Independent Actions of Pertussis Toxin (PTX)
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Pertussis Toxin
Pertussis Toxin Publication Number MAN0004270 Revision Date 09 May 2011 Catalog Number: PHZ1174 Quantity: 50 μg Lot Number: See product label. Appearance: Lyophilized solid. Origin: Bordetella pertussis. Purity: >99%. This preparation migrates as five distinct bands when analyzed by SDS–Urea PAGE. The five bands correspond to one A protomer subunit, designated S1 (Mr=26.2 kDa), and four B oligomer subunits, designated S2- S5 (Mr’s= 21.9, 21.9, 12.1, and 10.9 kDa, respectively). Summary: Islet-activating protein Pertussis toxin consists of an A protomer subunit (S1) which possesses both NAD+ glycohydrolase and ADP–ribosyltransferase activities, and B oligomer subunits (S2, S3, S4, and S5) which are responsible for attachment of the native toxin to eukaryotic cell surfaces. Pertussis toxin uncouples G proteins from receptors by ADP ribosylating a cysteine residue near the carboxyl terminus of the α subunit. Biological Activity: The lowest concentration which produces a clustered growth pattern with CHO cells is 0.03 ng/mL. The adenylate cyclase activity of this preparation is 20.1 picomoles/minute/μg in the presence of 1 μg calmodulin. Reconstitution Reconstitute the contents of this vial with 500 μL sterile, distilled water. The composition of the solution will be Recommendation: 50 μg Pertussis toxin, 10 mM sodium phosphate, pH 7.0, 50 mM sodium chloride. Because Pertussis toxin is relatively insoluble, the resulting suspension should be made uniform by gentle mixing prior to withdrawing aliquots. It is important to note that this suspension should not be sterile-filtered. This preparation is not activated. For use with intact cells or extracts, activation is not necessary. -
What Makes Some Bacterial Toxins So Dangerous? the Most Poisonous Substances Known
Features Poisons and antidotes What makes some bacterial toxins so dangerous? The most poisonous substances known David Moss, Ajit Basak We are used to thinking of proteins as beneficial, so it is surprising to realize that the most toxic sub- and Claire Naylor stances known to man are also protein molecules. These are the bacterial exotoxins, proteins secreted (Birkbeck College, London) by pathogenic bacteria. Toxicity is measured by the median lethal dose (LD50). An LD50 value is defined as the mass of toxin per kg of body weight required to wipe out half of an animal population. Whereas Downloaded from http://portlandpress.com/biochemist/article-pdf/32/4/4/5064/bio032040004.pdf by guest on 02 October 2021 classic poisons, such as potassium cyanide or arsenic trioxide, have LD50 values in the range 5–15 mg/ kg, the causative agent of botulism, botulinum toxin, has an LD50 in the range 1–3 ng/kg, a million times more toxic! Highly pathogenic toxins have a catalytic ing. Treatment has to be repeated every few months. and a binding domain or subunit In the early 19th Century, it was thought that chol- era was caused by ‘bad air’. It was then discovered that Nearly all of the most potent toxins have two compo- cholera was the result of ingesting faecal-contaminated nents, A and B, that are either separate polypeptide water and we now know that the pathogenic agent is an chains or are separate domains. Component A is an 87 kDa toxin secreted by the bacterium Vibrio cholerae3,4. enzyme which modifies an important protein inside the This bacterium is the cause of many deaths in the de- host cell, whereas B is a protein that enables the toxin veloping world in the aftermath of flooding. -
The Role of Streptococcal and Staphylococcal Exotoxins and Proteases in Human Necrotizing Soft Tissue Infections
toxins Review The Role of Streptococcal and Staphylococcal Exotoxins and Proteases in Human Necrotizing Soft Tissue Infections Patience Shumba 1, Srikanth Mairpady Shambat 2 and Nikolai Siemens 1,* 1 Center for Functional Genomics of Microbes, Department of Molecular Genetics and Infection Biology, University of Greifswald, D-17489 Greifswald, Germany; [email protected] 2 Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, CH-8091 Zurich, Switzerland; [email protected] * Correspondence: [email protected]; Tel.: +49-3834-420-5711 Received: 20 May 2019; Accepted: 10 June 2019; Published: 11 June 2019 Abstract: Necrotizing soft tissue infections (NSTIs) are critical clinical conditions characterized by extensive necrosis of any layer of the soft tissue and systemic toxicity. Group A streptococci (GAS) and Staphylococcus