scholarly journals Novel Pathway for Arsenic Detoxification in the Legume Symbiont Sinorhizobium meliloti

2005 ◽  
Vol 187 (20) ◽  
pp. 6991-6997 ◽  
Author(s):  
Hung-Chi Yang ◽  
Jiujun Cheng ◽  
Turlough M. Finan ◽  
Barry P. Rosen ◽  
Hiranmoy Bhattacharjee
Keyword(s):  
Sinorhizobium Meliloti ◽  
Physiological Function ◽  
Phosphate Transport ◽  
Transport Systems ◽  
Intact Cells ◽  
Arsenic Resistance ◽  
E Coli ◽  
Arsenic Detoxification ◽  
Selective Sensitivity

ABSTRACT We report a novel pathway for arsenic detoxification in the legume symbiont Sinorhizobium meliloti. Although a majority of ars operons consist of three genes, arsR (transcriptional regulator), arsB [As(OH)3/H+ antiporter], and arsC (arsenate reductase), the S. meliloti ars operon includes an aquaglyceroporin (aqpS) in place of arsB. The presence of AqpS in an arsenic resistance operon is interesting, since aquaglyceroporin channels have previously been shown to adventitiously facilitate uptake of arsenite into cells, rendering them sensitive to arsenite. To understand the role of aqpS in arsenic resistance, S. meliloti aqpS and arsC were disrupted individually. Disruption of aqpS resulted in increased tolerance to arsenite but not arsenate, while cells with an arsC disruption showed selective sensitivity to arsenate. The results of transport experiments in intact cells suggest that AqpS is the only protein of the S. meliloti ars operon that facilitates transport of arsenite. Coexpression of S. meliloti aqpS and arsC in a strain of E. coli lacking the ars operon complemented arsenate but not arsenite sensitivity. These results imply that, when S. meliloti is exposed to environmental arsenate, arsenate enters the cell through phosphate transport systems and is reduced to arsenite by ArsC. Internally generated arsenite flows out of the cell by downhill movement through AqpS. Thus, AqpS confers arsenate resistance together with ArsC-catalyzed reduction. This is the first report of an aquaglyceroporin with a physiological function in arsenic resistance.

Transcriptional regulation of a hybrid cluster (prismane) protein

2005 ◽  
Vol 33 (1) ◽  
pp. 195-197 ◽  
Author(s):  
N.A. Filenko ◽  
D.F. Browning ◽  
J.A. Cole
Keyword(s):  
Physiological Function ◽  
Response Regulator ◽  
Anaerobic Growth ◽  
Hybrid Cluster ◽  
E Coli ◽  
Mutant Strains ◽  
Translation Initiation Codon ◽  
Nitric Oxide Reduction ◽  
Nitrate And Nitrite

HCP (hybrid-cluster protein) contains two Fe/S clusters, one of which is a hybrid [4Fe-2S-2O] cluster. Despite intensive study, its physiological function has not been reported. The Escherichia coli hcp gene is located in a two-gene operon with hcr, which encodes an NADH-dependent HCP reductase. E. coli HCP is detected after anaerobic growth with nitrate or nitrite: possible roles for it in hydroxylamine or nitric oxide reduction have been proposed. To study the regulation and role of HCP, an hcp::lacZ fusion was constructed and transformed into fnr, arcA and norR mutant strains of E. coli. Transcription from the hcp promoter was induced during anaerobic growth. Only the fnr mutant was defective in hcp expression. Nitrate- and nitrite-induced transcription from the hcp promoter was activated by the response regulator proteins NarL and NarP. Gel retardation assays were used to show that FNR (fumarate-nitrate regulation) and NarL form a complex with the hcp promoter. Transcription of the hcp-hcr operon initiates at a thymine nucleotide located 31 bp upstream of the translation-initiation codon. HCP has been overexpressed from a recombinant plasmid for physiological studies.

