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Helene F Rosenberg - One of the best experts on this subject based on the ideXlab platform.
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Review Eosinophil-Derived Neurotoxin (EDN/RNase 2) and the Mouse Eosinophil-Associated RNases (mEars): Expanding Roles in Promoting Host Defense
2016Co-Authors: Helene F RosenbergAbstract:Abstract: The Eosinophil-Derived Neurotoxin (EDN/RNase2) and its divergent orthologs, the mouse eosinophil-associated RNases (mEars), are prominent secretory proteins of eosinophilic leukocytes and are all members of the larger family of RNase A-type ribonucleases. While EDN has broad antiviral activity, targeting RNA viruses via mechanisms that may require enzymatic activity, more recent studies have elucidated how these RNases may generate host defense via roles in promoting leukocyte activation, maturation, and chemotaxis. This review provides an update on recent discoveries, and highlights the versatility of this family in promoting innate immunity
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gata transcription factors regulate the expression of the human eosinophil derived Neurotoxin rnase 2 gene
Journal of Biological Chemistry, 2009Co-Authors: Zhijun Qiu, Kimberly D Dyer, Zhihui Xie, Madeleine Radinger, Helene F RosenbergAbstract:The transcription factors GATA-1 and GATA-2 have been implicated in promoting differentiation of eosinophilic leukocytes. In this study, we examined the roles of GATA-1 and GATA-2 in activating transcription of the secretory ribonuclease, the Eosinophil-Derived Neurotoxin (EDN/RNase 2). Augmented expression of both GATA-1 and GATA-2 was detected in eosinophil promyelocyte HL-60 clone 15 cells in response to biochemical differentiation with butyric acid. Deletion or mutation of one or both of the two consensus GATA-binding sites in the extended 1000-bp 5′ promoter of the EDN gene resulted in profound reduction in reporter gene activity. Antibody-augmented electrophoretic mobility shift and chromatin immunoprecipitation analyses indicate that GATA-1 and GATA-2 proteins bind to both functional GATA consensus sequences in the EDN promoter. Interestingly, RNA silencing of GATA-1 alone had no impact on EDN expression; silencing of GATA-2 resulted in diminished expression of EDN, and also diminished expression of GATA-1 in both butyric acid-induced HL-60 clone 15 cells and in differentiating human eosinophils derived from CD34+ hematopoietic progenitors. Likewise, overexpression of GATA-2 in uninduced HL-60 clone 15 cells resulted in augmented transcription of both EDN and GATA-1. Taken together, our data suggest that GATA-2 functions directly via interactions with the EDN promoter and also indirectly, via its ability to regulate the expression of GATA-1 in differentiating eosinophils and eosinophil cell lines.
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eosinophil derived Neurotoxin acts as an alarmin to activate the tlr2 myd88 signal pathway in dendritic cells and enhances th2 immune responses
Journal of Experimental Medicine, 2008Co-Authors: Dexuan Yang, Helene F Rosenberg, Qianqian Chen, Ping Zhang, Kahori Kurosaka, Rachel R Caspi, Suzanne M Michalek, Ning Zhang, Joost J OppenheimAbstract:Eosinophil-Derived Neurotoxin (EDN) is an eosinophil granule–derived secretory protein with ribonuclease and antiviral activity. We have previously shown that EDN can induce the migration and maturation of dendritic cells (DCs). Here, we report that EDN can activate myeloid DCs by triggering the Toll-like receptor (TLR)2–myeloid differentiation factor 88 signaling pathway, thus establishing EDN as an endogenous ligand of TLR2. EDN activates TLR2 independently of TLR1 or TLR6. When mice were immunized with ovalbumin (OVA) together with EDN or with EDN-treated OVA-loaded DCs, EDN enhanced OVA-specific T helper (Th)2-biased immune responses as indicated by predominant production of OVA-specific interleukin (IL)-5, IL-6, IL-10, and IL-13, as well as higher levels of immunoglobulin (Ig)G1 than IgG2a. Based on its ability to serve as a chemoattractant and activator of DCs, as well as the capacity to enhance antigen-specific immune responses, we consider EDN to have the properties of an endogenous alarmin that alerts the adaptive immune system for preferential enhancement of antigen-specific Th2 immune responses.
