scholarly journals Novel MenA Inhibitors Are Bactericidal againstMycobacterium tuberculosisand Synergize with Electron Transport Chain Inhibitors

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
Bryan J. Berube ◽  
Dara Russell ◽  
Lina Castro ◽  
Seoung-ryoung Choi ◽  
Prabagaran Narayanasamy ◽  
...  
Keyword(s):  
Electron Transport ◽  
Drug Treatment ◽  
Electron Transport Chain ◽  
Drug Targets ◽  
Content Type ◽  
Atp Production ◽  
Highly Active ◽  
Enhanced Activity ◽  
Transport Chain ◽  
Long Course

ABSTRACTMycobacterium tuberculosisis the leading cause of morbidity and death resulting from infectious disease worldwide. The incredible disease burden, combined with the long course of drug treatment and an increasing incidence of antimicrobial resistance amongM. tuberculosisisolates, necessitates novel drugs and drug targets for treatment of this deadly pathogen. Recent work has produced several promising clinical candidates targeting components of the electron transport chain (ETC) ofM. tuberculosis, highlighting this pathway’s potential as a drug target. Menaquinone is an essential component of theM. tuberculosisETC, as it functions to shuttle electrons through the ETC to produce the electrochemical gradient required for ATP production for the cell. We show that inhibitors of MenA, a component of the menaquinone biosynthetic pathway, are highly active againstM. tuberculosis. MenA inhibitors are bactericidal againstM. tuberculosisunder both replicating and nonreplicating conditions, with 10-fold higher bactericidal activity against nutrient-starved bacteria than against replicating cultures. MenA inhibitors have enhanced activity in combination with bedaquiline, clofazimine, and inhibitors of QcrB, a component of the cytochromebc1oxidase. Together, these data support MenA as a viable target for drug treatment againstM. tuberculosis. MenA inhibitors not only killM. tuberculosisin a variety of physiological states but also show enhanced activity in combination with ETC inhibitors in various stages of clinical trial testing.

scholarly journals The Agr Quorum-Sensing System Regulates Fibronectin Binding but Not Hemolysis in the Absence of a Functional Electron Transport Chain

2014 ◽  
Vol 82 (10) ◽  
pp. 4337-4347 ◽  
Author(s):  
Vera Pader ◽  
Ellen H. James ◽  
Kimberley L. Painter ◽  
Sivaramesh Wigneshweraraj ◽  
Andrew M. Edwards
Keyword(s):  
Electron Transport ◽  
Electron Transport Chain ◽  
Content Type ◽  
Atp Production ◽  
Transport Chain ◽  
Mutant Strains ◽  
Enhanced Expression ◽  
Fibronectin Binding Protein ◽  
Small Colony ◽  
Fibronectin Binding

ABSTRACTStaphylococcus aureusis responsible for numerous chronic and recurrent infections, which are frequently associated with the emergence of small-colony variants (SCVs) that lack a functional electron transport chain. SCVs exhibit enhanced expression of fibronectin-binding protein (FnBP) and greatly reduced hemolysin production, although the basis for this is unclear. One hypothesis is that these phenotypes are a consequence of the reduced Agr activity of SCVs, while an alternative is that the lack of a functional electron transport chain and the resulting reduction in ATP production are responsible. Disruption of the electron transport chain ofS. aureusgenetically (hemBandmenD) or chemically, using 2-n-heptyl-4-hydroxyquinolineN-oxide (HQNO), inhibited both growth and Agr activity and conferred an SCV phenotype. Supplementation of the culture medium with synthetic autoinducing peptide (sAIP) significantly increased Agr expression in bothhemBmutant strains andS. aureusgrown with HQNO and significantly reduced staphylococcal adhesion to fibronectin. However, sAIP did not promote hemolysin expression inhemBmutant strains orS. aureusgrown with HQNO. Therefore, while Agr regulates fibronectin binding in SCVs, it cannot promote hemolysin production in the absence of a functional electron transport chain.

