DOCSLIB.ORG

Endogenously Bound Ligands Crystal Structure of Bovine Cd1b3 With

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Endogenously Bound Ligands Crystal Structure of Bovine Cd1b3 With Crystal Structure of Bovine CD1b3 with Endogenously Bound Ligands Enrico Girardi, Jing Wang, Thien-Thi Mac, Cees Versluis, Veemal Bhowruth, Gurdyal Besra, Albert J.R. Heck, Ildiko This information is current as Van Rhijn and Dirk M. Zajonc of October 1, 2021. J Immunol published online 2 June 2010 http://www.jimmunol.org/content/early/2010/06/02/jimmun ol.1000042 Downloaded from Supplementary http://www.jimmunol.org/content/suppl/2010/06/02/jimmunol.100004 Material 2.DC1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average by guest on October 1, 2021 Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published June 2, 2010, doi:10.4049/jimmunol.1000042 The Journal of Immunology Crystal Structure of Bovine CD1b3 with Endogenously Bound Ligands Enrico Girardi,* Jing Wang,* Thien-Thi Mac,* Cees Versluis,†,‡ Veemal Bhowruth,x Gurdyal Besra,x Albert J.R. Heck,†,‡ Ildiko Van Rhijn,{,‖ and Dirk M. Zajonc* The CD1 family of Ag-presenting molecules is able to display lipids to T cells by binding them within a hydrophobic groove con- nected to the protein surface. In particular, the CD1b isotype is capable of binding ligands with greatly varying alkyl chain lengths through a complex network of interconnected hydrophobic pockets. Interestingly, mycobacterial lipids such as glucose monomy- colate exclusively bind to CD1b. We determined the crystal structure of one of the three expressed bovine CD1b proteins, CD1b3, in complex with endogenous ligands, identified by mass spectrometry as a mixture of phosphatidylcholine and phosphatidylethanol- amine, and analyzed the ability of the protein to bind glycolipids in vitro. The structure reveals a complex binding groove archi- tecture, similar to the human ortholog but with consequential differences. Intriguingly, in bovine CD1b3 only the A’, C’ and F’ Downloaded from pockets are present, whereas the T’ pocket previously described in human CD1b is closed. This different pocket conformation could affect the ability of boCD1b3 to recognize lipids with long acyl chains such as glucose monomycolate. However, even in the absence of a T’ tunnel, bovine CD1b3 is able to bind mycolates from Rhodococcus ruber in vitro. The Journal of Immunology, 2010, 185: 000–000. uberculosis, the disease caused in humans by infection not directly encoded by genes, and the low level of polymorphism with Mycobacterium tuberculosis, represents a major exhibited by the CD1 family of Ag-presenting molecules that dis- http://www.jimmunol.org/ T cause of mortality worldwide, with increasing reports of play these Ags to T cells (3). During mycobacterial infection in multidrug-resistant strains appearing in several countries (1). In humans and animals, CD1-restricted and glycolipid-specific T cell addition, bovine tuberculosis, caused by Mycobacterium bovis, responses can be detected ex vivo, suggesting a physiologically is of economic and zoonotic concern. Unfortunately, the attenuated relevant role for this mode of Ag presentation. Despite the fact that bacillus Calmette-Guerin vaccine currently available is of little pro- all these features would make CD1-presented lipids excellent vac- tective efficacy in humans and animals (2), highlighting the need of cine subunit candidates, the CD1 system is often seen as static, not alternative approaches for the development of an effective vaccine subject to evolutionary pressure exerted by the changing environ- against tuberculosis. In recent years, mycobacterial glycolipid and mental challenges of an evolving species, and often classified as by guest on October 1, 2021 lipid Ags have been the focus of much attention in this context part of the innate immune system. because of the T cell stimulatory activity of these compounds, the CD1 molecules, similar to MHC class I molecules, exist on the stable chemical structure of the lipid backbone, because lipids are surface of APCs as noncovalent heterodimers formed by a CD1 H chain and a b2-microglobulin (b2m) L chain (4). Whereas the b2m chain consists of a single Ig-like domain, the CD1 H chain *Division of Cell Biology, La Jolla Institute for Allergy and Immunology, La Jolla, CA; †Biomolecular Mass Spectrometry and Proteomics Group, Bijvoet Center for Biomo- is anchored to the membrane via a single-pass transmembrane lecular Research and Utrecht Institute for Pharmaceutical Sciences and {Department of domain and contains a short intracellular cytoplasmic tail and Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht Univer- sity; ‡Netherlands Proteomics Centre, Utrecht, The Netherlands; xSchool of Biosciences, three extracellular domains (denominated a1, a2anda3). The College of Life and Environmental