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proteomic based discovery and characterization of glycosylated eosinophil derived Neurotoxin and cooh terminal osteopontin fragments for ovarian cancer in urine
Clinical Cancer Research, 2006Co-Authors: Steven J Skates, Helene F Rosenberg, Samuel C Mok, Nora Horick, Allison F Vitonis, Dale Edwards, Patrick M Sluss, Won K Han, Ross S Berkowitz, Daniel W CramerAbstract:Purpose: The objective was to identify and characterize low molecular weight proteins/peptides in urine and their posttranslational modifications that might be used as a screening tool for ovarian cancer. Experimental Design: Urine samples collected preoperatively from postmenopausal women with ovarian cancer and benign conditions and from nonsurgical controls were analyzed by surface-enhanced laser desorption/ionization mass spectrometry and two-dimensional gel electrophoresis. Selected proteins from mass profiles were purified by chromatography and followed by liquid chromatography-tandem mass spectrometry sequence analysis. Specific antibodies were generated for further characterization, including immunoprecipitation and glycosylation. Quantitative and semiquantitative ELISAs were developed for preliminary validation in patients of 128 ovarian cancer, 52 benign conditions, 44 other cancers, and 188 healthy controls. Results: A protein ( m/z ∼17,400) with higher peak intensities in cancer patients than in benign conditions and controls was identified and subsequently defined as Eosinophil-Derived Neurotoxin (EDN). A glycosylated form of EDN was specifically elevated in ovarian cancer patients. A cluster of COOH-terminal osteopontin was identified from two-dimensional gels of urine from cancer patients. Modified forms EDN and osteopontin fragments were elevated in early-stage ovarian cancers and a combination of both resulted to 93% specificity and 72% sensitivity. Conclusions: Specific elevated posttranslationally modified urinary EDN and osteopontin COOH-terminal fragments in ovarian cancer might lead to potential noninvasive screening tests for early diagnosis. Urine with less complexity than serum and relatively high thermodynamic stability of peptides or metabolites is a promising study medium for discovery of the novel biomarkers which may present in many nonurinary tract neoplastic diseases.
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human ribonuclease a superfamily members eosinophil derived Neurotoxin and pancreatic ribonuclease induce dendritic cell maturation and activation
Journal of Immunology, 2004Co-Authors: De Yang, Helene F Rosenberg, Qian Chen, Susanna M Rybak, Dianne L Newton, Zhao Yuan Wang, Velizar T Tchernev, Minjuan Wang, Barry Schweitzer, Stephen F KingsmoreAbstract:A number of mammalian antimicrobial proteins produced by neutrophils and cells of epithelial origin have chemotactic and activating effects on host cells, including cells of the immune system. Eosinophil granules contain an antimicrobial protein known as Eosinophil-Derived Neurotoxin (EDN), which belongs to the RNase A superfamily. EDN has antiviral and chemotactic activities in vitro. In this study, we show that EDN, and to a lesser extent human pancreatic RNase (hPR), another RNase A superfamily member, activates human dendritic cells (DCs), leading to the production of a variety of inflammatory cytokines, chemokines, growth factors, and soluble receptors. Human angiogenin, a RNase evolutionarily more distant to EDN and hPR, did not display such activating effects. Additionally, EDN and hPR also induced phenotypic and functional maturation DCs. These RNases were as efficacious as TNF-α, but induced a different set of cytokine mediators. Furthermore, EDN production by human macrophages could be induced by proinflammatory stimuli. The results reveal the DC-activating activity of EDN and hPR and suggest that they are likely participants of inflammatory and immune responses. A number of endogenous mediators in addition to EDN have been reported to have both chemotactic and activating effects on APCs, and can thus amplify innate and Ag-specific immune responses to danger signals. We therefore propose these mediators be considered as endogenous multifunctional immune alarmins.
Hirohito Kita - One of the best experts on this subject based on the ideXlab platform.
