scholarly journals Dinitrogenase-Driven Photobiological Hydrogen Production Combats Oxidative Stress in Cyanothece sp. Strain ATCC 51142

2016 ◽  
Vol 82 (24) ◽  
pp. 7227-7235 ◽  
Author(s):  
Natalie C. Sadler ◽  
Hans C. Bernstein ◽  
Matthew R. Melnicki ◽  
Moiz A. Charania ◽  
Eric A. Hill ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Production ◽  
System Level ◽  
Oxygenic Photosynthesis ◽  
Hydrogenase Activity ◽  
Enzyme Complex ◽  
Strain Atcc ◽  
Content Type ◽  
Thiol Redox

ABSTRACTPhotobiologically synthesized hydrogen (H2) gas is carbon neutral to produce and clean to combust, making it an ideal biofuel.Cyanothecesp. strain ATCC 51142 is a cyanobacterium capable of performing simultaneous oxygenic photosynthesis and H2production, a highly perplexing phenomenon because H2evolving enzymes are O2sensitive. We employed a system-levelin vivochemoproteomic profiling approach to explore the cellular dynamics of protein thiol redox and how thiol redox mediates the function of the dinitrogenase NifHDK, an enzyme complex capable of aerobic hydrogenase activity. We found that NifHDK responds to intracellular redox conditions and may act as an emergency electron valve to prevent harmful reactive oxygen species formation in concert with other cell strategies for maintaining redox homeostasis. These results provide new insight into cellular redox dynamics useful for advancing photolytic bioenergy technology and reveal a new understanding for the biological function of NifHDK.IMPORTANCEHere, we demonstrate that high levels of hydrogen synthesis can be induced as a protection mechanism against oxidative stress via the dinitrogenase enzyme complex inCyanothecesp. strain ATCC 51142. This is a previously unknown feature of cyanobacterial dinitrogenase, and we anticipate that it may represent a strategy to exploit cyanobacteria for efficient and scalable hydrogen production. We utilized a chemoproteomic approach to capture thein situdynamics of reductant partitioning within the cell, revealing proteins and reactive thiols that may be involved in redox sensing and signaling. Additionally, this method is widely applicable across biological systems to achieve a greater understanding of how cells navigate their environment and how redox chemistry can be utilized to alter metabolism and achieve homeostasis.

Hydrogen Evolution Catalyzed by Hydrogenase in Cultures of Cyanobacteria

1981 ◽  
Vol 36 (1-2) ◽  
pp. 87-92 ◽  
Author(s):  
Patrick C. Hallenbeck ◽  
Leon V. Kochian ◽  
John R. Benemann
Keyword(s):  
Hydrogen Production ◽  
Hydrogen Evolution ◽  
High Frequency ◽  
Nitrogenase Activity ◽  
Inhibitory Effect ◽  
Hydrogenase Activity ◽  
Oxygen Inhibition ◽  
Oxygen Sensitive

Abstract Cultures of Anabaena cylindrica, grown on media containing 5 mᴍ NH4Cl (which represses heterocyst formation), evolved hydrogen after a period of dark incubation under an argon atmosphere. This hydrogen production was not due to nitrogenase activity, which was nearly undetectable, but was due to a hydrogenase. Cultures grown on media with tungsten substituted for molybdenum had a high frequency of heterocysts (15%) and inactive nitrogenase after nitrogen starvation. The hydrogenase activity of these cultures was three-fold greater than the activity of non-heterocystous cultures. The effects of oxygen inhibition on hydrogen evolution by hetero-cystous cultures suggest that two pools of hydrogenase activity exist - an oxygen sensitive hydrogen evolution in vegetative cells and a relatively oxygen-resistent hydrogen evolution in heterocysts. In either case, inhibition by oxygen was reversible. Light had an inhibitory effect on net hydrogen evolution. Hydrogen production in vitro was much higher than in vivo, indicating that in vivo hydrogenase activity is limited by endogenous reductant supply.

