Iron acquisition by oral hemolytic spirochetes: isolation of a hemin-binding protein and identification of iron reductase activity

1996 ◽  
Vol 42 (10) ◽  
pp. 1072-1079 ◽  
Author(s):  
D. Scott ◽  
R. Siboo ◽  
E. C. S. Chan
Keyword(s):  
Binding Protein ◽  
Reductase Activity ◽  
Iron Acquisition ◽  
Ammonium Citrate ◽  
Ferric Ammonium Citrate ◽  
Affinity Column ◽  
Treponema Denticola ◽  
Apparent Homogeneity ◽  
Iron Reductase

Oral anaerobic spirochetes (OAS) have been implicated in the etiology of periodontal disease. To adapt to the environment of the subgingiva, OAS must be able to acquire iron from limited sources. OAS have previously been shown not to produce siderophores but are β-hemolytic and can bind hemin via a proteinaceous 47-kDa outer membrane sheath (OMS) receptor. Present studies show that [3H]hemin is not transported into the cytoplasm, that hemin and ferric ammonium citrate, as the sole iron sources, can support the growth of OAS and that protoporphyrin IX and Congo red are inhibitory, thereby implying an important in vivo role for hemin as an iron source. Treponema denticola ATCC 35405 produces an iron reductase. The iron reductase can reduce the central ferric iron moiety of hemin. The 47-kDa OMS hemin-binding protein has been purified to apparent homogeneity by methanol–chloroform extraction of cellular lipoproteins and the use of a hemin–agarose bead affinity column. A model of iron acquisition by OAS is presented.Key words: Treponema denticola, hemin-binding protein, iron limitation, iron reductase.

ANTIBACTERIAL ACTION OF A REACTION PRODUCT OF CYSTEINE AND IRON: III. IN VIVO ACTION ON PNEUMOCOCCUS INFECTION IN MICE

1954 ◽  
Vol 1 (3) ◽  
pp. 211-215 ◽  
Author(s):  
N. A. Hinton ◽  
Jack Konowalchuk ◽  
G. B. Reed
Keyword(s):  
Lethal Dose ◽  
Intravenous Dose ◽  
Ammonium Citrate ◽  
Antibacterial Action ◽  
Ferric Ammonium Citrate ◽  
Dilute Solution ◽  
Complete Protection ◽  
Ferric Ammonium ◽  
Colloidal Sulphur

A colloidal sulphur preparation, formed by autoclaving a dilute solution of cysteine and ferric ammonium citrate, was shown to have no toxicity for mice after eleven 1-mgm. intravenous doses on alternate days. Single doses up to 2 mgm. subcutaneously were not toxic but larger doses by this route produced necrosis. A single 1-mgm. dose of the preparation given to mice intravenously afforded no protection against a lethal dose of Diplococcus pneumoniae Type III when given simultaneously. However, groups of mice given a 1-mgm. intravenous dose of the complex and challenged with a lethal dose of pneumococcus at intervals up to 168 hr. after the therapy show no protection for the first 24 hr. following the therapy, increasing protection from 24 to 78 hr., complete protection at 78 hr., and decreasing protection from 78 to 168 hr.

scholarly journals Nonbinding Site-Directed Mutants of Transferrin Binding Protein B Exhibit Enhanced Immunogenicity and Protective Capabilities

2014 ◽  
Vol 83 (3) ◽  
pp. 1030-1038 ◽  
Author(s):  
Rafael Frandoloso ◽  
Sonia Martínez-Martínez ◽  
Charles Calmettes ◽  
Jamie Fegan ◽  
Estela Costa ◽  
...  
Keyword(s):  
Immune Response ◽  
Binding Protein ◽  
Iron Acquisition ◽  
Critical Role ◽  
T Cell Response ◽  
Content Type ◽  
Growth And Survival ◽  
Mutant Proteins ◽  
Protein B