Sciences, University of Birmingham, Edgbaston, C-terminal cytoplasmic tail usually contains trafficking motifs ‖ Birmingham, United Kingdom; and Division of Rheumatology, Immunology and Al- responsible for the intracellular localization of the protein (5). lergy, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA 02115 The a1anda2 ectodomains form the Ag-binding groove respon- Received for publication January 8, 2010. Accepted for publication April 16, 2010. sible for the recognition of self and nonself Ags, whereas the This work was supported in part by Grant AI 074952 from the National Institutes of a3 domain associates noncovalently with b2m. The CD1 binding Health (to D.M.Z.). D.M.Z. is the recipient of an investigator award from the Cancer Research Institute. G.S.B. acknowledges support in the form of a Personal Research groove is highly hydrophobic and defined by two a helices that Chair from Mr. James Badrick, Royal Society Wolfson Research Merit Award, as sit above an antiparallel b-sheet platform. Ags are generally a former Lister Institute-Jenner Research Fellow, the Medical Council and The Well- come Trust (084923/B/08/7). I.V.R. is supported by a Meervoud subsidy of the bound with the alkyl chains buried in the deep hydrophobic Nederlandse Organisatie voor Wetenschappelijk Onderzoek. groove, whereas the glycosidic or polar portion of the molecule Structure factors and coordinates presented in this article have been deposited into is exposed at the surface for recognition by a TCR (4). However, the Protein Data Bank database with code 3L9R. while the overall architecture of these Ag-presenting molecules Address correspondence and reprint requests to Dirk M. Zajonc, La Jolla Institute for is conserved, the size, shape, and number of individual pockets Allergy and Immunology, 9420 Athena Circle, La Jolla CA, 92037. E-mail address: differ between the various CD1 isotypes. [email protected] In humans, all group 1 CD1 proteins (CD1a, CD1b, and CD1c) The online version of this article contains supplemental material. are highly expressed on immature thymocytes and immature and Abbreviations used in this paper: b2m, b2-microglobulin; bo, bovine; ESI, electrospray ionization; GMM, glucose monomycolate; huCD1b, human CD1b; mature dendritic cells, whereas group 2 CD1 (CD1d) is expressed IEF, isoelectric focusing; MS, mass spectrometry; PDB, Protein Data Bank; PC, at lower levels on many cell types (3). A fifth CD1 molecule, phosphatidylcholine; PE, phosphatidylethanolamine; pK, negative logarithm of the CD1e, is not expressed on the cell surface but is found to be in- dissociation constant K. volved in facilitating lipid loading onto CD1b (6). CD1d is the Copyright Ó 2010 by The American Association of Immunologists, Inc. 0022-1767/10/$16.00 restricting element for NKT cells, a cell population with a limited www.jimmunol.org/cgi/doi/10.4049/jimmunol.1000042 2 STRUCTURAL BASIS OF LIPID-Ag RECOGNITION IN CATTLE T cell repertoire that can quickly release large amounts of IFN-g for mouse CD1d (24). The protein expressed in insect cells was purified by and other cytokines (7). Group 1 CD1 proteins are known for their Nickel affinity chromatography (HisTrap FF; GE Healthcare, Piscataway, ability to present lipid Ags to T cells with a diverse T cell repertoire NJ) and eluted with a linear gradient of 20–250 mM imidazole, followed by anion-exchange chromatography on a MonoQ 5/50 column (GE Health- (8, 9). Each of the five human CD1 isotypes possesses typical care) equilibrated with 10 mM Tris-HCl (pH 8.0) and elution using a linear characteristics in terms of size and shape of the Ag-binding groove gradient of 0–350 mM NaCl. The fractions containing the boCD1b3-b2m and its subcellular location. This finding has led to the general idea heterodimer were pooled together and further purified by gel filtra- that human group 1 CD1 represents all possible options to present tion on a Superdex200 10/300 column (GE Healthcare) equilibrated in 50 mM HEPES (pH 7.5), 150 mM NaCl. The fractions containing the an optimal variety of lipid Ags to T cells; CD1b, with the largest complex were then concentrated to 10 mg/ml in 10 mM HEPES Ag-binding groove, is the only isotype that can bind mycolates and (pH 7.5), 30 mM NaCl for subsequent crystallization and analysis. uses lysosomal factors to present exogenous Ags; CD1a has the smallest groove and functions without intersecting with the lyso- Mass spectrometry analysis of boCD1b3 some, whereas CD1c is intermediate in both regards (10, 11). Be- The intact noncovalent complex of boCD1b3 with the lipids was analyzed cause all CD1 molecules except CD1d have been deleted in muroid with native nano electrospray ionization (ESI) mass spectrometry (MS) rodents, the human family of CD1 proteins is often considered to be using borosilicate capillaries to introduce the sample.