scholarly journals Biochemical Studies on Active Transport

1968 ◽  
Vol 52 (1) ◽  
pp. 279-295 ◽  
Author(s):  
Arthur B. Pardee
Keyword(s):  
Active Transport ◽  
Cell Function ◽  
Specific Protein ◽  
Transport Systems ◽  
Initial State ◽  
Intact Cells ◽  
The Past ◽  
Biochemical Studies ◽  
Active Transport Process

The active transport process, so important in cell function, has been studied in the past with intact cells. Models which have arisen from this work all depend on: first, a specific protein to recognize the substrate; second, translocation of the substrate across the cell membrane; third, release of substrate within the cell and restoration of the system to its initial state. These steps are adequate for facilitated transport, but in active transport an energy input is required to maintain a concentration gradient. Parts of transport systems have been isolated recently. A protein which specifically recognizes ß-galactosides has been partially purified. In another case, a protein that appears to be the recognition part of the sulfate transport system of Salmonella typhimurium has been crystallized, and many of its properties have been described. The role of this protein in recognition and in translocation is discussed. Also proteins that phosphorylate a variety of sugars as they enter the cell's interior provide a mechanism for concentrating sugars as their phosphates, against a gradient.

scholarly journals Functional Promiscuity of Homologues of the Bacterial ArsA ATPases

2010 ◽  
Vol 2010 ◽  
pp. 1-21 ◽  
Author(s):  
Rostislav Castillo ◽  
Milton H. Saier
Keyword(s):  
Phylogenetic Trees ◽  
Essential Role ◽  
Carbon Starvation ◽  
Arsenic Resistance ◽  
E Coli ◽  
Published Evidence ◽  
Vesicle Biogenesis ◽  
Arsenic Detoxification

The ArsA ATPase ofE. coliplays an essential role in arsenic detoxification. Published evidence implicates ArsA in the energization of As(III) efflux via the formation of an oxyanion-translocating complex with ArsB. In addition, eukaryotic ArsA homologues have several recognized functions unrelated to arsenic resistance. By aligning ArsA homologues, constructing phylogenetic trees, examining ArsA encoding operons, and estimating the probable coevolution of these homologues with putative transporters and auxiliary proteins unrelated to ArsB, we provide evidence for new functions for ArsA homologues. They may play roles in carbon starvation, gas vesicle biogenesis, and arsenic resistance. The results lead to the proposal that ArsA homologues energize four distinct and nonhomologous transporters, ArsB, ArsP, CstA, and Acr3.

The role of Na+ in membrane transport and respiration in the marine bacterium Deleya aesta 134

1991 ◽  
Vol 37 (6) ◽  
pp. 433-439 ◽  
Author(s):  
Marc Berthelet ◽  
Robert A. MacLeod
Keyword(s):  
Membrane Transport ◽  
Maximum Rate ◽  
Marine Bacterium ◽  
Transport Systems ◽  
Intact Cells ◽  
Effective Competition ◽  
Glucose Accumulation ◽  
Quinone Acceptor ◽  
The Relationship

Deleya aesta required Na+ for the uptake of 8 of 11 metabolites tested; the other three were transported at low rates in the absence of Na+ but at much higher rates in its presence. The optimal concentration of Na+ for maximum rate of transport of all the metabolites was 200 to 300 mM. Higher concentrations added as NaCl inhibited transport to the same extent as equiosomolar concentrations of other salts and sucrose. Li+ but not K+ could replace Na+ for the uptake of some metabolites but was only one-half as effective. Competition studies indicated that a number of different transport systems were involved in uptake. Inhibitor studies of succinate, L-alanine, and D-glucose accumulation (the last in both the presence and absence of Na+) suggested that for each a membrane potential was required. The relationship between rates of transport and oxidation of succinate by intact cells at various Na+ concentrations indicated that the Na+ requirement for oxidation reflected the Na+ requirement for transport. For D-glucose, the relationship was more complex, sinced over a narrow, low concentration range, Na+ inhibited respiration but not transport. Evidence for the presence in D. aesta 134 of a Na+-activated NADH–quinone acceptor oxidoreductase was obtained. Key words: Deleya aesta, marine bacteria, sodium, membrane transport, respiration.