Host-adapted Gram-negative bacterial pathogens from thePasteurellaceae,Neisseriaceae, andMoraxellaceaefamilies normally reside in the upper respiratory or genitourinary tracts of their hosts and rely on utilizing iron from host transferrin (Tf) for growth and survival. The surface receptor proteins that mediate this critical iron acquisition pathway have been proposed as ideal vaccine targets due to the critical role that they play in survival and disease pathogenesisin vivo. In particular, the surface lipoprotein component of the receptor, Tf binding protein B (TbpB), had received considerable attention as a potential antigen for vaccines in humans and food production animals but this has not translated into the series of successful vaccine products originally envisioned. Preliminary immunization experiments suggesting that host Tf could interfere with development of the immune response prompted us to directly address this question with site-directed mutant proteins defective in binding Tf. Site-directed mutants with dramatically reduced binding of porcine transferrin and nearly identical structure to the native proteins were prepared. A mutantHaemophilus parasuisTbpB was shown to induce an enhanced B-cell and T-cell response in pigs relative to native TbpB and provide superior protection from infection than the native TbpB or a commercial vaccine product. The results indicate that binding of host transferrin modulates the development of the immune response against TbpBs and that strategies designed to reduce or eliminate binding can be used to generate superior antigens for vaccines.

Web-Based Bioinformatics Analysis of TbpB from Actinobacillus pleuropneumonie L20 Strain

2012 ◽  
Vol 466-467 ◽  
pp. 256-261
Author(s):  
Ren Feng Li ◽  
Xiang Qin Tian ◽  
Kun Zhao ◽  
Jin Qing Jiang ◽  
San Hu Wang
Keyword(s):  
Membrane Protein ◽  
Bioinformatics Analysis ◽  
Binding Protein ◽  
Iron Acquisition ◽  
Protein A ◽  
Actinobacillus Pleuropneumoniae ◽  
Biological Characterization ◽  
Web Based

The transferring(Tf) receptor from Actinobacillus pleuropneumoniae (App) is comprised of a surface exposed lipoprotein, Tf-binding protein B (TbpB), and an integral outer-membrane protein, Tf-binding protein A (TbpA), both of which are essential for survival in the host, and TbpB is required for the iron acquisition process in vivo. In this study. We analyzed the salient features of the TbpB gene and encoded protein of App L20 strain by bioinformatics tools and highlighted its important biological characterization, which. providing insights into the mechanism of Tf binding and the role of TbpB,

scholarly journals Differences in iron requirements by concanavalin A-treated and anti-CD3-treated murine splenic lymphocytes

2002 ◽  
Vol 88 (1) ◽  
pp. 67-72 ◽  
Author(s):  
Solo R. Kuvibidila ◽  
Maria Velez ◽  
Lolie Yu ◽  
Raj P. Warrier ◽  
B. Surendra Baliga
Keyword(s):  
Dna Synthesis ◽  
T Cell Activation ◽  
Concanavalin A ◽  
Lymphocyte Proliferation ◽  
Reductase Activity ◽  
Interleukin 2 ◽  
Calf Serum ◽  
Ammonium Citrate ◽  
Ferric Ammonium Citrate ◽  
Splenic Lymphocytes