scholarly journals Identification of 4-Amino-Thieno[2,3-d]Pyrimidines as QcrB Inhibitors in Mycobacterium tuberculosis

mSphere ◽  
2019 ◽  
Vol 4 (5) ◽  
Author(s):  
Gregory A. Harrison ◽  
Anne E. Mayer Bridwell ◽  
Megh Singh ◽  
Keshav Jayaraman ◽  
Leslie A. Weiss ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Electron Transport ◽  
Electron Transport Chain ◽  
Drug Target ◽  
Drug Targets ◽  
Bacterial Infections ◽  
Nonsynonymous Mutation ◽  
Content Type ◽  
Transport Chain

ABSTRACT Antibiotic resistance is a global crisis that threatens our ability to treat bacterial infections, such as tuberculosis, caused by Mycobacterium tuberculosis. Of the 10 million cases of tuberculosis in 2017, approximately 19% of new cases and 43% of previously treated cases were caused by strains of M. tuberculosis resistant to at least one frontline antibiotic. There is a clear need for new therapies that target these genetically resistant strains. Here, we report the discovery of a new series of antimycobacterial compounds, 4-amino-thieno[2,3-d]pyrimidines, that potently inhibit the growth of M. tuberculosis. To elucidate the mechanism by which these compounds inhibit M. tuberculosis, we selected for mutants resistant to a representative 4-amino-thieno[2,3-d]pyrimidine and sequenced these strains to identify the mutations that confer resistance. We isolated a total of 12 resistant mutants, each of which harbored a nonsynonymous mutation in the gene qcrB, which encodes a subunit of the electron transport chain (ETC) enzyme cytochrome bc1 oxidoreductase, leading us to hypothesize that 4-amino-thieno[2,3-d]pyrimidines target this enzyme complex. We found that addition of 4-amino-thieno[2,3-d]pyrimidines to M. tuberculosis cultures resulted in a decrease in ATP levels, supporting our model that these compounds inhibit the M. tuberculosis ETC. Furthermore, 4-amino-thieno[2,3-d]pyrimidines had enhanced activity against a mutant of M. tuberculosis deficient in cytochrome bd oxidase, which is a hallmark of cytochrome bc1 inhibitors. Therefore, 4-amino-thieno[2,3-d]pyrimidines represent a novel series of QcrB inhibitors that build on the growing number of chemical scaffolds that are able to inhibit the mycobacterial cytochrome bc1 complex. IMPORTANCE The global tuberculosis (TB) epidemic has been exacerbated by the rise in drug-resistant TB cases worldwide. To tackle this crisis, it is necessary to identify new vulnerable drug targets in Mycobacterium tuberculosis, the causative agent of TB, and develop compounds that can inhibit the bacterium through novel mechanisms of action. The QcrB subunit of the electron transport chain enzyme cytochrome bc1 has recently been validated to be a potential drug target. In the current work, we report the discovery of a new class of QcrB inhibitors, 4-amino-thieno[2,3-d]pyrimidines, that potently inhibit M. tuberculosis growth in vitro. These compounds are chemically distinct from previously reported QcrB inhibitors, and therefore, 4-amino-thieno[2,3-d]pyrimidines represent a new scaffold that can be exploited to inhibit this drug target.

scholarly journals Physiological Evidence for Isopotential Tunneling in the Electron Transport Chain of Methane-Producing Archaea

2017 ◽  
Vol 83 (18) ◽  
Author(s):  
Nikolas Duszenko ◽  
Nicole R. Buan
Keyword(s):  
Escherichia Coli ◽  
Electron Transport ◽  
Stationary Phase ◽  
Exponential Growth ◽  
Electron Transport Chain ◽  
Methanosarcina Barkeri ◽  
Metabolic Efficiency ◽  
Methanosarcina Acetivorans ◽  
Content Type ◽  
Transport Chain