Relationship of the Escherichia coli TrkA System of Potassium Ion Uptake with the F0F1-ATPase Under Growth Conditions Without Anaerobic or Aerobic Respiration

1998 ◽  
Vol 18 (3) ◽  
pp. 143-154 ◽  
Author(s):  
Armen Trchounian ◽  
Yelena Ohanjanyan ◽  
Karine Bagramyan ◽  
Vitya Vardanian ◽  
Eleonora Zakharyan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Growth Conditions ◽  
Transport Systems ◽  
Aerobic Respiration ◽  
Anaerobic Conditions ◽  
Intact Cells ◽  
Potassium Ion Uptake ◽  
Relationship Of ◽  
The Relationship

K+ uptake by the Escherichia coli TrkA system is unusual in that it requires both ATP and ΔμH; a relation withH+ circulation through the membrane is thereforesuggested. The relationship of this system with the F0F1-ATPase was studied in intact cells grownunder different conditions. A significant increase of the N,N'-dicyclohexylcarbodiimide(DCCD)-inhibitedH+ efflux through the F0F1 by 5 mMK+, but not by Na+ added into thepotassium-free medium was revealed only in fermenting wild-type orparent cells, that were grown under anaerobic conditions withoutanaerobic or aerobic respiration and with the production of H2. Such an increase disappeared in the Δunc or the trkA mutants that have alteredF0F1 or defective TrkA, respectively. This finding indicates a closed relationship between TrkA andF0F1, with these transport systems beingassociated in a single mechanism that functions as an ATP-driven H+–K+-exchanging pump. ADCCD-inhibited H+–K+-exchangethrough these systems with the fixed stoichiometry of H+and K+ fluxes(2H+/K+) and a higherK+ gradient between the cytoplasm and the externalmedium were also found in these bacteria. They were not observed incells cultured under anaerobic conditions in the presence of nitrate orunder aerobic conditions with respiration and without production of H2. The role of anaerobic or aerobic respiration as adeterminant of the relationship of the TrkA with the F0F1 is postulated. Moreover, an increase of DCCD-inhibited H+ efflux by added K+, aswell as the characteristics of DCCD-sensitiveH+–K+-exchange found in a parentstrain, were lost in the arcA mutant with a defective Arc system, suggesting a repression of enzymes in respiratorypathways. In addition, K+ influx in the latest mutantwas not markedly changed by valinomycin or with temperature. The arcA gene product or the Arc system is proposed to beimplicated in the regulation of the relationship between TrkA and F0F1.

Phospholipid synthesis in Borrelia burgdorferi: BB0249 and BB0721 encode functional phosphatidylcholine synthase and phosphatidylglycerolphosphate synthase proteins

Microbiology ◽  
2004 ◽  
Vol 150 (2) ◽  
pp. 391-397 ◽  
Author(s):  
Xing-Guo Wang ◽  
Joanna P. Scagliotti ◽  
Linden T. Hu
Keyword(s):  
Borrelia Burgdorferi ◽  
Sinorhizobium Meliloti ◽  
Biosynthetic Pathways ◽  
Phospholipid Synthesis ◽  
Membrane Phospholipids ◽  
Bacterial Membranes ◽  
E Coli ◽  
Methylation Pathway ◽  
And Function