Load more

Recommended publications
  • Investigation of Antibiotic Targets in the Decaprenyl
    INVESTIGATION OF ANTIBIOTIC TARGETS IN THE DECAPRENYL-PHOSPHORYLARABINOSE BIOSYNTHESIS IN M. TUBERCULOSIS by SZILVIA TÓTH a thesis submitted to University of Birmingham for the degree of DOCTOR OF PHILOSOPHY School of Biosciences College of Life and Environmental Sciences University of Birmingham July 2018 University of Birmingham Research Archive e-theses repository This unpublished thesis/dissertation is copyright of the author and/or third parties. The intellectual property rights of the author or third parties in respect of this work are as defined by The Copyright Designs and Patents Act 1988 or as modified by any successor legislation. Any use made of information contained in this thesis/dissertation must be in accordance with that legislation and must be properly acknowledged. Further distribution or reproduction in any format is prohibited without the permission of the copyright holder. Declaration The work presented in this thesis is original except where citing relevant references. Studies were conducted from 2014 to 2017 in the School of Biosciences, University of Birmingham, B15 2TT, UK and also in research and development facility of GlaxoSmithKline, laboratory for Diseases of the Developing World, Tres Cantos, Spain. No part of this work has been submitted for a degree or a diploma to this or any other university. ii Abstract An estimated 1.67 million people died of tuberculosis (TB) in 2016 and it is a threat to human life on a global-scale. To shorten current treatments and battle drug resistant strains it is important to discover and develop new drugs against the causative agent, Mycobacterium tuberculosis. Phenotypic screens have delivered potent hit and lead molecules in the past but the need to target new pathways in M.
    [Show full text]
  • Cell Activation Dictates Localization in Vivo the Mechanism of Splenic Invariant
    The Mechanism of Splenic Invariant NKT Cell Activation Dictates Localization In Vivo Irah L. King, Eyal Amiel, Mike Tighe, Katja Mohrs, Natacha Veerapen, Gurdyal Besra, Markus Mohrs and This information is current as Elizabeth A. Leadbetter of September 27, 2021. J Immunol 2013; 191:572-582; Prepublished online 19 June 2013; doi: 10.4049/jimmunol.1300299 http://www.jimmunol.org/content/191/2/572 Downloaded from Supplementary http://www.jimmunol.org/content/suppl/2013/06/17/jimmunol.130029 Material 9.DC1 http://www.jimmunol.org/ References This article cites 46 articles, 16 of which you can access for free at: http://www.jimmunol.org/content/191/2/572.full#ref-list-1 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision by guest on September 27, 2021 • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2013 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology The Mechanism of Splenic Invariant NKT Cell Activation Dictates Localization In Vivo Irah L.