scholarly journals In Vivo Fitness Adaptations of Colistin-Resistant Acinetobacter baumannii Isolates to Oxidative Stress

2016 ◽  
Vol 61 (3) ◽  
Author(s):  
Crystal L. Jones ◽  
Shweta S. Singh ◽  
Yonas Alamneh ◽  
Leila G. Casella ◽  
Robert K. Ernst ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Acinetobacter Baumannii ◽  
Catalase Activity ◽  
Content Type ◽  
Increased Susceptibility

ABSTRACT The loss of fitness in colistin-resistant (CR) Acinetobacter baumannii was investigated using longitudinal isolates from the same patient. Early CR isolates were outcompeted by late CR isolates for growth in broth and survival in the lungs of mice. Fitness loss was associated with an increased susceptibility to oxidative stress since early CR strains had reduced in vitro survival in the presence of hydrogen peroxide and decreased catalase activity compared to that of late CR and colistin-susceptible (CS) strains.

scholarly journals Sigma S-Dependent Antioxidant Defense Protects Stationary-Phase Escherichia coli against the Bactericidal Antibiotic Gentamicin

2014 ◽  
Vol 58 (10) ◽  
pp. 5964-5975 ◽  
Author(s):  
Jing-Hung Wang ◽  
Rachna Singh ◽  
Michael Benoit ◽  
Mimi Keyhan ◽  
Matthew Sylvester ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Stationary Phase ◽  
Antioxidant Defense ◽  
Physiological State ◽  
Pentose Phosphate ◽  
Phase Cell ◽  
Content Type

ABSTRACTStationary-phase bacteria are important in disease. The σs-regulated general stress response helps them become resistant to disinfectants, but the role of σsin bacterial antibiotic resistance has not been elucidated. Loss of σsrendered stationary-phaseEscherichia colimore sensitive to the bactericidal antibiotic gentamicin (Gm), and proteomic analysis suggested involvement of a weakened antioxidant defense. Use of the psfiAgenetic reporter, 3′-(p-hydroxyphenyl) fluorescein (HPF) dye, and Amplex Red showed that Gm generated more reactive oxygen species (ROS) in the mutant. HPF measurements can be distorted by cell elongation, but Gm did not affect stationary-phase cell dimensions. Coadministration of the antioxidantN-acetyl cysteine (NAC) decreased drug lethality particularly in the mutant, as did Gm treatment under anaerobic conditions that prevent ROS formation. Greater oxidative stress, due to insufficient quenching of endogenous ROS and/or respiration-linked electron leakage, therefore contributed to the greater sensitivity of the mutant; infection by a uropathogenic strain in mice showed this to be the case alsoin vivo. Disruption of antioxidant defense by eliminating the quencher proteins, SodA/SodB and KatE/SodA, or the pentose phosphate pathway proteins, Zwf/Gnd and TalA, which provide NADPH for ROS decomposition, also generated greater oxidative stress and killing by Gm. Thus, besides its established mode of action, Gm also kills stationary-phase bacteria by generating oxidative stress, and targeting the antioxidant defense ofE. colican enhance its efficacy. Relevant aspects of the current controversy on the role of ROS in killing by bactericidal drugs of exponential-phase bacteria, which represent a different physiological state, are discussed.

scholarly journals Evidence that Oxidative Stress InducesspxA2Transcription in Bacillus anthracis Sterne through a Mechanism Requiring SpxA1 and Positive Autoregulation

2016 ◽  
Vol 198 (21) ◽  
pp. 2902-2913 ◽  
Author(s):  
Skye Barendt ◽  
Cierra Birch ◽  
Lea Mbengi ◽  
Peter Zuber
Keyword(s):  
Oxidative Stress ◽  
Bacillus Anthracis ◽  
Transcription Initiation ◽  
Bacterial Species ◽  
Negative Control ◽  
Mobility Shift ◽  
Content Type ◽  
Oxidative Protein Damage