Phospholipids are an important component of bacterial membranes. Borrelia burgdorferi differs from many other bacteria in that it contains only two major membrane phospholipids: phosphatidylglycerol (PG) and phosphatidylcholine (PC). B. burgdorferi appears to lack enzymes required for synthesis of PC through the well-described methylation pathway. However, B. burgdorferi does contain a gene (BB0249) with significant identity to a recently described phosphatidylcholine synthase gene (pcs) of Sinorhizobium meliloti. B. burgdorferi also contains a gene (BB0721) with significant identity to the gene (pgs) encoding phosphatidylglycerolphosphate synthase, an enzyme in the synthetic pathway of PG. Activity of BB0249 was confirmed by cloning the gene into Escherichia coli, which does not produce PC. Transformation with a plasmid carrying BB0249 resulted in production of PC by E. coli, but only in the presence of exogenously supplied choline, as would be predicted for a Pcs. Because loss of Pgs activity is lethal to E. coli, activity of BB0721 was confirmed by the ability of BB0721 to complement an E. coli Pgs− mutant. A plasmid containing BB0721 was transformed into a Pgs− mutant of E. coli containing a copy of the native gene on a temperature-regulated plasmid. The temperature-regulated plasmid was exchanged for a plasmid containing BB0721 and it was shown that BB0721 was able to replace the lost Pgs function and restore bacterial growth. This study has established the existence and function of two critical enzymes in the synthesis of PC and PG in B. burgdorferi. Understanding of the biosynthetic pathways of PC and PG in B. burgdorferi is the first step in delineating the role of these phospholipids in the pathogenesis of Lyme disease.

scholarly journals Role of oxyR from Sinorhizobium meliloti in Regulating the Expression of Catalases

2005 ◽  
Vol 37 (6) ◽  
pp. 421-428 ◽  
Author(s):  
Li Luo ◽  
Ming-Sheng Qi ◽  
Shi-Yi Yao ◽  
Hai-Ping Cheng ◽  
Jia-Bi Zhu ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Sinorhizobium Meliloti ◽  
Wild Type Strain ◽  
Reduced Form ◽  
Insertion Mutant ◽  
Transcriptional Level ◽  
Oxygen Species ◽  
E Coli ◽  
Catalase Peroxidase

AbstractThe process of symbiotic nitrogen fixation results in the generation of reactive oxygen species such as the superoxide anion (O2-) and hydrogen peroxide (H2O2). The response of rhizobia to these toxic oxygen species is an important factor in nodulation and nitrogen fixation. In Sinorhizobium meliloti, one oxyR homologue and three catalase genes, katA, katB, and katC were detected by sequence analysis. This oxyR gene is located next to and divergently from katA on the chromosome. To investigate the possible roles of oxyR in regulating the expression of catalases at the transcriptional level in S. meliloti, an insertion mutant of this gene was constructed. The mutant was more sensitive and less adaptive to H2O2 than the wild type strain, and total catalase/peroxidase activity was reduced approximately fourfold with the OxyR mutation relative to controls. The activities of KatA and KatB and the expression of katA::lacZ and katB::lacZ promoter fusions were increased in the mutant strain compared with the parental strain grown in the absence of H2O2, indicating that katA and katB are repressed by OxyR. However, when exposed to H2O2, katA expression was also increased in both S. meliloti and Escherichia coli. When exposed to H2O2, OxyR is converted from a reduced to an oxidized form in E. coli. We concluded that the reduced form of OxyR functions as a repressor of katA and katB expression. Thus, in the presence of H2O2, reduced OxyR is converted to the oxidized form of OxyR that then results in increased katA expression. We further showed that oxyR expression is autoregulated via negative feedback.

scholarly journals The TolC Homologue of Brucella suis Is Involved in Resistance toAntimicrobial Compounds and Virulence

2006 ◽  
Vol 75 (1) ◽  
pp. 379-389 ◽  
Author(s):  
Diana M. Posadas ◽  
Fernando A. Martín ◽  
Julia V. Sabio y García ◽  
Juan M. Spera ◽  
M. Victoria Delpino ◽  
...  
Keyword(s):  
Infection Process ◽  
Transport Systems ◽  
Large Protein ◽  
Gene Encoding ◽  
Toxic Compounds ◽  
E Coli ◽  
Brucella Suis ◽  
Protein Toxins ◽  
Mouse Model Of Infection