aureus are two major pathogens associated with monomicrobial NSTIs. In the tissue environment, both Gram-positive bacteria secrete a variety of molecules, including pore-forming exotoxins, superantigens, and proteases with cytolytic and immunomodulatory functions. The present review summarizes the current knowledge about streptococcal and staphylococcal toxins in NSTIs with a special focus on their contribution to disease progression, tissue pathology, and immune evasion strategies. Keywords: Streptococcus pyogenes; group A streptococcus; Staphylococcus aureus; skin infections; necrotizing soft tissue infections; pore-forming toxins; superantigens; immunomodulatory proteases; immune responses Key Contribution: Group A streptococcal and Staphylococcus aureus toxins manipulate host physiological and immunological responses to promote disease severity and progression. 1. Introduction Necrotizing soft tissue infections (NSTIs) are rare and represent a more severe rapidly progressing form of soft tissue infections that account for significant morbidity and mortality [1]. -
Review Cholera Toxin Structure, Gene Regulation and Pathophysiological
Cell. Mol. Life Sci. 65 (2008) 1347 – 1360 1420-682X/08/091347-14 Cellular and Molecular Life Sciences DOI 10.1007/s00018-008-7496-5 Birkhuser Verlag, Basel, 2008 Review Cholera toxin structure, gene regulation and pathophysiological and immunological aspects J. Sncheza and J. Holmgrenb,* a Facultad de Medicina, UAEM, Av. Universidad 1001, Col. Chamilpa, CP62210 (Mexico) b Department of Microbiology and Immunology and Gothenburg University Vaccine Research Institute (GUVAX), University of Gçteborg, Box 435, Gothenburg, 405 30 (Sweden), e-mail: [email protected] Received 25 October 2007; accepted 12 December 2007 Online First 19 February 2008 Abstract. Many notions regarding the function, struc- have recently been discovered. Regarding the cell ture and regulation of cholera toxin expression have intoxication process, the mode of entry and intra- remained essentially unaltered in the last 15 years. At cellular transport of cholera toxin are becoming the same time, recent findings have generated addi- clearer. In the immunological field, the strong oral tional perspectives. For example, the cholera toxin immunogenicity of the non-toxic B subunit of cholera genes are now known to be carried by a non-lytic toxin (CTB) has been exploited in the development of bacteriophage, a previously unsuspected condition. a now widely licensed oral cholera vaccine. Addition- Understanding of how the expression of cholera toxin ally, CTB has been shown to induce tolerance against genes is controlled by the bacterium at the molecular co-administered (linked) foreign antigens in some level has advanced significantly and relationships with autoimmune and allergic diseases. cell-density-associated (quorum-sensing) responses Keywords. -
How Do Pathogenic Microorganisms Develop Cross-Kingdom Host Jumps? Peter Van Baarlen1, Alex Van Belkum2, Richard C
Molecular mechanisms of pathogenicity: how do pathogenic microorganisms develop cross-kingdom host jumps? Peter van Baarlen1, Alex van Belkum2, Richard C. Summerbell3, Pedro W. Crous3 & Bart P.H.J. Thomma1 1Laboratory of Phytopathology, Wageningen University, Wageningen, The Netherlands; 2Department of Medical Microbiology and Infectious Diseases, Erasmus MC, University Medical Centre Rotterdam, Rotterdam, The Netherlands; and 3CBS Fungal Biodiversity Centre, Utrecht, The Netherlands Correspondence: Bart P.H.J. Thomma, Abstract Downloaded from https://academic.oup.com/femsre/article/31/3/239/2367343 by guest on 27 September 2021 Laboratory of Phytopathology, Wageningen University, Binnenhaven 5, 6709 PD It is common knowledge that pathogenic viruses can change hosts, with avian Wageningen, The Netherlands. Tel.: 10031 influenza, the HIV, and the causal agent of variant Creutzfeldt–Jacob encephalitis 317 484536; fax: 10031 317 483412; as well-known examples. Less well known, however, is that host jumps also occur e-mail: [email protected] with more complex pathogenic microorganisms such as bacteria and fungi. In extreme cases, these host jumps even cross kingdom of life barriers. A number of Received 3 July 2006; revised 22 December requirements need to be met to enable a microorganism to cross such kingdom 2006; accepted 23 December 2006. barriers. Potential cross-kingdom pathogenic microorganisms must be able to First published online 26 February 2007. come into close and frequent contact with potential hosts, and must be able to overcome or evade host defences. Reproduction on, in, or near the new host will DOI:10.1111/j.1574-6976.2007.00065.x ensure the transmission or release of successful genotypes. -