ABSTRACT Many, but not all, organisms use quinones to conserve energy in their electron transport chains. Fermentative bacteria and methane-producing archaea (methanogens) do not produce quinones but have devised other ways to generate ATP. Methanophenazine (MPh) is a unique membrane electron carrier found in Methanosarcina species that plays the same role as quinones in the electron transport chain. To extend the analogy between quinones and MPh, we compared the MPh pool sizes between two well-studied Methanosarcina species, Methanosarcina acetivorans C2A and Methanosarcina barkeri Fusaro, to the quinone pool size in the bacterium Escherichia coli. We found the quantity of MPh per cell increases as cultures transition from exponential growth to stationary phase, and absolute quantities of MPh were 3-fold higher in M. acetivorans than in M. barkeri. The concentration of MPh suggests the cell membrane of M. acetivorans, but not of M. barkeri, is electrically quantized as if it were a single conductive metal sheet and near optimal for rate of electron transport. Similarly, stationary (but not exponentially growing) E. coli cells also have electrically quantized membranes on the basis of quinone content. Consistent with our hypothesis, we demonstrated that the exogenous addition of phenazine increases the growth rate of M. barkeri three times that of M. acetivorans. Our work suggests electron flux through MPh is naturally higher in M. acetivorans than in M. barkeri and that hydrogen cycling is less efficient at conserving energy than scalar proton translocation using MPh. IMPORTANCE Can we grow more from less? The ability to optimize and manipulate metabolic efficiency in cells is the difference between commercially viable and nonviable renewable technologies. Much can be learned from methane-producing archaea (methanogens) which evolved a successful metabolic lifestyle under extreme thermodynamic constraints. Methanogens use highly efficient electron transport systems and supramolecular complexes to optimize electron and carbon flow to control biomass synthesis and the production of methane. Worldwide, methanogens are used to generate renewable methane for heat, electricity, and transportation. Our observations suggest Methanosarcina acetivorans, but not Methanosarcina barkeri, has electrically quantized membranes. Escherichia coli, a model facultative anaerobe, has optimal electron transport at the stationary phase but not during exponential growth. This study also suggests the metabolic efficiency of bacteria and archaea can be improved using exogenously supplied lipophilic electron carriers. The enhancement of methanogen electron transport through methanophenazine has the potential to increase renewable methane production at an industrial scale.

scholarly journals Age-Dependent Loss of Mitochondrial Function in Epithelial Tissue Can Be Reversed by Coenzyme Q10

2018 ◽  
Vol 2018 ◽  
pp. 1-8 ◽  
Author(s):  
Daniel Schniertshauer ◽  
Daniel Gebhard ◽  
Jörg Bergemann
Keyword(s):  
Electron Transport ◽  
Coenzyme Q10 ◽  
Mitochondrial Respiration ◽  
Electron Transport Chain ◽  
Epithelial Tissue ◽  
Atp Production ◽  
Transport Chain ◽  
Respiratory Parameters ◽  
Age Dependent ◽  
Theory Of Aging

The process of aging is characterized by the increase of age-associated disorders as well as severe diseases. Due to their role in the oxidative phosphorylation and thus the production of ATP which is crucial for many cellular processes, one reason for this could be found in the mitochondria. The accumulation of reactive oxygen species damaged mitochondrial DNA and proteins can induce mitochondrial dysfunction within the electron transport chain. According to the “mitochondrial theory of aging,” understanding the impact of harmful external influences on mitochondrial function is therefore essential for a better view on aging in general, but the measurement of mitochondrial respiration in skin cells from cell cultures cannot completely reflect the real situation in skin. Here, we describe a new method to measure the mitochondrial respiratory parameters in epithelial tissue derived from human skin biopsies using a XF24 extracellular flux analyzer to evaluate the effect of coenzyme Q10. We observed a decrease in mitochondrial respiration and ATP production with donor age corresponding to the “mitochondrial theory of aging.” For the first time ex vivo in human epidermis, we could show also a regeneration of mitochondrial respiratory parameters if the reduced form of coenzyme Q10, ubiquinol, was administered. In conclusion, an age-related decrease in mitochondrial respiration and ATP production was confirmed. Likewise, an increase in the respiratory parameters by the addition of coenzyme Q10 could also be shown. The fact that there is a significant effect of administered coenzyme Q10 on the respiratory parameters leads to the assumption that this is mainly caused by an increase in the electron transport chain. This method offers the possibility of testing age-dependent effects of various substances and their influence on the mitochondrial respiration parameters in human epithelial tissue.

