation by compounds binding to the transmembrane domains (TMDs). Recent chemical screens have identified intriguing LPS transport inhibitors targeting MsbA: the ATPase stimulator TBT1 and the ATPase inhibitor G247. Despite preliminary biochemical and structural data, it is unclear how TBT1 and G247 bind to the MsbA TMDs yet induce opposite allosteric effect in the nucleotide-binding domains (NBDs). Through single-particle EM, mutagenesis and activity assay, we show that TBT1 and G247 bind adjacent yet separate locations in the TMDs, inducing drastic changes in TMD conformation and NBD positioning. Two TBT1 molecules asymmetrically occupy the LPS binding site to break the symmetry of MsbA, resulting in disordered transmembrane helices and decreased NBD distance. In this novel inhibited ABC transporter state, decreased distance between the NBDs causes stimulation of ATP hydrolysis yet LPS transport blockage. In contrast, G247 acts as a TMDs wedge, symmetrically increasing NBD separation and preventing conformational transition of MsbA. Our study uncovers the distinct mechanisms of the first-generation MsbA-specific inhibitors and demonstrates that rational design of substrate-mimicking compounds can be exploited to develop useful ABC transporter modulators.

scholarly journals The VirAB-VirSR-AnrAB Multicomponent System Is Involved in Resistance of Listeria monocytogenes EGD-e to Cephalosporins, Bacitracin, Nisin, Benzalkonium Chloride, and Ethidium Bromide

2019 ◽  
Vol 85 (20) ◽  
Author(s):  
Xiaobing Jiang ◽  
Yimin Geng ◽  
Siyu Ren ◽  
Tao Yu ◽  
Yi Li ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Abc Transporter ◽  
Ethidium Bromide ◽  
Abc Transporters ◽  
Benzalkonium Chloride ◽  
Multicomponent System ◽  
Tetracycline Resistance ◽  
Dilution Method ◽  
Agar Dilution Method ◽  
Content Type

ABSTRACT In Listeria monocytogenes, it has been proposed that the VirSR two-component signal transduction systems (TCSs) and two ATP-binding cassette (ABC) transporters, VirAB and AnrAB, constitute a complex TCS/ABC transporter system which has been recognized as a unique resistance mode. The role of the putative VirAB-VirSR-AnrAB system in antimicrobial resistance and the respective contributions of the members of the system to resistance were investigated in this study. We constructed gene deletion mutants of L. monocytogenes EGD-e and complemented strains of the mutants and determined MICs of antimicrobial agents against these strains against using the agar dilution method. We assessed the relative expression levels of target genes by reverse transcription-quantitative PCR (RT-qPCR) and measured promoter activity levels by β-galactosidase assays. Our results showed that the VirAB-VirSR-AnrAB system mediates not only nisin and bacitracin resistance but also resistance to cephalosporins, ethidium bromide (EtBr), and benzalkonium chloride (BC). In this system, two ABC transporters, VirAB and AnrAB, play distinct roles in cefotaxime resistance: the former is responsible only for antimicrobial sensing and signaling by VirSR, while the latter contributes to transportation of antimicrobials. Notably, VirAB itself, rather than the VirAB-VirSR-AnrAB system as a whole, contributes to kanamycin and tetracycline resistance. On the basis of the results obtained from this study, we speculate that VirAB acts as a sensor for VirSR in response to cephalosporins, bacitracin, nisin, EtBr, and BC, while VirAB itself plays a role in response to kanamycin and tetracycline in L. monocytogenes EGD-e. IMPORTANCE This report describes TCS/ABC transporter modules characterized in Listeria monocytogenes EGD-e. The modules consist of the VirSR TCS and the VirAB and AnrAB ABC transporters. Our results showed that this system is involved in nisin and bacitracin resistance, as well as resistance to cephalosporins, ethidium bromide (EtBr), and benzalkonium chloride (BC). In this system, VirAB is responsible only for antimicrobial sensing and signaling by VirSR, while AnrAB contributes to transportation of antimicrobials. Interestingly, VirAB itself, rather than the VirAB-VirSR-AnrAB system as a whole, contributes to kanamycin and tetracycline resistance.

scholarly journals The Human ATP-Binding Cassette (ABC) Transporter Superfamily

2021 ◽  
Author(s):  
Michael Dean ◽  
Karobi Moitra ◽  
Rando Allikmets
Keyword(s):  
Abc Transporter ◽  
Abc Transporters ◽  
Rare Variants ◽  
Mendelian Inheritance ◽  
Atp Binding ◽  
Evolutionary Diversification ◽  
Binding Domains ◽  
Cholesterol Levels ◽  
Human Disorders ◽  
Atp Binding Cassette