Identi®Cation and Role of Adenylyl Cyclase in Auxin Signalling in Higher
letters to nature + + + P.P. thank the Academy of Finland and the Deutsche Forschungsgemeinschaft, respectively, for ®nancial CO , 53), 77 (C6H5 , 60), 73 (TMSi , 84); 6-methyl-4-hydroxy-2-pyrone: RRt 0.35, 198 (M+, 18), 183 ([M-Me]+, 16), 170 ([M-CO]+, 54), 155 ([M-CO-Me]+, support. + + Correspondence and requests for materials should be addressed to J.S. (e-mail: [email protected] 15), 139 ([M-Me-CO2] , 10), 127 ([M-Me-2CO] , 13), 99 (12), 84 (13), 73 + + freiburg.de). (TMSi , 100), 43 (CH3CO , 55). The numbers show m/z values, and the key fragments and their relative intensities are indicated in parentheses. Received 4 August; accepted 14 October 1998. erratum 1. Helariutta, Y. et al. Chalcone synthase-like genes active during corolla development are differentially expressed and encode enzymes with different catalytic properties in Gerbera hybrida (Asteraceae). Plant Mol. Biol. 28, 47±60 (1995). 2. Helariutta, Y. et al. Duplication and functional divergence in the chalcone synthase gene family of 8 Asteraceae: evolution with substrate change and catalytic simpli®cation. Proc. Natl Acad. Sci. USA 93, Crystal structure of the complex 9033±9038 (1996). 3. Thaisrivongs, S. et al. Structure-based design of HIV protease inhibitors: 5,6-dihydro-4-hydroxy-2- of the cyclin D-dependent pyrones as effective, nonpeptidic inhibitors. J. Med. Chem. 39, 4630±4642 (1996). 4. Hagen, S. E. et al. Synthesis of 5,6-dihydro-4-hydroxy-2-pyrones as HIV-1 protease inhibitors: the kinase Cdk6 bound to the profound effect of polarity on antiviral activity. J. Med. Chem. -
Biological Toxins Fact Sheet
Work with FACT SHEET Biological Toxins The University of Utah Institutional Biosafety Committee (IBC) reviews registrations for work with, possession of, use of, and transfer of acute biological toxins (mammalian LD50 <100 µg/kg body weight) or toxins that fall under the Federal Select Agent Guidelines, as well as the organisms, both natural and recombinant, which produce these toxins Toxins Requiring IBC Registration Laboratory Practices Guidelines for working with biological toxins can be found The following toxins require registration with the IBC. The list in Appendix I of the Biosafety in Microbiological and is not comprehensive. Any toxin with an LD50 greater than 100 µg/kg body weight, or on the select agent list requires Biomedical Laboratories registration. Principal investigators should confirm whether or (http://www.cdc.gov/biosafety/publications/bmbl5/i not the toxins they propose to work with require IBC ndex.htm). These are summarized below. registration by contacting the OEHS Biosafety Officer at [email protected] or 801-581-6590. Routine operations with dilute toxin solutions are Abrin conducted using Biosafety Level 2 (BSL2) practices and Aflatoxin these must be detailed in the IBC protocol and will be Bacillus anthracis edema factor verified during the inspection by OEHS staff prior to IBC Bacillus anthracis lethal toxin Botulinum neurotoxins approval. BSL2 Inspection checklists can be found here Brevetoxin (http://oehs.utah.edu/research-safety/biosafety/ Cholera toxin biosafety-laboratory-audits). All personnel working with Clostridium difficile toxin biological toxins or accessing a toxin laboratory must be Clostridium perfringens toxins Conotoxins trained in the theory and practice of the toxins to be used, Dendrotoxin (DTX) with special emphasis on the nature of the hazards Diacetoxyscirpenol (DAS) associated with laboratory operations and should be Diphtheria toxin familiar with the signs and symptoms of toxin exposure. -
Predicting Coupling Probabilities of G-Protein Coupled Receptors Gurdeep Singh1,2,†, Asuka Inoue3,*,†, J