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Eosinophil granule proteins in peripheral blood granulocytes
2015Co-Authors: A Abu-ghazaleh I. S, Hirohito Kita, D A Loegering, James L Checkel, Ra L. Dunnette, Larry L. Thomas, Gerald J. GleichtiAbstract:Abstract: Eosinophils contain four principal cationic proteins, major basic protein (MBP), Eosinophil-Derived Neurotoxin (EDN), eosinophil cationic protein (ECP), and eosinophil peroxidase (EPO). To determine the quantities of these proteins in granulocytes and whether they are specific to eosinophils, their concentrations in ly-sates of human granulocytes were measured using specific radioimmunoassays. The effect of different methods for eosinophil lysis on the recovery of the proteins was also studied. Maximal recovery occurred at pH 2 for MBP and pH 5.6 for the other granule proteins. The proteins cosedimented with eosinophils and their concentrations (X ± SEM) in ng/106 eosinophils (and in nM/106 eo
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accuracy safety and tolerability of tissue collection by cytosponge vs endoscopy for evaluation of eosinophilic esophagitis
Clinical Gastroenterology and Hepatology, 2015Co-Authors: David A Katzka, Hirohito Kita, Thomas C Smyrk, Debra M Geno, Anupama Ravi, Pierre Laosirieix, Ahmed Miramedi, Irene Debiram, Maria Odonovan, Gail M KephartAbstract:Background & Aims Management of eosinophilic esophagitis (EoE) requires repeated endoscopic collection of mucosal samples to assess disease activity and response to therapy. An easier and less expensive means of monitoring of EoE is required. We compared the accuracy, safety, and tolerability of sample collection via Cytosponge (an ingestible gelatin capsule comprising compressed mesh attached to a string) with those of endoscopy for assessment of EoE. Methods Esophageal tissues were collected from 20 patients with EoE (all with dysphagia, 15 with stricture, 13 with active EoE) via Cytosponge and then by endoscopy. Number of eosinophils/high-power field and levels of Eosinophil-Derived Neurotoxin were determined; hematoxylin-eosin staining was performed. We compared the adequacy, diagnostic accuracy, safety, and patient preference for sample collection via Cytosponge vs endoscopy procedures. Results All 20 samples collected by Cytosponge were adequate for analysis. By using a cutoff value of 15 eosinophils/high power field, analysis of samples collected by Cytosponge identified 11 of the 13 individuals with active EoE (83%); additional features such as abscesses were also identified. Numbers of eosinophils in samples collected by Cytosponge correlated with those in samples collected by endoscopy (r = 0.50, P = .025). Analysis of tissues collected by Cytosponge identified 4 of the 7 patients without active EoE (57% specificity), as well as 3 cases of active EoE not identified by analysis of endoscopy samples. Including information on level of Eosinophil-Derived Neurotoxin did not increase the accuracy of diagnosis. No complications occurred during the Cytosponge procedure, which was preferred by all patients, compared with endoscopy. Conclusions In a feasibility study, the Cytosponge is a safe and well-tolerated method for collecting near mucosal specimens. Analysis of numbers of eosinophils/high-power field identified patients with active EoE with 83% sensitivity. Larger studies are needed to establish the efficacy and safety of this method of esophageal tissue collection. ClinicalTrials.gov number: NCT01585103.
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Allergen-specific IgG1 and IgG3 through FcRIl Induce
2014Co-Authors: Eosinophil Degranulation, Mark C. Swanson, Masayuki Kaneko, Hirohito KitaAbstract:Evidence suggests that eosinophils contribute to inflammation in bronchial asthma by releasing chemical mediators and cytotoxic granule proteins. To investigate the mechanism of eosinophil degranulation in asthma, we established an in vitro model of allergen-induced degranulation. We treated tissue culture plates with short ragweed pollen (SRW) extract and sera from either normal donors or SRW-sensitive patients with asthma. Eosinophils were incubated in the wells and degranulation was assessed by measurement of Eosinophil-Derived Neurotoxin in supernatants. We detected degranulation only when sera from SRW-sensitive patients were reacted with SRW. Anti-IgG and anti-FcRII mAb, but not anti-IgE or anti-FcRU mAb, abolished the degranulation. IgG-depleted serum did not induce degranulation; IgE-depleted serum triggered as much degranulation as untreated serum. Furthermore, serum levels of SRW-specific IgG1 or IgG3 correlated with the amounts of released Eosinophil-Derived Neurotoxin. When eosinophils were cultured in wells coated with purified IgG or IgE, eosinophil degranulation was observed only with IgG. Finally, human IgGl and IgG3, and less consistently IgG2, but not IgG4, induced degranulation. Thus, sera from patients with SRW-sensitive asthma induce eosinophil degranulation in vitro through antigen-specific IgG1 and IgG3 antibodies. These antibodies may be responsible for degranulation of eosinophils in inflammatory reactions, such as bronchial asthma. (J. Clin. Invest. 1995.95:2813-2821.) Key words: bronchial asthma. eosinophils * immunoglobulin. allergen-degranulatio