scholarly journals Vibrio cholerae VciB Mediates Iron Reduction

2017 ◽  
Vol 199 (12) ◽  
Author(s):  
Eric D. Peng ◽  
Shelley M. Payne
Keyword(s):  
Electron Transport ◽  
Vibrio Cholerae ◽  
Ferrous Iron ◽  
Electron Transport Chain ◽  
Iron Reduction ◽  
Iron Transport ◽  
Ferric Iron ◽  
Transport Systems ◽  
Content Type ◽  
Transport Chain

ABSTRACT Vibrio cholerae is the causative agent of the severe diarrheal disease cholera. V. cholerae thrives within the human host, where it replicates to high numbers, but it also persists within the aquatic environments of ocean and brackish water. To survive within these nutritionally diverse environments, V. cholerae must encode the necessary tools to acquire the essential nutrient iron in all forms it may encounter. A prior study of systems involved in iron transport in V. cholerae revealed the existence of vciB, which, while unable to directly transport iron, stimulates the transport of iron through ferrous (Fe2+) iron transport systems. We demonstrate here a role for VciB in V. cholerae in which VciB stimulates the reduction of Fe3+ to Fe2+, which can be subsequently transported into the cell with the ferrous iron transporter Feo. Iron reduction is independent of functional iron transport but is associated with the electron transport chain. Comparative analysis of VciB orthologs suggests a similar role for other proteins in the VciB family. Our data indicate that VciB is a dimer located in the inner membrane with three transmembrane segments and a large periplasmic loop. Directed mutagenesis of the protein reveals two highly conserved histidine residues required for function. Taken together, our results support a model whereby VciB reduces ferric iron using energy from the electron transport chain. IMPORTANCE Vibrio cholerae is a prolific human pathogen and environmental organism. The acquisition of essential nutrients such as iron is critical for replication, and V. cholerae encodes a number of mechanisms to use iron from diverse environments. Here, we describe the V. cholerae protein VciB that increases the reduction of oxidized ferric iron (Fe3+) to the ferrous form (Fe2+), thus promoting iron acquisition through ferrous iron transporters. Analysis of VciB orthologs in Burkholderia and Aeromonas spp. suggest that they have a similar activity, allowing a functional assignment for this previously uncharacterized protein family. This study builds upon our understanding of proteins known to mediate iron reduction in bacteria.

scholarly journals Exploration of Nitrate Reductase Metabolic Pathway inCorynebacterium pseudotuberculosis

2017 ◽  
Vol 2017 ◽  
pp. 1-12 ◽  
Author(s):  
Sintia Almeida ◽  
Cassiana Sousa ◽  
Vinícius Abreu ◽  
Carlos Diniz ◽  
Elaine M. S. Dorneles ◽  
...  
Keyword(s):  
Electron Transport ◽  
Nitrate Reductase ◽  
Virulence Factors ◽  
Electron Transport Chain ◽  
Drug Targets ◽  
Carbon Sources ◽  
The Other ◽  
Ecological Niches ◽  
Transport Chain ◽  
In Silico Methods

Based on the ability of nitrate reductase synthesis,Corynebacterium pseudotuberculosisis classified into two biovars: Ovis and Equi. Due to the presence of nitrate reductase, the Equi biovar can survive in absence of oxygen. On the other hand, Ovis biovar that does not have nitrate reductase is able to adapt to various ecological niches and can grow on certain carbon sources. Apart from these two biovars, some other strains are also able to carry out the reduction of nitrate. The enzymes that are involved in electron transport chain are also identified by in silico methods. Findings about pathogen metabolism can contribute to the identification of relationship between nitrate reductase and theC. pseudotuberculosispathogenicity, virulence factors, and discovery of drug targets.

scholarly journals The dynamin-related GTPase Opa1 is required for glucose-stimulated ATP production in pancreatic beta cells