Fe availability is critical for optimal lymphocyte proliferation; however, the minimum required levels are unknown. Such information is valuable when assessingin vitroimmune responses in Fe-deficient subjects, because serum (Fe) added to the culture medium may replete lymphocytes. To address this issue, splenic lymphocytes obtained from seventeen 3-month-old C57BL/6 mice were incubated without and with 1 mg/l concanavalin A or 50 μg/l anti-CD3 antibody in media that contained between 0·113 and 9·74 μmol Fe/l. Fe was provided by either fetal calf serum (FCS, 0–100 ml/l), newborn calf serum (NBCS, 0–100 ml/l), or NBCS (10 ml/l) plus ferric ammonium citrate. As expected, the rate of DNA synthesis increased with Fe levels (P<0·01). Maximum DNA synthesis was obtained with 2·26 μmol Fe/l (50 ml FCS/l) for concanavalin A and 0·895 μmol/l (20 ml FCS/l) for anti-CD3-treated cells. In serum-free media (0·113 μmol Fe/l), the proliferative responses to concanavalin A were below the background, while they rose 5·5-fold in anti-CD3-treated cells (P<0·05). In apotransferrin-supplemented media (0·13 μmol Fe/l), the proliferative responses to concanavalin A and anti-CD3 antibody were 18·6 and 71 %, respectively, of that obtained with 4·66 μmol Fe/l (100 ml FCS/l). Interleukin 2 secretion also followed the same trend as lymphocyte proliferation. Since differences between both mitogens persisted after FCS was substituted with NBCS, we can rule out an effect on ribonucleotide reductase activity, or by other serum growth factors. We speculate an Fe effect at an early step of T-cell activation. Data suggest that the minimum Fe concentration required for lymphocyte proliferation varies with the mitogen.

Iron overload increases hepatic development ofPlasmodium yoeliiin mice

Parasitology ◽  
1996 ◽  
Vol 112 (2) ◽  
pp. 165-168 ◽  
Author(s):  
J. Goma ◽  
L. Rénia ◽  
F. Miltgen ◽  
D. Mazier
Keyword(s):  
Iron Overload ◽  
Malaria Infection ◽  
Iron Supplementation ◽  
Plasmodium Yoelii ◽  
Ammonium Citrate ◽  
Ferric Ammonium Citrate ◽  
Ferric Ammonium ◽  
Hepatic Development

SummaryIron overload in BALB/c mice by treatment with ferric ammonium citrate promotes the hepatic development ofPlasmodium yoelii in vivoand invitro. This was the result of increased penetration of the parasite into hepatocytes since no effect was observed on parasite transformation or maturation. These results could explain why in endemic regions iron supplementation led, in certain studies, to an increase in clinical episodes of malaria and in the prevalence of malaria infection.

Evidence that mammalian ribonucleotide reductase is a nuclear membrane associated glycoprotein

1990 ◽  
Vol 68 (5) ◽  
pp. 880-888 ◽  
Author(s):  
Marianna Sikorska ◽  
Linda M. Brewer ◽  
Tony Youdale ◽  
Robert Richards ◽  
James F. Whitfield ◽  
...  
Keyword(s):  
Rat Liver ◽  
Ribonucleotide Reductase ◽  
Nuclear Membrane ◽  
Mammalian Cells ◽  
Reductase Activity ◽  
Cell Types ◽  
Liver Cells ◽  
Affinity Column ◽  
Rat Liver Cells

Epitope-specific antibodies to the M1 and M2 subunits of mammalian ribonucleotide reductase were prepared using peptides predicted to have a high antigenic index. Western blotting demonstrated that the anti-M1 antibody was specific for the 89-kilodalton M1 subunit (and its degradation fragments) and the anti-M2 antibody specifically recognized the 45-kilodalton M2 subunit. Both antibodies inhibited the CDP-reductase activity of the holoenzyme. Using these antibodies, both the M1 and M2 subunits were shown to be localized in the cytoplasm and in the nuclear regions of a number of cell types, including B77 avian sarcoma virus transformed NRK cells, T51B rat liver cells, 5123tc hepatoma cells, and rat liver cells in vivo. In addition, the M1 subunit was found to be localized as a halo around isolated rat liver nuclei. Biochemical analysis of the cytoplasmic fraction of liver cells and a Triton X-100 wash of nuclei from these cells confirmed the location of the enzyme activity in these cellular compartments. The M1 subunit appears to be glycosylated, as indicated by its retention on a Affi-Gel – concanavalin A affinity column. Therefore, in mammalian cells ribonucleotide reductase appears to be not only in the cytoplasm, but is also associated with the nuclear membrane or nuclear lamina. The activity of the enzyme in the membrane fraction changes dynamically during the cell cycle.Key words: replication, DNA, synthesis, glycosylation, liver.