ABSTRACTBacillus anthracispossesses two paralogs of the transcriptional regulator, Spx. SpxA1 and SpxA2 interact with RNA polymerase (RNAP) to activate the transcription of genes implicated in the prevention and alleviation of oxidative protein damage. ThespxA2gene is highly upregulated in infected macrophages, but how this is achieved is unknown. Previous studies have shown that thespxA2gene was under negative control by the Rrf2 family repressor protein, SaiR, whose activity is sensitive to oxidative stress. These studies also suggested thatspxA2was under positive autoregulation. In the present study, we show byin vivoandin vitroanalyses thatspxA2is under direct autoregulation but is also dependent on the SpxA1 paralogous protein. The deletion of eitherspxA1orspxA2reduced the diamide-inducible expression of anspxA2-lacZconstruct.In vitrotranscription reactions using purifiedB. anthracisRNAP showed that SpxA1 and SpxA2 protein stimulates transcription from a DNA fragment containing thespxA2promoter. Ectopically positionedspxA2-lacZfusion requires both SpxA1 and SpxA2 for expression, but the requirement for SpxA1 is partially overcome whensaiRis deleted. Electrophoretic mobility shift assays showed that SpxA1 and SpxA2 enhance the affinity of RNAP forspxA2promoter DNA and that this activity is sensitive to reductant. We hypothesize that the previously observed upregulation ofspxA2in the oxidative environment of the macrophage is at least partly due to SpxA1-mediated SaiR repressor inactivation and the positive autoregulation ofspxA2transcription.IMPORTANCERegulators of transcription initiation are known to govern the expression of genes required for virulence in pathogenic bacterial species. Members of the Spx family of transcription factors function in control of genes required for virulence and viability in low-GC Gram-positive bacteria. InBacillus anthracis, thespxA2gene is highly induced in infected macrophages, which suggests an important role in the control of virulence gene expression during the anthrax disease state. We provide evidence that elevated concentrations of oxidized, active SpxA2 result from an autoregulatory positive-feedback loop drivingspxA2transcription.

scholarly journals Quantifying changes in the thiol redox proteome upon oxidative stress in vivo

2008 ◽  
Vol 105 (24) ◽  
pp. 8197-8202 ◽  
Author(s):  
L. I. Leichert ◽  
F. Gehrke ◽  
H. V. Gudiseva ◽  
T. Blackwell ◽  
M. Ilbert ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Thiol Redox

scholarly journals Bacterioferritin ofMagnetospirillum gryphiswaldenseIs a Heterotetraeicosameric Complex Composed of Functionally Distinct Subunits but Is Not Involved in Magnetite Biomineralization

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
René Uebe ◽  
Frederik Ahrens ◽  
Jörg Stang ◽  
Katharina Jäger ◽  
Lars H. Böttger ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Magnetotactic Bacteria ◽  
Functional Differentiation ◽  
Spectroscopic Techniques ◽  
Content Type ◽  
Magnetospirillum Gryphiswaldense ◽  
Ferroxidase Activity ◽  
Magnetite Biomineralization ◽  
Field Lines