2011 ◽  
Vol 22 (13) ◽  
pp. 2235-2245 ◽  
Author(s):  
Zhongyan Zhang ◽  
Nobunao Wakabayashi ◽  
Junko Wakabayashi ◽  
Yasushi Tamura ◽  
Woo-Jin Song ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Electron Transport ◽  
Beta Cells ◽  
Pancreatic Beta Cells ◽  
Electron Transport Chain ◽  
Chain Complex ◽  
Atp Production ◽  
Transport Chain

Previous studies using in vitro cell culture systems have shown the role of the dynamin-related GTPase Opa1 in apoptosis prevention and mitochondrial DNA (mtDNA) maintenance. However, it remains to be tested whether these functions of Opa1 are physiologically important in vivo in mammals. Here, using the Cre-loxP system, we deleted mouse Opa1 in pancreatic beta cells, in which glucose-stimulated ATP production in mitochondria plays a key role in insulin secretion. Beta cells lacking Opa1 maintained normal copy numbers of mtDNA; however, the amount and activity of electron transport chain complex IV were significantly decreased, leading to impaired glucose-stimulated ATP production and insulin secretion. In addition, in Opa1-null beta cells, cell proliferation was impaired, whereas apoptosis was not promoted. Consequently, mice lacking Opa1 in beta cells develop hyperglycemia. The data suggest that the function of Opa1 in the maintenance of the electron transport chain is physiologically relevant in beta cells.

scholarly journals Genomic, Proteomic, and Biochemical Analysis of the Organohalide Respiratory Pathway in Desulfitobacterium dehalogenans

2014 ◽  
Vol 197 (5) ◽  
pp. 893-904 ◽  
Author(s):  
Thomas Kruse ◽  
Bram A. van de Pas ◽  
Ariane Atteia ◽  
Klaas Krab ◽  
Wilfred R. Hagen ◽  
...  
Keyword(s):  
Electron Transport ◽  
Electron Acceptor ◽  
Electron Transport Chain ◽  
Biochemical Analysis ◽  
Active Components ◽  
Organohalide Respiration ◽  
Content Type ◽  
Membrane Complex ◽  
Redox Active ◽  
Transport Chain

Desulfitobacterium dehalogenansis able to grow by organohalide respiration using 3-chloro-4-hydroxyphenyl acetate (Cl-OHPA) as an electron acceptor. We used a combination of genome sequencing, biochemical analysis of redox active components, and shotgun proteomics to study elements of the organohalide respiratory electron transport chain. The genome ofDesulfitobacterium dehalogenansJW/IU-DC1Tconsists of a single circular chromosome of 4,321,753 bp with a GC content of 44.97%. The genome contains 4,252 genes, including six rRNA operons and six predicted reductive dehalogenases. One of the reductive dehalogenases, CprA, is encoded by a well-characterizedcprTKZEBACDgene cluster. Redox active components were identified in concentrated suspensions of cells grown on formate and Cl-OHPA or formate and fumarate, using electron paramagnetic resonance (EPR), visible spectroscopy, and high-performance liquid chromatography (HPLC) analysis of membrane extracts. In cell suspensions, these components were reduced upon addition of formate and oxidized after addition of Cl-OHPA, indicating involvement in organohalide respiration. Genome analysis revealed genes that likely encode the identified components of the electron transport chain from formate to fumarate or Cl-OHPA. Data presented here suggest that the first part of the electron transport chain from formate to fumarate or Cl-OHPA is shared. Electrons are channeled from an outward-facing formate dehydrogenase via menaquinones to a fumarate reductase located at the cytoplasmic face of the membrane. When Cl-OHPA is the terminal electron acceptor, electrons are transferred from menaquinones to outward-facing CprA, via an as-yet-unidentified membrane complex, and potentially an extracellular flavoprotein acting as an electron shuttle between the quinol dehydrogenase membrane complex and CprA.

scholarly journals Electron Transport Chain Inhibition Suppresses CD40 Expression and Sensitizes Chronic Lymphocytic Leukaemia Cells to Venetoclax