Promising Anti-stroke Signature of Voglibose: Investigation through In- Silico Molecular Docking and Virtual Screening in In-Vivo Animal Studies

2020 ◽  
Vol 20 (3) ◽  
pp. 223-235
Author(s):  
Pooja Shah ◽  
Vishal Chavda ◽  
Snehal Patel ◽  
Shraddha Bhadada ◽  
Ghulam Md. Ashraf
Keyword(s):  
Molecular Docking ◽  
Virtual Screening ◽  
In Silico ◽  
Reductase Activity ◽  
Infarct Volume ◽  
Docking Studies ◽  
Serum Glucose ◽  
In Vivo Studies ◽  
In Silico Molecular Docking

Background: Postprandial hyperglycemia considered to be a major risk factor for cerebrovascular complications. Objective: The current study was designed to elucidate the beneficial role of voglibose via in-silico in vitro to in-vivo studies in improving the postprandial glycaemic state by protection against strokeprone type 2 diabetes. Material and Methods: In-Silico molecular docking and virtual screening were carried out with the help of iGEMDOCK+ Pymol+docking software and Protein Drug Bank database (PDB). Based on the results of docking studies, in-vivo investigation was carried out for possible neuroprotective action. T2DM was induced by a single injection of streptozotocin (90mg/kg, i.v.) to neonates. Six weeks after induction, voglibose was administered at the dose of 10mg/kg p.o. for two weeks. After eight weeks, diabetic rats were subjected to middle cerebral artery occlusion, and after 72 hours of surgery, neurological deficits were determined. The blood was collected for the determination of serum glucose, CK-MB, LDH and lipid levels. Brains were excised for determination of brain infarct volume, brain hemisphere weight difference, Na+-K+ ATPase activity, ROS parameters, NO levels, and aldose reductase activity. Results: In-silico docking studies showed good docking binding score for stroke associated proteins, which possibly hypotheses neuroprotective action of voglibose in stroke. In the present in-vivo study, pre-treatment with voglibose showed a significant decrease (p<0.05) in serum glucose and lipid levels. Voglibose has shown significant (p<0.05) reduction in neurological score, brain infarct volume, the difference in brain hemisphere weight. On biochemical evaluation, treatment with voglibose produced significant (p<0.05) decrease in CK-MB, LDH, and NO levels in blood and reduction in Na+-K+ ATPase, oxidative stress, and aldose reductase activity in brain homogenate. Conclusion: In-silico molecular docking and virtual screening studies and in-vivo studies in MCAo induced stroke, animal model outcomes support the strong anti-stroke signature for possible neuroprotective therapeutics.

scholarly journals ATF3 contributes to brucine-triggered glioma cell ferroptosis via promotion of hydrogen peroxide and iron

2021 ◽  
Author(s):  
Shan Lu ◽  
Xuan-zhong Wang ◽  
Chuan He ◽  
Lei Wang ◽  
Shi-peng Liang ◽  
...  
Keyword(s):  
Lipid Peroxidation ◽  
Cell Death ◽  
Er Stress ◽  
Glioma Cell ◽  
Nuclear Translocation ◽  
Indole Alkaloid ◽  
Ammonium Citrate ◽  
Ferric Ammonium Citrate ◽  
Activating Transcription Factor 3 ◽  
Intracellular Iron