ABSTRACTThe biomineralization pathway of magnetite in magnetotactic bacteria is still poorly understood and a matter of intense debates. In particular, the existence, nature, and location of possible mineral precursors of magnetite are not clear. One possible precursor has been suggested to be ferritin-bound ferrihydrite. To clarify its role for magnetite biomineralization, we analyzed and characterized ferritin-like proteins from the magnetotactic alphaproteobacteriumMagnetospirillum gryphiswaldenseMSR-1, employing genetic, biochemical, and spectroscopic techniques. Transmission Mössbauer spectroscopy of the wild type (WT) and a bacterioferritin (bfr) deletion strain uncovered that the presence of ferrihydrite in cells is coupled to the presence of Bfr. However,bfranddpsdeletion mutants, encoding another ferritin-like protein, or even mutants with their codeletion had no impact on magnetite formation in MSR-1. Thus, ferritin-like proteins are not involved in magnetite biomineralization and Bfr-bound ferrihydrite is not a precursor of magnetite biosynthesis. Using transmission electron microscopy and bacterial two-hybrid and electrophoretic methods, we also show that MSR-1 Bfr is an atypical representative of the Bfr subfamily, as it forms tetraeicosameric complexes from two distinct subunits. Furthermore, our analyses revealed that these subunits are functionally divergent, with Bfr1 harboring a ferroxidase activity while only Bfr2 contributes to heme binding. Because of this functional differentiation and the poor formation of homooligomeric Bfr1 complexes, only heterooligomeric Bfr protects cells from oxidative stressin vivo.In summary, our results not only provide novel insights into the biomineralization of magnetite but also reveal the unique properties of so-far-uncharacterized heterooligomeric bacterioferritins.IMPORTANCEMagnetotactic bacteria likeMagnetospirillum gryphiswaldenseare able to orient along magnetic field lines due to the intracellular formation of magnetite nanoparticles. Biomineralization of magnetite has been suggested to require a yet-unknown ferritin-like ferrihydrite component. Here, we report the identification of a bacterioferritin as the source of ferrihydrite inM. gryphiswaldenseand show that, contrary to previous reports, bacterioferritin is not involved in magnetite biomineralization but required for oxidative stress resistance. Additionally, we show that bacterioferritin ofM. gryphiswaldenseis an unusual member of the bacterioferritin subfamily as it is composed of two functionally distinct subunits. Thus, our findings extend our understanding of the bacterioferritin subfamily and also solve a longstanding question about the magnetite biomineralization pathway.

scholarly journals Dps and DpsL Mediate SurvivalIn VitroandIn Vivoduring the Prolonged Oxidative Stress Response in Bacteroides fragilis

2015 ◽  
Vol 197 (20) ◽  
pp. 3329-3338 ◽  
Author(s):  
Michael I. Betteken ◽  
Edson R. Rocha ◽  
C. Jeffrey Smith
Keyword(s):  
Oxidative Stress ◽  
Stress Response ◽  
Peritoneal Cavity ◽  
Bacteroides Fragilis ◽  
Oxidative Stress Response ◽  
Iron Storage ◽  
Content Type ◽  
Iron Storage Proteins

ABSTRACTBacteroides fragilisis a Gram-negative anaerobe and member of the human intestinal tract microbiome, where it plays many beneficial roles. However, translocation of the organism to the peritoneal cavity can lead to peritonitis, intra-abdominal abscess formation, bacteremia, and sepsis. During translocation,B. fragilisis exposed to increased oxidative stress from the oxygenated tissues of the peritoneal cavity and the immune response. In order to survive,B. fragilismounts a robust oxidative stress response consisting of an acute and a prolonged oxidative stress (POST) response. This report demonstrates that the ability to induce high levels of resistance totert-butyl hydroperoxide (tBOOH) after extended exposure to air can be linked to the POST response. Disk diffusion assays comparing the wild type to a Δdpsmutant and a ΔdpsΔbfrmutant showed greater sensitivity of the mutants to tBOOH after exposure to air, suggesting that Dps and DpsL play a role in the resistance phenotype. Complementation studies withdpsorbfr(encoding DpsL) restored tBOOH resistance, suggesting a role for both of these ferritin-family proteins in the response. Additionally, cultures treated with the iron chelator dipyridyl were not killed by tBOOH, indicating Dps and DpsL function by sequestering iron to prevent cellular damage. Anin vivoanimal model showed that the ΔdpsΔbfrmutant was attenuated, indicating that management of iron is important for survival within the abscess. Together, these data demonstrate a role for Dps and DpsL in the POST response which mediates survivalin vitroandin vivo.IMPORTANCEB. fragilisis the anaerobe most frequently isolated from extraintestinal opportunistic infections, but there is a paucity of information about the factors that allow this organism to survive outside its normal intestinal environment. This report demonstrates that the iron storage proteins Dps and DpsL protect against oxidative stress and that they contribute to survival bothin vitroandin vivo. Additionally, this work demonstrates an important role for the POST response inB. fragilissurvival and provides insight into the complex regulation of this response.