The ATP-binding cassette (ABC) transporter superfamily comprises membrane proteins that efflux various substrates across extra- and intra-cellular membranes. Mutations in ABC genes cause 21 human disorders or phenotypes with Mendelian inheritance, including cystic fibrosis, adrenoleukodystrophy, retinal degeneration, cholesterol, and bile transport defects. Common polymorphisms and rare variants in ABC genes are associated with several complex phenotypes such as gout, gallstones, and cholesterol levels. Overexpression or amplification of specific drug efflux genes contributes to chemotherapy multidrug resistance. Conservation of the ATP-binding domains of ABC transporters defines the superfamily members, and phylogenetic analysis groups the 48 human ABC transporters into seven distinct subfamilies. While the conservation of ABC genes across most vertebrate species is high, there is also considerable gene duplication, deletion, and evolutionary diversification.

scholarly journals Combining Mutations That Inhibit Two Distinct Steps of the ATP Hydrolysis Cycle Restores Wild-Type Function in the Lipopolysaccharide Transporter and Shows that ATP Binding Triggers Transport

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Brent W. Simpson ◽  
Karanbir S. Pahil ◽  
Tristan W. Owens ◽  
Emily A. Lundstedt ◽  
Rebecca M. Davis ◽  
...  
Keyword(s):  
Abc Transporter ◽  
Conformational Changes ◽  
Abc Transporters ◽  
Atp Binding ◽  
Gram Negative Bacteria ◽  
Wild Type ◽  
Lethal Effects ◽  
Gram Negative ◽  
Binding Domains ◽  
Transport Cycle

ABSTRACT ATP-binding cassette (ABC) transporters constitute a large family of proteins present in all domains of life. They are powered by dynamic ATPases that harness energy from binding and hydrolyzing ATP through a cycle that involves the closing and reopening of their two ATP-binding domains. The LptB2FGC exporter is an essential ABC transporter that assembles lipopolysaccharides (LPS) on the surface of Gram-negative bacteria to form a permeability barrier against many antibiotics. LptB2FGC extracts newly synthesized LPS molecules from the inner membrane and powers their transport across the periplasm and through the outer membrane. How LptB2FGC functions remains poorly understood. Here, we show that the C-terminal domain of the dimeric LptB ATPase is essential for LPS transport in Escherichia coli. Specific changes in the C-terminal domain of LptB cause LPS transport defects that can be repaired by intragenic suppressors altering the ATP-binding domains. Surprisingly, we found that each of two lethal changes in the ATP-binding and C-terminal domains of LptB, when present in combined form, suppressed the defects associated with the other to restore LPS transport to wild-type levels both in vivo and in vitro. We present biochemical evidence explaining the effect that each of these mutations has on LptB function and how the observed cosuppression results from the opposing lethal effects these changes have on the dimerization state of the LptB ATPase. We therefore propose that these sites modulate the closing and reopening of the LptB dimer, providing insight into how the LptB2FGC transporter cycles to export LPS to the cell surface and how to inhibit this essential envelope biogenesis process. IMPORTANCE Gram-negative bacteria are naturally resistant to many antibiotics because their surface is covered by the glycolipid LPS. Newly synthesized LPS is transported across the cell envelope by the multiprotein Lpt machinery, which includes LptB2FGC, an unusual ABC transporter that extracts LPS from the inner membrane. Like in other ABC transporters, the LptB2FGC transport cycle is driven by the cyclical conformational changes that a cytoplasmic, dimeric ATPase, LptB, undergoes when binding and hydrolyzing ATP. How these conformational changes are controlled in ABC transporters is poorly understood. Here, we identified two lethal changes in LptB that, when combined, remarkably restore wild-type transport function. Biochemical studies revealed that the two changes affect different steps in the transport cycle, having opposing, lethal effects on LptB’s dimerization cycle. Our work provides mechanistic details about the LptB2FGC extractor that could be used to develop Lpt inhibitors that would overcome the innate antibiotic resistance of Gram-negative bacteria.

scholarly journals Chemo-mechanical Coupling in the Transport Cycle of a Type II ABC Transporter

2018 ◽  
Author(s):  
Koichi Tamura ◽  
Hiroshi Sugimoto ◽  
Yoshitsugu Shiro ◽  
Yuji Sugita
Keyword(s):  
Atpase Activity ◽  
Abc Transporter ◽  
Conformational Changes ◽  
Abc Transporters ◽  
Md Simulations ◽  
Coupling Mechanism ◽  
Transmembrane Helices ◽  
Mechanical Coupling ◽  
Binding Domains ◽  
Wide Range