    [Show full text]
  • Wellcome Investigators March 2011
    Wellcome Trust Investigator Awards Feedback from Expert Review Group members 28 March 2011 1 Roughly 7 months between application and final outcome The Expert Review Groups 1. Cell and Developmental Biology 2. Cellular and Molecular Neuroscience (Zaf Bashir) 3. Cognitive Neuroscience and Mental Health 4. Genetics, Genomics and Population Research (George Davey Smith) 5. Immune Systems in Health and Disease (David Wraith) 6. Molecular Basis of Cell Function 7. Pathogen Biology and Disease Transmission 8. Physiology in Health and Disease (Paul Martin) 9. Population and Public Health (Rona Campbell) 2 Summary Feedback from ERG Panels • The bar is very high across all nine panels • Track record led - CV must demonstrate a substantial impact of your research (e.g. high impact journals, record of ground breaking research, clear upward trajectory etc) To paraphrase Walport ‘to support scientists with the best track records, obviously appropriate to the stage in their career’ • Notable esteem factors (but note ‘several FRSs were not shortlisted’) • Your novelty of your research vision is CRUCIAL. Don’t just carry on doing more of the same • The Trust is not averse to risk (but what about ERG panel members?) • Success rate for short-listing for interview is ~15-25% at Senior Investigator level (3-5 proposals shortlisted from each ERG) • Fewer New Investigator than Senior Investigator applications – an opportunity? • There are fewer applications overall for the second round, but ‘the bar will not be lowered’ The Challenge UoB has roughly 45 existing
    [Show full text]
  • The Molecular Bases of / T Cell–Mediated
    Article The molecular bases of / T cell–mediated antigen recognition Daniel G. Pellicci,1,2* Adam P. Uldrich,1,2* Jérôme Le Nours,3,4 Fiona Ross,1,2 Eric Chabrol,3 Sidonia B.G. Eckle,1 Renate de Boer,5 Ricky T. Lim,1 Kirsty McPherson,1 Gurdyal Besra,6 Amy R. Howell,7 Lorenzo Moretta,8 James McCluskey,1 Mirjam H.M. Heemskerk,5 Stephanie Gras,3,4 Jamie Rossjohn,3,4,9** and Dale I. Godfrey1,2** 1Department of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity and 2Australian Research Council Centre of Excellence in Advanced Molecular Imaging, University of Melbourne, Parkville, Victoria 3010, Australia 3Department of Biochemistry and Molecular Biology, School of Biomedical Sciences and 4Australian Research Council Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, Victoria 3800, Australia 5Department of Hematology, Leiden University Medical Center, 2300 RC Leiden, Netherlands 6School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, England, UK 7Department of Chemistry, University of Connecticut, Storrs, CT 06269 8Istituto Giannina Gaslini, 16147 Genova, Italy 9Institute of Infection and Immunity, School of Medicine, Cardiff University, Heath Park, Cardiff CF14 4XN, Wales, UK and T cells are disparate T cell lineages that can respond to distinct antigens (Ags) via the use of the and T cell Ag receptors (TCRs), respectively. Here we characterize a population of human T cells, which we term / T cells, expressing TCRs comprised of a TCR- variable gene (V1) fused to joining and constant domains, paired with an array of TCR- chains. We demonstrate that these cells, which represent 50% of all V1+ human T cells, can recognize peptide- and lipid-based Ags presented by human leukocyte antigen (HLA) and CD1d, respectively.
    [Show full text]
  • 1A.Jan2011-1
    “...scholarships for researchand education...” Newsletter January 2011 Arctic climate change 1750 to 1850 Changing Of whales Planetary codes and men atmospheres No malice…. and Ancient numeracy According to recent findings, one adult in five in England today is the consequences innumerate (“Wecan’t allow people to say ‘I’m no good at maths’”, Arough but possible division of research styles is Guardian 27/4/2010). The figure is particularly stunning when compared to adult illiteracy rates, which are closer to 1%. As the that between the impact-driven and the opportunity- article remarks, for aperson to admit to illiteracy carries social driven. In the first, we tackle recognised objectives stigma, whereas many people will not have aproblem in proclaiming such as the elimination of aparticular disease or the that they cannot count, do not have ahead for figures –that they are establishment of areliable renewable energy source; ‘no good at maths’. Numeracy and literacy,both parts of basic the target is known, it is widely recognised and education, are notably asymmetrical in terms of prestige, and of the progress towards it can be sensibly measured. In the status attached to possessing, or not possessing, them. second, we assess the existing knowledge base and How did things get to be like this? Why is there such agap between we explore the avenues available for its extension; perceptions of numeracy and of literacy –when was it decided that the target is new understanding, it is frequently an being apoet is more glamorous than being an actuary? Ipropose to individual’s quest, and progress lies in the promise take these questions back to the ancient world, because, for all the for new and meaningful investigation that is revealed.