scholarly journals Deletion ofsll1541inSynechocystissp. Strain PCC 6803 Allows Formation of a Far-Red-Shiftedholo-ProteorhodopsinIn Vivo

2018 ◽  
Vol 84 (9) ◽  
Author(s):  
Que Chen ◽  
Jeroen B. van der Steen ◽  
Jos C. Arents ◽  
Aloysius F. Hartog ◽  
Srividya Ganapathy ◽  
...  
Keyword(s):  
Proton Pump ◽  
Essential Gene ◽  
Critical Role ◽  
Action Spectrum ◽  
Atp Synthesis ◽  
Chemical Oxidation ◽  
Oxygenic Photosynthesis ◽  
Content Type ◽  
Pcc 6803

ABSTRACTIn many pro- and eukaryotes, a retinal-based proton pump equips the cell to drive ATP synthesis with (sun)light. Such pumps, therefore, have been proposed as a plug-in for cyanobacteria to artificially increase the efficiency of oxygenic photosynthesis. However, little information on the metabolism of retinal, their chromophore, is available for these organisms. We have studied thein vivoroles of five genes (sll1541,slr1648,slr0091,slr1192, andslr0574) potentially involved in retinal metabolism inSynechocystissp. strain PCC 6803. With a gene deletion approach, we have shown thatSynechocystis apo-carotenoid-15,15-oxygenase (SynACO), encoded by genesll1541, is an indispensable enzyme for retinal synthesis inSynechocystis, presumably via asymmetric cleavage of β-apo-carotenal. The second carotenoid oxygenase (SynDiox2), encoded by geneslr1648, competes with SynACO for substrate(s) but only measurably contributes to retinal biosynthesis in stationary phase via an as-yet-unknown mechanism.In vivodegradation of retinal may proceed through spontaneous chemical oxidation and via enzyme-catalyzed processes. Deletion of geneslr0574(encoding CYP120A1), but not ofslr0091or ofslr1192, causes an increase (relative to the level in wild-typeSynechocystis) in the retinal content in both the linear and stationary growth phases. These results suggest that CYP120A1 does contribute to retinal degradation. Preliminary data obtained using13C-labeled retinal suggest that conversion to retinol and retinoic acid and subsequent further oxidation also play a role. Deletion ofsll1541leads to deficiency in retinal synthesis and allows thein vivoreconstitution of far-red-absorbingholo-proteorhodopsin with exogenous retinal analogues, as demonstrated here for all-trans3,4-dehydroretinal and 3-methylamino-16-nor-1,2,3,4-didehydroretinal.IMPORTANCERetinal is formed by many cyanobacteria and has a critical role in most forms of life for processes such as photoreception, growth, and stress survival. However, the metabolic pathways in cyanobacteria for synthesis and degradation of retinal are poorly understood. In this paper we identify genes involved in its synthesis, characterize their role, and provide an initial characterization of the pathway of its degradation. This led to the identification ofsll1541(encoding SynACO) as the essential gene for retinal synthesis. Multiple pathways for retinal degradation presumably exist. These results have allowed us to construct a strain that expresses a light-dependent proton pump with an action spectrum extending beyond 700 nm. The availability of this strain will be important for further work aimed at increasing the overall efficiency of oxygenic photosynthesis.

scholarly journals Phosphatidylinositol-Specific Phospholipase C Contributes to Survival of Staphylococcus aureus USA300 in Human Blood and Neutrophils