AbstractFerroptotic cell death is characterized by iron-dependent lipid peroxidation that is initiated by ferrous iron and H2O2 via Fenton reaction, in which the role of activating transcription factor 3 (ATF3) remains elusive. Brucine is a weak alkaline indole alkaloid extracted from the seeds of Strychnos nux-vomica, which has shown potent antitumor activity against various tumors, including glioma. In this study, we showed that brucine inhibited glioma cell growth in vitro and in vivo, which was paralleled by nuclear translocation of ATF3, lipid peroxidation, and increases of iron and H2O2. Furthermore, brucine-induced lipid peroxidation was inhibited or exacerbated when intracellular iron was chelated by deferoxamine (500 μM) or improved by ferric ammonium citrate (500 μM). Suppression of lipid peroxidation with lipophilic antioxidants ferrostatin-1 (50 μM) or liproxstatin-1 (30 μM) rescued brucine-induced glioma cell death. Moreover, knockdown of ATF3 prevented brucine-induced accumulation of iron and H2O2 and glioma cell death. We revealed that brucine induced ATF3 upregulation and translocation into nuclei via activation of ER stress. ATF3 promoted brucine-induced H2O2 accumulation via upregulating NOX4 and SOD1 to generate H2O2 on one hand, and downregulating catalase and xCT to prevent H2O2 degradation on the other hand. H2O2 then contributed to brucine-triggered iron increase and transferrin receptor upregulation, as well as lipid peroxidation. This was further verified by treating glioma cells with exogenous H2O2 alone. Moreover, H2O2 reversely exacerbated brucine-induced ER stress. Taken together, ATF3 contributes to brucine-induced glioma cell ferroptosis via increasing H2O2 and iron.

scholarly journals C4b-binding protein α-chain enhances antitumor immunity by facilitating the accumulation of tumor-infiltrating lymphocytes in the tumor microenvironment in pancreatic cancer

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Kosuke Sasaki ◽  
Shigetsugu Takano ◽  
Satoshi Tomizawa ◽  
Yoji Miyahara ◽  
Katsunori Furukawa ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Microenvironment ◽  
Binding Protein ◽  
Combined Treatment ◽  
Tumor Infiltrating Lymphocytes ◽  
Pdac Cell ◽  
Α Chain ◽  
Infiltrating Lymphocytes

Abstract Background Recent studies indicate that complement plays pivotal roles in promoting or suppressing cancer progression. We have previously identified C4b-binding protein α-chain (C4BPA) as a serum biomarker for the early detection of pancreatic ductal adenocarcinoma (PDAC). However, its mechanism of action remains unclear. Here, we elucidated the functional roles of C4BPA in PDAC cells and the tumor microenvironment. Methods We assessed stromal C4BPA, the C4BPA binding partner CD40, and the number of CD8+ tumor-infiltrating lymphocytes in resected human PDAC tissues via immunohistochemical staining. The biological functions of C4BPA were investigated in peripheral blood mononuclear cells (PBMCs) and human PDAC cell lines. Mouse C4BPA (mC4BPA) peptide, which is composed of 30 amino acids from the C-terminus and binds to CD40, was designed for further in vitro and in vivo experiments. In a preclinical experiment, we assessed the efficacy of gemcitabine plus nab-paclitaxel (GnP), dual immune checkpoint blockades (ICBs), and mC4BPA peptide in a mouse orthotopic transplantation model. Results Immunohistochemical analysis revealed that high stromal C4BPA and CD40 was associated with favorable PDAC prognosis (P=0.0005). Stromal C4BPA strongly correlated with the number of CD8+ tumor-infiltrating lymphocytes (P=0.001). In in vitro experiments, flow cytometry revealed that recombinant human C4BPA (rhC4BPA) stimulation increased CD4+ and CD8+ T cell numbers in PBMCs. rhC4BPA also promoted the proliferation of CD40-expressing PDAC cells. By contrast, combined treatment with gemcitabine and rhC4BPA increased PDAC cell apoptosis rate. mC4BPA peptide increased the number of murine T lymphocytes in vitro and the number of CD8+ tumor-infiltrating lymphocytes surrounding PDAC tumors in vivo. In a preclinical study, GnP/ICBs/mC4BPA peptide treatment, but not GnP treatment, led to the accumulation of a greater number of CD8+ T cells in the periphery of PDAC tumors and to greater tumor regression than did control treatment. Conclusions These findings demonstrate that the combination of GnP therapy with C4BPA inhibits PDAC progression by promoting antitumor T cell accumulation in the tumor microenvironment.

Sign in / Sign up

Export Citation Format

Share Document