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 35-35
Author(s):  
Zhenghao Chen ◽  
Gaspard Cretenet ◽  
Valeria Carnazzo ◽  
Gerritje J. W. van der Windt ◽  
Arnon P. Kater ◽  
...  
Keyword(s):  
Electron Transport ◽  
Research Funding ◽  
Electron Transport Chain ◽  
Specific Inhibitor ◽  
Cd40 Ligation ◽  
Atp Production ◽  
Transport Chain ◽  
Cd40 Stimulation ◽  
Cd40 Expression

Alterations in expression of specifically BCL-XL and MCL-1 dictate sensitivity of CLL cells to the Bcl-2 specific inhibitor venetoclax (VEN). We and others have shown upregulation of these anti-apoptotic proteins by interaction of CLL cells with CD4+ T helper cells within their lymph node microenvironment (LN-ME) mediated by CD40 signalling. We also reported significant metabolic changes of LN-ME activated CLL cells but whether metabolic alterations can be linked to VEN resistance remains unclear. As VEN is increasingly used in early stages of CLL, better understanding and tools to circumvent VEN resistance are highly needed. We aim to reveal the metabolic adaption of CLL to CD40 signalling in connection with VEN resistance. After in vitro CD40 signalling stimulation of peripheral blood (PB) CLL cells, mitochondrial mass and glucose uptake were measured by flow cytometry, oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured on Seahorse XF Analyser. The result demonstrated that CD40 stimulation enhances both oxidative phosphorylation (OXPHOS) and glycolysis. This was also confirmed by microarray and metabolomics analyses, as genes and metabolites involved in these two metabolic pathways are significantly upregulated by CD40 stimulation. To find out whether these pathways are linked to VEN resistance, PB CLL cells were treated with OXPHOS or glycolysis inhibitors during CD40 stimulation. Remarkably, OXPHOS inhibition by electron transport chain (ETC) inhibitors (rotenone, antimycin A and oligomycin) counteracted strongly for VEN resistance, while glycolysis inhibition by 2-Deoxy-D-glucose (2DG) did not. The three ETC inhibitors also attenuated CLL activation, ATP production and NAD levels. Interestingly, complex II inhibition of the ETC (TTFA and DMM) did not affect VEN resistance. Regarding BCL-2 family members induced by CD40 ligation, both MCL-1 and BCL-XL were downregulated by these ETC inhibitors. In addition, OXPHOS inhibition strongly elevates glycolysis, and vice versa, which illustrates a strong metabolic plasticity of CLL cells. To further investigate the cross-talk between CD40 signalling, VEN resistance and mitochondrial metabolism, the three main fuels of the TCA cycle were inhibited: pyruvate (by UK5099), glutamine (by DON) and fatty acids (by etomoxir). Even though the OCR and ECAR were slightly decreased by (combinations of) these fuel inhibitors, neither CD40 signalling nor VEN sensitivity was affected. Next, we inhibited PI3K by idelalisib, BTK by ibrutinib and mTOR by rapamycin, which are three downstream targets of CD40 signalling. The results showed that only rapamycin inhibited CD40 activation and metabolic activities, and none of the three inhibitors counteracts VEN resistance. Lastly, we investigated CD40 splicing and overall expression. Interestingly, CD40 stimulation has a huge impact on CD40 expression itself, and these changes were blocked by ETC inhibition. These data indicate that ETC inhibition affects CD40 signals to counteract VEN resistance, by directly affecting the expression of CD40 protein on the cell membrane. In conclusion, after CD40 stimulation, CLL cells become metabolically activated and highly flexible in the use of mitochondrial fuels. The enhanced OXPHOS but not glycolysis contributes to VEN resistance, while ETC inhibition reverses CLL VEN resistance by directly suppressing CD40 expression on CLL. These findings link CLL metabolism directly to CD40 transcription and signalling, which may contribute to clinical VEN resistance. Disclosures van der Windt: genmab: Current Employment. Kater:Abbvie: Research Funding; Roche: Research Funding; Celgene: Research Funding; Janssen: Research Funding; Genentech: Research Funding. Eldering:Genentech: Research Funding; Celgene: Research Funding; Janssen: Research Funding.

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