2014 ◽  
Vol 82 (4) ◽  
pp. 1559-1571 ◽  
Author(s):  
Mark J. White ◽  
Jeffrey M. Boyd ◽  
Alexander R. Horswill ◽  
William M. Nauseef
Keyword(s):  
Oxidative Stress ◽  
Staphylococcus Aureus ◽  
Phospholipase C ◽  
Whole Blood ◽  
Dependent Manner ◽  
Binding Studies ◽  
Content Type ◽  
Positive Regulators

ABSTRACTStaphylococcus aureusis an important human pathogen that employs a large repertoire of secreted virulence factors to promote disease pathogenesis. Many strains ofS. aureuspossess aplcgene that encodes a phosphatidylinositol (PI)-specific phospholipase C (PI-PLC) capable of hydrolyzing PI and cleaving glycosyl-PI (GPI)-linked proteins from cell surfaces. Despite being secreted by virulent staphylococci, the contribution of PI-PLC to the capacity ofS. aureusto cause disease remains undefined. Our goal in these studies was to understand PI-PLC in the context ofS. aureusbiology. Among a collection of genetically diverse clinical isolates ofS. aureus, community-associated methicillin-resistantS. aureus(CA-MRSA) USA300 secreted the most PI-PLC. Screening a collection of two-component system (TCS) mutants ofS. aureus, we identified both theagrquorum-sensing system and the SrrAB TCS to be positive regulators ofplcgene expression. Real-time PCR and PI-PLC enzyme assays of the TCS mutants, coupled with SrrA promoter binding studies, demonstrated that SrrAB was the predominant transcriptional activator ofplc. Furthermore,plcregulation was linked to oxidative stress bothin vitroandin vivoin a SrrAB-dependent manner. A Δplcmutant in a CA-MRSA USA300 background exhibited a survival defect in human whole blood and in isolated neutrophils. However, the same mutant strain displayed no survival defect in murine models of infection or murine whole blood. Overall, these data identify potential links between bacterial responses to the host innate immune system and to oxidative stress and suggest how PI-PLC could contribute to the pathogenesis ofS. aureusinfections.

scholarly journals Alkyl Hydroperoxide Reductase Is Required for Helicobacter cinaedi Intestinal Colonization and Survival under Oxidative Stress in BALB/c and BALB/c Interleukin-10−/−Mice

2011 ◽  
Vol 80 (3) ◽  
pp. 921-928 ◽  
Author(s):  
Nisanart Charoenlap ◽  
Zeli Shen ◽  
Megan E. McBee ◽  
Suresh Muthupalani ◽  
Gerald N. Wogan ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Serum Levels ◽  
Interleukin 10 ◽  
Intestinal Bacterium ◽  
Bacterial Survival ◽  
Alkyl Hydroperoxide Reductase ◽  
Content Type ◽  
Helicobacter Cinaedi ◽  
Alkyl Hydroperoxide

Helicobacter cinaedi, a common human intestinal bacterium, has been implicated in various enteric and systemic diseases in normal and immunocompromised patients. Protection against oxidative stress is a crucial component of bacterium-host interactions. Alkyl hydroperoxide reductase C (AhpC) is an enzyme responsible for detoxification of peroxides and is important in protection from peroxide-induced stress.H. cinaedipossesses a singleahpC, which was investigated with respect to its role in bacterial survival during oxidative stress. TheH. cinaedi ahpCmutant had diminished resistance to organic hydroperoxide toxicity but increased hydrogen peroxide resistance compared with the wild-type (WT) strain. The mutant also exhibited an oxygen-sensitive phenotype and was more susceptible to killing by macrophages than the WT strain.In vivoexperiments in BALB/c and BALB/c interleukin-10 (IL-10)−/−mice revealed that the cecal colonizing ability of theahpCmutant was significantly reduced. The mutant also had diminished ability to induce bacterium-specific immune responsesin vivo, as shown by immunoglobulin (IgG2a and IgG1) serum levels. Collectively, these data suggest thatH. cinaedi ahpCnot only contributes to protecting the organism against oxidative stress but also alters its pathogenic propertiesin vivo.

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