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human airway eosinophils respond to chemoattractants with greater eosinophil derived Neurotoxin release adherence to fibronectin and activation of the ras erk pathway when compared with blood eosinophils
Journal of Immunology, 2010Co-Authors: Mary Ellen Bates, Hirohito Kita, Julie B Sedgwick, Yiming Zhu, Lin Ying Liu, Rose G Heuser, Nizar N Jarjour, Paul J BerticsAbstract:Human blood eosinophils exposed ex vivo to hematopoietic cytokines (e.g., IL-5 or GM-CSF) subsequently display enhanced responsiveness to numerous chemoattractants, such as chemokines, platelet-activating factor, or FMLP, through a process known as priming. Airway eosinophils, obtained by bronchoalveolar lavage after segmental Ag challenge, also exhibit enhanced responsiveness to selected chemoattractants, suggesting that they are primed during cell trafficking from the blood to the airway. Earlier work has shown that chemoattractants stimulate greater activation of ERK1 and ERK2 following IL-5 priming in vitro, thus revealing that ERK1/ERK2 activity can be a molecular readout of priming under these circumstances. Because few studies have examined the intracellular mechanisms regulating priming as it relates to human airway eosinophils, we evaluated the responsiveness of blood and airway eosinophils to chemoattractants (FMLP, platelet-activating factor, CCL11, CCL5, CXCL8) with respect to degranulation, adherence to fibronectin, or Ras-ERK signaling cascade activation. When compared with blood eosinophils, airway eosinophils exhibited greater FMLP-stimulated Eosinophil-Derived Neurotoxin release as well as augmented FMLP- and CCL11-stimulated adherence to fibronectin. In airway eosinophils, FMLP, CCL11, and CCL5 stimulated greater activation of Ras or ERK1/ERK2 when compared with baseline. Ras activation by FMLP in blood eosinophils was also enhanced following IL-5 priming. These studies are consistent with a model of in vivo priming of eosinophils by IL-5 or related cytokines following allergen challenge, and further demonstrate the key role of priming in the chemoattractant-stimulated responses of eosinophils. These data also demonstrate the importance of the Ras-ERK signaling pathway in the regulation of eosinophil responses to chemoattractants in the airway. Human airway eosinophils respond to several chemoattractants with increased activation of the Ras-ERK cascade, Eosinophil-Derived Neurotoxin release, and adherence to fibronectin relative to blood eosinophils.
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marked deposition of eosinophil derived Neurotoxin in adult patients with eosinophilic esophagitis
The American Journal of Gastroenterology, 2010Co-Authors: Gail M Kephart, Jeffrey A Alexander, Amindra S Arora, Yvonne Romero, Thomas C Smyrk, Nicholas J Talley, Hirohito KitaAbstract:Marked Deposition of Eosinophil-Derived Neurotoxin in Adult Patients With Eosinophilic Esophagitis
Gerald J Gleich - One of the best experts on this subject based on the ideXlab platform.
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eosinophil ribonucleases and their cutaneous lesion forming activity
Journal of Immunology, 2009Co-Authors: Douglas A Plager, Gerald J Gleich, Mark D.p. Davis, Kristin M. Leiferman, Terry J George, Amy G Andrews, Michael J CoenenAbstract:Eosinophil granule proteins are deposited in cutaneous lesions in many human diseases, but how these proteins contribute to pathophysiology is obscure. We injected eosinophil cationic protein (ECP or RNase 3), Eosinophil-Derived Neurotoxin (EDN or RNase 2), eosinophil peroxidase (EPO), and major basic protein-1 (MBP1) intradermally into guinea pig and rabbit skin. ECP and EDN each induced distinct skin lesions at ≥2.5 μM that began at 2 days, peaking at ∼7 days and persisting up to 6 wk. These lesions were ulcerated (ECP) or crusted (EDN) with marked cellular infiltration. EPO and MBP1 (10 μM) each produced perceptible induration and erythema with moderate cellular infiltration resolving within 2 wk. ECP and EDN localized to dermal cells within 2 days, whereas EPO and MBP1 remained extracellular. Overall, cellular localization and RNase activity of ECP and EDN were critical for lesion formation; differential glycosylation, net cationic charge, or RNase activity alone did not account for lesion formation. Ulcerated lesions from patients with the hypereosinophilic syndrome showed ECP and EDN deposition comparable to that in guinea pig skin. In conclusion, ECP and EDN disrupt skin integrity and cause inflammation. Their presence in ulcerative skin lesions may explain certain findings in human eosinophil-associated diseases.