AbstractAT P -binding cassette (ABC) transporters are integral membrane proteins that translocate a wide range of substrates across biological membranes, harnessing free energy from the binding and hydrolysis of ATP. To understand the mechanism of the inward- to outward-facing transition that could be achieved by tight regulation of ATPase activity through extensive conformational changes of the protein, we applied template-based iterative all-atom molecular dynamics (MD) simulation to the heme ABC transporter BhuUV-T. The simulations, together with biased MDs, predict two new conformations of the protein, namely, occluded (Occ) and outward-facing (OF) conformations. The comparison between the inward-facing crystal structure and the predicted two structures shows atomic details of the gating motions at the transmembrane helices and dimerization of the nucleotide-binding domains (NBDs). The MD simulations further reveal a novel role of the ABC signature motifs (LSGG[Q/E]) at the NBDs in decelerating ATPase activity in the Occ form through sporadic flipping of the side chains of the LSGG[Q/E] catalytic serine residues. The orientational changes are coupled to loose NBD dimerization in the Occ state, whereas they are blocked in the OF form where the NBDs are tightly dimerized. The chemo-mechanical coupling mechanism may apply to other types of ABC transporters having the conserved LSGG[Q/E] signature motifs.

scholarly journals ABCA5 Resides in Lysosomes, and ABCA5 Knockout Mice Develop Lysosomal Disease-Like Symptoms

2005 ◽  
Vol 25 (10) ◽  
pp. 4138-4149 ◽  
Author(s):  
Yoshiyuki Kubo ◽  
Sayaka Sekiya ◽  
Megumi Ohigashi ◽  
Chiemi Takenaka ◽  
Kyoko Tamura ◽  
...  
Keyword(s):  
Thyroid Gland ◽  
Abc Transporter ◽  
Neural Cells ◽  
Newborn Mouse ◽  
Amino Acid Residues ◽  
Lysosomal Disease ◽  
Transmembrane Segments ◽  
Reverse Transcription Pcr ◽  
Localization Analysis ◽  
Immunohistochemical Studies

ABSTRACT ABCA5 is a member of the ABC transporter A subfamily, and a mouse orthologue (mABCA5) in newborn mouse brain and neural cells was identified by reverse transcription-PCR. Full-length cDNA cloning revealed that mABCA5 consists of 1,642 amino acid residues and that its putative structure is that of a full-type ABC transporter having two sets of six transmembrane segments and a nucleotide binding domain. Immunohistochemical studies revealed that mABCA5 is expressed in brain, lung, heart, and thyroid gland. A subcellular localization analysis showed that mABCA5 is a resident of lysosomes and late endosomes. Abca5 − / − mice exhibited symptoms similar to those of several lysosomal diseases in heart, although no prominent abnormalities were found in brain or lung. They developed a dilated cardiomyopathy-like heart after reaching adulthood and died due to depression of the cardiovascular system. In addition, Abca5 − / − mice also exhibited exophthalmos and collapse of the thyroid gland. Therefore, ABCA5 is a protein related to a lysosomal disease and plays important roles, especially in cardiomyocytes and follicular cells.

scholarly journals Tn5406, a New Staphylococcal Transposon Conferring Resistance to Streptogramin A and Related Compounds Including Dalfopristin

2002 ◽  
Vol 46 (8) ◽  
pp. 2337-2343 ◽  
Author(s):  
Julien Haroche ◽  
Jeanine Allignet ◽  
Névine El Solh
Keyword(s):  
Staphylococcus Aureus ◽  
Insertion Site ◽  
Single Copy ◽  
Amino Acid Identity ◽  
Plasmid Copy ◽  
Acid Identity ◽  
Abc Protein ◽  
Single Plasmid ◽  
Insertion Sites ◽  
Related Compounds

ABSTRACT We characterized a new transposon, Tn5406 (5,467 bp), in a clinical isolate of Staphylococcus aureus (BM3327). It carries a variant of vgaA, which encodes a putative ABC protein conferring resistance to streptogramin A but not to mixtures of streptogramins A and B. It also carries three putative genes, the products of which exhibit significant similarities (61 to 73% amino acid identity) to the three transposases of the staphylococcal transposon Tn554. Like Tn554, Tn5406 failed to generate target repeats. In BM3327, the single copy of Tn5406 was inserted into the chromosomal att554 site, which is the preferential insertion site of Tn554. In three other independent S. aureus clinical isolates, Tn5406 was either present as a single plasmid copy (BM3318), as two chromosomal copies (BM3252), or both in the chromosome and on a plasmid (BM3385). The Tn5406-carrying plasmids also contain two other genes, vgaB and vatB. The insertion sites of Tn5406 in BM3252 were studied: one copy was in att554, and one copy was in the additional SCCmec element. Amplification experiments revealed circular forms of Tn5406, indicating that this transposon might be active. To our knowledge, a transposon conferring resistance to streptogramin A and related compounds has not been previously described.

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