Published online 30 May 2019 Nucleic Acids Research, 2019, Vol. 47, Web Server issue W395–W401 doi: 10.1093/nar/gkz392 PRECOG: PREdicting COupling probabilities of G-protein coupled receptors Gurdeep Singh1,2,†, Asuka Inoue3,*,†, J. Silvio Gutkind4, Robert B. Russell1,2,* and Francesco Raimondi1,2,* 1CellNetworks, Bioquant, Heidelberg University, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany, 2Biochemie Zentrum Heidelberg (BZH), Heidelberg University, Im Neuenheimer Feld 328, 69120 Heidelberg, Germany, 3Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Miyagi 980-8578, Japan and 4Department of Pharmacology and Moores Cancer Center, University of California, San Diego, La Jolla, CA 92093, USA Received February 10, 2019; Revised April 13, 2019; Editorial Decision April 24, 2019; Accepted May 01, 2019 ABSTRACT great use in tinkering with signalling pathways in living sys- tems (5). G-protein coupled receptors (GPCRs) control multi- Ligand binding to GPCRs induces conformational ple physiological states by transducing a multitude changes that lead to binding and activation of G-proteins of extracellular stimuli into the cell via coupling to situated on the inner cell membrane. Most of mammalian intra-cellular heterotrimeric G-proteins. Deciphering GPCRs couple with more than one G-protein giving each which G-proteins couple to each of the hundreds receptor a distinct coupling profile (6) and thus specific of GPCRs present in a typical eukaryotic organism downstream cellular responses. Determining these coupling is therefore critical to understand signalling. Here, profiles is critical to understand GPCR biology and phar- we present PRECOG (precog.russelllab.org): a web- macology. Despite decades of research and hundreds of ob- server for predicting GPCR coupling, which allows served interactions, coupling information is still missing for users to: (i) predict coupling probabilities for GPCRs many receptors and sequence determinants of coupling- specificity are still largely unknown. -
Reporting the Implementation of the Three Rs in European Primate and Mouse Research Papers: Are We Making Progress?
ATLA 38, 495–517, 2010 495 Reporting the Implementation of the Three Rs in European Primate and Mouse Research Papers: Are We Making Progress? Katy Taylor British Union for the Abolition of Vivisection, London, UK Summary — It is now more than 20 years since both Council of Europe Convention ETS123 and EU Directive 86/609?EEC were introduced, to promote the implementation of the Three Rs in animal experi- mentation and to provide guidance on animal housing and care. It might therefore be expected that reports of the implementation of the Three Rs in animal research papers would have increased during this period. In order to test this hypothesis, a literature survey of animal-based research was conducted. A ran- domly-selected sample from 16 high-profile medical journals, of original research papers arising from European institutions that featured experiments which involved either mice or primates, were identified for the years 1986 and 2006 (Total sample = 250 papers). Each paper was scored out of 10 for the incidence of reporting on the implementation of Three Rs-related factors corresponding to Replacement (justification of non-use of non-animal methods), Reduction (statistical analysis of the number of animals needed) and Refinement (housing aspects, i.e. increased cage size, social housing, enrichment of cage environment and food; and procedural aspects, i.e. the use of anaesthesia, analgesia, humane endpoints, and training for procedures with positive reinforcement). There was no significant increase in overall reporting score over time, for either mouse or primate research. By 2006, mouse research papers scored an average of 0 out of a possible 10, and primate research papers scored an average of 1.5. -
Targeted Neuronal Cell Ablation in the Drosophila Embryo: Pathfinding by Follower Growth Cones in the Absence of Pioneers
Neuron, Vol. 14, 707-715, April, 1995, Copyright© 1995 by Cell Press Targeted Neuronal Cell Ablation in the Drosophila Embryo: Pathfinding by Follower Growth Cones in the Absence of Pioneers David M. Lin,* Vanessa J. Auld,*t reach their targets in the CNS (Wigglesworth, 1953). The and Corey S. Goodman ability of the initial axons to guide later growth cones has Howard Hughes Medical Institute led to the suggestion that pioneering growth cones might Division of Neurobiology be endowed with special pathfinding abilities. Do subse- Department of Molecular and Cell Biology quent growth cones possess the same pathfinding abili- Life Science Addition, Room 519 ties, or are they different from the pioneers? University of California, Berkeley Ablation experiments in a variety of organisms thus far Berkeley, California 94720 have provided a range of conflicting answers to this ques- tion. In the grasshopper embryo CNS, the G growth cone turns anteriorly along the P axons in the NP fascicle, a Summary pathway formed by the three descending P and the two ascending A axons. Although the A and P axons normally We developed a rapid method that uses diphtheria fasciculate and follow one another, either can pioneer the toxin, the flp recognition target sequences, and the complete pathway on its own. Furthermore, any one of GAL4-UAS activation system, to ablate specific neu- the P axons will suffice, since when any two of them are rons in the Drosophila embryo and to examine the con- ablated the remaining P pioneers the pathway. When the sequences in large numbers of embryos at many time three P axons are ablated, the G growth cone stalls, and points. -