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eosinophil associated inflammation and elaboration of eosinophil derived proteins in 2 children with raccoon roundworm baylisascaris procyonis encephalitis
Pediatrics, 2001Co-Authors: Christopher L Moertel, Hirohito Kita, Kevin R Kazacos, Joseph H Butterfield, Jan Watterson, Gerald J GleichAbstract:Objective. Eosinophil-associated proteins, especially Eosinophil-Derived Neurotoxin, may be important contributors to the neurologic pathology and symptoms caused by Baylisascaris procyonis infection. Methods. Two cases of severe B procyonis encephalitis with evidence of marked eosinophil degranulation in the central nervous system are presented. Serial cerebrospinal fluid (CSF) specimens were collected from each patient during the course of their illness. Antibodies against B procyonis were measured in the patients9 serum and CSF. Levels of the eosinophilopoietin interleukin-5 (IL-5) and 2 important eosinophil proteins, Eosinophil-Derived Neurotoxin and major basic protein, were assayed in the CSF. Results. Both patients had rapidly progressive central nervous system disease with evidence of eosinophilic meningoencephalitis. Both tested positive for antibodies to B procyonis in serum and CSF and had progressively worsening deep white matter changes on magnetic resonance images of the brain. CSF levels of IL-5, Eosinophil-Derived Neurotoxin, and major basic protein were markedly elevated over controls. Conclusions. This is the first report of the measurement of IL-5, Eosinophil-Derived Neurotoxin, and major basic protein in human CSF. In addition to traumatic damage and necrosis caused by migrating larvae, Eosinophil-Derived Neurotoxin from associated eosinophilic inflammation may be an important contributory factor in the pathogenesis of B procyonis encephalitis. parasite, Eosinophil-Derived-Neurotoxin, major basic protein, eosinophilia, hypereosinophilia, interleukin-5, encephalitis, child.
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indices of airway inflammation in induced sputum reproducibility and validity of cell and fluid phase measurements
American Journal of Respiratory and Critical Care Medicine, 1996Co-Authors: Emilio Pizzichini, Gerald J Gleich, Marcia Margaret Menezes Pizzichini, Ann Efthimiadis, S Evans, Marilyn M Morris, Diane L Squillace, J Dolovich, F E HargreaveAbstract:Methods to examine sputum for indices of airway inflammation are evolving. We have examined the repeatability and the validity of an improved method to measure sputum cells and fluid-phase eosinophil cationic protein (ECP), major basic protein (MBP), Eosinophil-Derived Neurotoxin (EDN), albumin, fibrinogen, tryptase, and interleukin-5 (IL-5). Sputum was induced with hypertonic saline twice within 6 d in 10 healthy subjects, 19 stable asthmatics, and 10 smokers with nonobstructive bronchitis. The method included the processing of freshly expectorated sputum separated from saliva, treatment with a fixed proportion of dithiothreitol 0.1% followed by Dulbecco's phosphate-buffered saline, making cytospins, and collecting the supernatant. The reproducibility of measurements, calculated by the intraclass correlation coefficient, was high for all indices measured with the exception of total cell counts and proportion of lymphocytes. Asthmatics, in comparison with healthy subjects and smokers with bronchitis, had ...
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human eosinophil major basic protein is an endogenous allosteric antagonist at the inhibitory muscarinic m2 receptor
Journal of Clinical Investigation, 1993Co-Authors: David B Jacoby, Gerald J Gleich, A D FryerAbstract:Abstract The effect of human eosinophil major basic protein (MBP) as well as other eosinophil proteins, on binding of [3H]N-methyl-scopolamine ([3H]NMS: 1 x 10(-10) M) to muscarinic M2 receptors in heart membranes and M3 receptors in submandibular gland membranes was studied. MBP inhibited specific binding of [3H]NMS to M2 receptors but not to M3 receptors. MBP also inhibited atropine-induced dissociation of [3H]NMS-receptor complexes in a dose-dependent fashion, demonstrating that the interaction of MBP with the M2 muscarinic receptor is allosteric. This effect of MBP suggests that it may function as an endogenous allosteric inhibitor of agonist binding to the M2 muscarinic receptor. Inhibition of [3H]NMS binding by MBP was reversible by treatment with heparin, which binds and neutralizes MBP. Eosinophil peroxidase (EPO) also inhibited specific binding of [3H]NMS to M2 receptors but not to M3 receptors and inhibited atropine-induced dissociation of [3H]NMS-receptor complexes. On a molar basis, EPO is less potent than MBP. Neither eosinophil cationic protein nor Eosinophil-Derived Neurotoxin affected binding of [3H]NMS to M2 receptors. Thus both MBP and EPO are selective allosteric antagonists at M2 receptors. The effects of these proteins may be important causes of M2 receptor dysfunction and enhanced vagally mediated bronchoconstriction in asthma.