ABSTRACT Brucella spp., like other pathogens, must cope with the environment of diverse host niches during the infection process. In doing this, pathogens evolved different type of transport systems to help them survive and disseminate within the host. Members of the TolC family have been shown to be involved in the export of chemically diverse molecules ranging from large protein toxins to small toxic compounds. The role of proteins from the TolC family in Brucella and otherα -2-proteobacteria has been explored little. The gene encoding the unique member of the TolC family from Brucella suis (BepC) was cloned and expressed in an Escherichia coli mutant disrupted in the gene encoding TolC, which has the peculiarity of being involved in diverse transport functions. BepC fully complemented the resistance to drugs such as chloramphenicol and acriflavine but was incapable of restoring hemolysin secretion in the tolC mutant of E. coli. An insertional mutation in the bepC gene strongly affected the resistance phenotype of B. suis to bile salts and toxic chemicals such as ethidium bromide and rhodamine and significantly decreased the resistance to antibiotics such as erythromycin, ampicillin, tetracycline, and norfloxacin. Moreover, the B. suis bepC mutant was attenuated in the mouse model of infection. Taken together, these results suggest that BepC-dependent efflux processes of toxic compounds contribute to B. suis survival inside the host.

Expression of theEscherichia colichromosomalarsoperon

1996 ◽  
Vol 42 (7) ◽  
pp. 662-671 ◽  
Author(s):  
Jie Cai ◽  
Michael S. DuBow
Keyword(s):  
Escherichia Coli ◽  
Northern Blotting ◽  
Open Reading Frames ◽  
Gene Fusions ◽  
Arsenic Resistance ◽  
E Coli ◽  
Ars Operon ◽  
Arsenic Detoxification ◽  
Transcriptional Fusions ◽  
Reading Frames

A chromosomally located operon (ars) of Escherichia coli has been previously shown to be functional in arsenic detoxification. DNA sequencing revealed three open reading frames homologous to the arsR, arsB, and arsC open reading frames of plasmid-based arsenic resistance operons isolated from both E. coli and staphylococcal species. To examine the outline of transcriptional regulation of the chromosomal ars operon, several transcriptional fusions, using the luciferase-encoding luxAB genes of Vibrio harveyi, were constructed. Measurement of the expression of these gene fusions demonstrated that the operon was rapidly induced by sodium arsenite and negatively regulated by the trans-acting arsR gene product. Northern blotting and primer extension analyses revealed that the chromosomal ars operon is most likely transcribed as a single mRNA of approximately 2100 nucleotides in length and processed into two smaller mRNA products in a manner similar to that found in the E. coli R773 plasmid-borne ars operon. However, transcription was found to initiate at a position that is relatively further upstream of the initiation codon of the arsR coding sequence than that determined for the E. coli R773 plasmid's ars operon.Key words: arsenic resistance, Escherichia coli, transcription, gene fusions.

scholarly journals Role of Potassium Uptake Systems in Sinorhizobium meliloti Osmoadaptation and Symbiotic Performance

2009 ◽  
Vol 191 (7) ◽  
pp. 2133-2143 ◽  
Author(s):  
Ana Domínguez-Ferreras ◽  
Socorro Muñoz ◽  
José Olivares ◽  
María J. Soto ◽  
Juan Sanjuán
Keyword(s):  
Sinorhizobium Meliloti ◽  
Growth Conditions ◽  
Potassium Uptake ◽  
Transport Systems ◽  
Osmotic Adaptation ◽  
Low Potassium ◽  
Genes Encoding ◽  
In Silico Analyses ◽  
Important System

ABSTRACT Stimulation of potassium uptake is the most rapid response to an osmotic upshock in bacteria. This cation accumulates by a number of different transport systems whose importance has not been previously addressed for rhizobia. In silico analyses reveal the presence of genes encoding four possible potassium uptake systems in the genome of Sinorhizobium meliloti 1021: Kup1, Kup2, Trk, and Kdp. The study of the relevance of these systems under a number of different growth conditions and in symbiosis showed that the integrity of Kup1 or Trk is essential for growth under laboratory conditions even in osmotically balanced media and the absence of both systems leads to a reduced infectivity and competitiveness of the bacteria in alfalfa roots. Trk is the main system involved in the accumulation of potassium after an osmotic upshift and the most important system for growth of S. meliloti under hyperosmotic conditions. The other three systems, especially Kup1, are also relevant during the osmotic adaptation of the cell, and the relative importance of the Kdp system increases at low potassium concentrations.

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