G-Protein ␥-Complex Is Crucial for Efficient Signal Amplification in Vision
The Journal of Neuroscience, June 1, 2011 • 31(22):8067–8077 • 8067 Cellular/Molecular G-Protein ␥-Complex Is Crucial for Efficient Signal Amplification in Vision Alexander V. Kolesnikov,1 Loryn Rikimaru,2 Anne K. Hennig,1 Peter D. Lukasiewicz,1 Steven J. Fliesler,4,5,6,7 Victor I. Govardovskii,8 Vladimir J. Kefalov,1 and Oleg G. Kisselev2,3 1Department of Ophthalmology and Visual Sciences, Washington University School of Medicine, St. Louis, Missouri 63110, Departments of 2Ophthalmology and 3Biochemistry and Molecular Biology, Saint Louis University School of Medicine, Saint Louis, Missouri 63104, 4Research Service, Veterans Administration Western New York Healthcare System, and Departments of 5Ophthalmology (Ross Eye Institute) and 6Biochemistry, University at Buffalo/The State University of New York (SUNY), and 7SUNY Eye Institute, Buffalo, New York 14215, and 8Sechenov Institute for Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, Saint Petersburg 194223, Russia A fundamental question of cell signaling biology is how faint external signals produce robust physiological responses. One universal mechanism relies on signal amplification via intracellular cascades mediated by heterotrimeric G-proteins. This high amplification system allows retinal rod photoreceptors to detect single photons of light. Although much is now known about the role of the ␣-subunit of the rod-specific G-protein transducin in phototransduction, the physiological function of the auxiliary ␥-complex in this process remains a mystery. Here, we show that elimination of the transducin ␥-subunit drastically reduces signal amplification in intact mouse rods. The consequence is a striking decline in rod visual sensitivity and severe impairment of nocturnal vision. Our findings demonstrate that transducin ␥-complex controls signal amplification of the rod phototransduction cascade and is critical for the ability of rod photoreceptors to function in low light conditions. -
Supplementary Materials
Supplementary Materials COMPARATIVE ANALYSIS OF THE TRANSCRIPTOME, PROTEOME AND miRNA PROFILE OF KUPFFER CELLS AND MONOCYTES Andrey Elchaninov1,3*, Anastasiya Lokhonina1,3, Maria Nikitina2, Polina Vishnyakova1,3, Andrey Makarov1, Irina Arutyunyan1, Anastasiya Poltavets1, Evgeniya Kananykhina2, Sergey Kovalchuk4, Evgeny Karpulevich5,6, Galina Bolshakova2, Gennady Sukhikh1, Timur Fatkhudinov2,3 1 Laboratory of Regenerative Medicine, National Medical Research Center for Obstetrics, Gynecology and Perinatology Named after Academician V.I. Kulakov of Ministry of Healthcare of Russian Federation, Moscow, Russia 2 Laboratory of Growth and Development, Scientific Research Institute of Human Morphology, Moscow, Russia 3 Histology Department, Medical Institute, Peoples' Friendship University of Russia, Moscow, Russia 4 Laboratory of Bioinformatic methods for Combinatorial Chemistry and Biology, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, Moscow, Russia 5 Information Systems Department, Ivannikov Institute for System Programming of the Russian Academy of Sciences, Moscow, Russia 6 Genome Engineering Laboratory, Moscow Institute of Physics and Technology, Dolgoprudny, Moscow Region, Russia Figure S1. Flow cytometry analysis of unsorted blood sample. Representative forward, side scattering and histogram are shown. The proportions of negative cells were determined in relation to the isotype controls. The percentages of positive cells are indicated. The blue curve corresponds to the isotype control. Figure S2. Flow cytometry analysis of unsorted liver stromal cells. Representative forward, side scattering and histogram are shown. The proportions of negative cells were determined in relation to the isotype controls. The percentages of positive cells are indicated. The blue curve corresponds to the isotype control. Figure S3. MiRNAs expression analysis in monocytes and Kupffer cells. Full-length of heatmaps are presented.