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eosinophil derived Neurotoxin and human liver ribonuclease identity of structure and linkage of neurotoxicity to nuclease activity
Journal of Biological Chemistry, 1992Co-Authors: S Sorrentino, Dohn G Glitz, K J Hamann, D A Loegering, J L Checkel, Gerald J GleichAbstract:Abstract Eosinophil-Derived Neurotoxin (EDN) and human liver RNase were found to be indistinguishable from each other but distinct from the pancreatic ribonucleases in their nucleolytic activity on polynucleotides or small defined substrates. Antibodies to EDN and liver RNase showed identical cross-reactivities in assays of nuclease inhibition and in a radioimmunoassay. In each instance, EDN and liver RNase were easily distinguished from bovine or human pancreatic RNase. When injected intrathecally into rabbits, 5-10 micrograms of EDN or liver RNase each was neurotoxic as judged by induction of the Gordon phenomenon. Human pancreatic RNase was less neurotoxic, and up to 20-fold higher levels of bovine pancreatic RNase showed no effect. Treatment of EDN, liver RNase, and eosinophil cationic protein with iodoacetic acid at pH 5.5 resulted in inactivation of their RNase activity and also destroyed their neurotoxicity. EDN conformation was not greatly affected by iodoacetate treatment since interaction of the modified protein with antibodies was only slightly altered. We conclude that RNase activity is necessary but not sufficient to induce neurotoxic action.
Nicolas Doucet - One of the best experts on this subject based on the ideXlab platform.
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sequence specific backbone resonance assignments and microsecond timescale molecular dynamics simulation of human eosinophil derived Neurotoxin
Biomolecular Nmr Assignments, 2017Co-Authors: Donald Gagne, Chitra Narayanan, Khushboo Bafna, Laurieanne Charest, Pratul K Agarwal, Nicolas DoucetAbstract:Eight active canonical members of the pancreatic-like ribonuclease A (RNase A) superfamily have been identified in human. All structural homologs share similar RNA-degrading functions, while also cumulating other various biological activities in different tissues. The functional homologs Eosinophil-Derived Neurotoxin (EDN, or RNase 2) and eosinophil cationic protein (ECP, or RNase 3) are known to be expressed and secreted by eosinophils in response to infection, and have thus been postulated to play an important role in host defense and inflammatory response. We recently initiated the biophysical and dynamical investigation of several vertebrate RNase homologs and observed that clustering residue dynamics appear to be linked with the phylogeny and biological specificity of several members. Here we report the 1H, 13C and 15N backbone resonance assignments of human EDN (RNase 2) and its molecular dynamics simulation on the microsecond timescale, providing means to pursue this comparative atomic-scale functional and dynamical analysis by NMR and computation over multiple time frames.
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Sequence-specific backbone (1)H, (13)C, and (15)N resonance assignments of human ribonuclease 4.
Biomolecular NMR Assignments, 2015Co-Authors: Donald Gagne, Nicolas DoucetAbstract:Human ribonuclease 4 (RNase 4) is the most evolutionarily conserved member of the 8 canonical human pancreatic-like RNases, showing more than 90 % identity with bovine and porcine homologues. The enzyme displays ribonucleolytic activity with a strong preference for uracil-containing RNA substrates, a feature only shared with human eosinophil derived-Neurotoxin (EDN, or RNase 2) and eosinophil cationic protein (ECP, or RNase 3). It is also the shortest member of the human family, with a significantly truncated C-terminal tail. Its unique active-site pocket and high degree of conservation among vertebrates suggest that the enzyme plays a crucial biological function. Here, we report on the (1)H, (13)C and (15)N backbone resonance assignments of RNase 4, providing means to characterize its molecular function at the atomic level by NMR.
K.r. Acharya - One of the best experts on this subject based on the ideXlab platform.
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Eosinophil-Associated Ribonuclease Inhibitor Design
2015Co-Authors: D E Holloway, M. D. Baker, S. Iyer, K.r. AcharyaAbstract:Eosinophils are leukocytes that store a battery of toxic proteins in cytoplasmic granules. These cells are recruited to tissues in response to parasitic infections, whereupon the toxic proteins are secreted as part of the innate host defense system [1]. The granule proteins include Eosinophil-Derived Neurotoxin (EDN; RNase 2) and eosinophil cationic protein (ECP; RNase 3), two closely-related members of the pancreatic ribonuclease superfamily [2]. ECP possesses potent helminthotoxic and bactericidal activities, while those of EDN are somewhat weaker [3, 4]. Each is an effective agent against ssRNA viruses such as respiratory syncytial virus, possibly mediating the antiviral activity of eosinophils against isolated virions in the respiratory tract [5]. Eosinophil infiltration and degranulation are also characteristic features of allergic inflammation [6]. Markedly elevated levels of EDN and ECP have been found in the inflamed tissues of patients suffering from allegic asthma, rhinitis, inflammatory bowel disease, atopic dermatitis and certain eye diseases [7-11]. These proteins are capable of damaging host tissues. For example, they kill cerebellar Purkinje cells in vitro [12] and produce the neurotoxic Gordon phenomenon when injected into rabbits [13]. ECP is also toxic to guinea pig tracheal epithelium in vitro [14]. It is therefore likely that EDN and ECP are involved in the pathophysiology of the allergic disorder
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molecular recognition of human eosinophil derived Neurotoxin rnase 2 by placental ribonuclease inhibitor
Journal of Molecular Biology, 2005Co-Authors: Shalini Iyer, D E Holloway, Robert Shapiro, K.r. AcharyaAbstract:Placental ribonuclease inhibitor (RI) binds diverse mammalian RNases with dissociation constants that are in the femtomolar range. Previous studies on the complexes of RI with RNase A and angiogenin revealed that RI utilises largely distinctive interactions to achieve high affinity for these two ligands. Here we report a 2.0 A resolution crystal structure of RI in complex with a third ligand, Eosinophil-Derived Neurotoxin (EDN), and a mutational analysis based on this structure. The RI–EDN interface is more extensive than those of the other two complexes and contains a considerably larger set of interactions. Few of the contacts present in the RI–angiogenin complex are replicated; the correspondence to the RI–RNase A complex is somewhat greater, but still modest. The energetic contributions of various interface regions differ strikingly from those in the earlier complexes. These findings provide insight into the structural basis for the unusual combination of high avidity and relaxed stringency that RI displays.
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toward rational design of ribonuclease inhibitors high resolution crystal structure of a ribonuclease a complex with a potent 3 5 pyrophosphate linked dinucleotide inhibitor
Biochemistry, 1999Co-Authors: D D Leonidas, Robert Shapiro, Nello Russo, L I Irons, K.r. AcharyaAbstract:The crystal structure of ribonuclease A (RNase A) in complex with pdUppA-3‘-p [5‘-phospho-2‘-deoxyuridine-3‘-pyrophosphate (P‘→5‘) adenosine 3‘-phosphate] has been determined at 1.7 A resolution. This dinucleotide is the most potent low molecular weight inhibitor of RNase A reported to date (Ki = 27 nM) and is also effective against two major nonpancreatic RNases: Eosinophil-Derived Neurotoxin and RNase-4; in all cases, tight binding in large part derives from the unusual 3‘,5‘-pyrophosphate internucleotide linkage [Russo, N., and Shapiro, R. (1999) J. Biol. Chem. 274, 14902−14908]. The design of pdUppA-3‘-p was based on the crystal structure of RNase A complexed with 5‘-diphosphoadenosine 3‘-phosphate (ppA-3‘-p) [Leonidas, D. D., Shapiro, R., Irons, L. I., Russo, N., and Acharya, K. R. (1997) Biochemistry 36, 5578−5588]. The adenosine of pdUppA-3‘-p adopts an atypical syn conformation not observed for standard adenosine nucleotides bound to RNase A. This conformation, which allows extensive interactions...