    [Show full text]
  • Mrp1 Is Involved in Lipid Presentation and Inkt Cell Activation by Streptococcus Pneumoniae
    ARTICLE DOI: 10.1038/s41467-018-06646-8 OPEN Mrp1 is involved in lipid presentation and iNKT cell activation by Streptococcus pneumoniae Shilpi Chandra1, James Gray2, William B. Kiosses1, Archana Khurana1, Kaori Hitomi1, Catherine M. Crosby1, Ashu Chawla3, Zheng Fu 3, Meng Zhao1, Natacha Veerapen4, Stewart K. Richardson5, Steven A. Porcelli6, Gurdyal Besra 4, Amy R. Howell5, Sonia Sharma2,7, Bjoern Peters8,9 & Mitchell Kronenberg 1,10 Invariant natural killer T cells (iNKT cells) are activated by lipid antigens presented by CD1d, 1234567890():,; but the pathway leading to lipid antigen presentation remains incompletely characterized. Here we show a whole-genome siRNA screen to elucidate the CD1d presentation pathway. A majority of gene knockdowns that diminish antigen presentation reduced formation of glycolipid-CD1d complexes on the cell surface, including members of the HOPS and ESCRT complexes, genes affecting cytoskeletal rearrangement, and ABC family transporters. We validated the role in vivo for the multidrug resistance protein 1 (Mrp1) in CD1d antigen presentation. Mrp1 deficiency reduces surface clustering of CD1d, which decreased iNKT cell activation. Infected Mrp1 knockout mice show decreased iNKT cell responses to antigens from Streptococcus pneumoniae and were associated with increased mortality. Our results highlight the unique cellular events involved in lipid antigen presentation and show how modification of this pathway can lead to lethal infection. 1 Division of Developmental Immunology, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA. 2 The Functional Genomics Center, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA. 3 Bioinformatics Core, La Jolla Institute for Allergy and Immunology, La Jolla, CA 92037, USA.
    [Show full text]
  • Pnas11052ackreviewers 5098..5136
    Acknowledgment of Reviewers, 2013 The PNAS editors would like to thank all the individuals who dedicated their considerable time and expertise to the journal by serving as reviewers in 2013. Their generous contribution is deeply appreciated. A Harald Ade Takaaki Akaike Heather Allen Ariel Amir Scott Aaronson Karen Adelman Katerina Akassoglou Icarus Allen Ido Amit Stuart Aaronson Zach Adelman Arne Akbar John Allen Angelika Amon Adam Abate Pia Adelroth Erol Akcay Karen Allen Hubert Amrein Abul Abbas David Adelson Mark Akeson Lisa Allen Serge Amselem Tarek Abbas Alan Aderem Anna Akhmanova Nicola Allen Derk Amsen Jonathan Abbatt Neil Adger Shizuo Akira Paul Allen Esther Amstad Shahal Abbo Noam Adir Ramesh Akkina Philip Allen I. Jonathan Amster Patrick Abbot Jess Adkins Klaus Aktories Toby Allen Ronald Amundson Albert Abbott Elizabeth Adkins-Regan Muhammad Alam James Allison Katrin Amunts Geoff Abbott Roee Admon Eric Alani Mead Allison Myron Amusia Larry Abbott Walter Adriani Pietro Alano Isabel Allona Gynheung An Nicholas Abbott Ruedi Aebersold Cedric Alaux Robin Allshire Zhiqiang An Rasha Abdel Rahman Ueli Aebi Maher Alayyoubi Abigail Allwood Ranjit Anand Zalfa Abdel-Malek Martin Aeschlimann Richard Alba Julian Allwood Beau Ances Minori Abe Ruslan Afasizhev Salim Al-Babili Eric Alm David Andelman Kathryn Abel Markus Affolter Salvatore Albani Benjamin Alman John Anderies Asa Abeliovich Dritan Agalliu Silas Alben Steven Almo Gregor Anderluh John Aber David Agard Mark Alber Douglas Almond Bogi Andersen Geoff Abers Aneel Aggarwal Reka Albert Genevieve Almouzni George Andersen Rohan Abeyaratne Anurag Agrawal R. Craig Albertson Noga Alon Gregers Andersen Susan Abmayr Arun Agrawal Roy Alcalay Uri Alon Ken Andersen Ehab Abouheif Paul Agris Antonio Alcami Claudio Alonso Olaf Andersen Soman Abraham H.
    [Show full text]
  • Monoclonal Invariant
    Monoclonal Invariant NKT (iNKT) Cell Mice Reveal a Role for Both Tissue of Origin and the TCR in Development of iNKT Functional Subsets This information is current as of October 29, 2020. Eleanor Clancy-Thompson, Gui Zhen Chen, Paul M. Tyler, Mariah M. Servos, Marta Barisa, Patrick J. Brennan, Hidde L. Ploegh and Stephanie K. Dougan J Immunol 2017; 199:159-171; Prepublished online 2 June 2017; Downloaded from doi: 10.4049/jimmunol.1700214 http://www.jimmunol.org/content/199/1/159 Supplementary http://www.jimmunol.org/content/suppl/2017/05/31/jimmunol.170021 http://www.jimmunol.org/ Material 4.DCSupplemental References This article cites 58 articles, 28 of which you can access for free at: http://www.jimmunol.org/content/199/1/159.full#ref-list-1 Why The JI? Submit online. by guest on October 29, 2020 • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2017 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Monoclonal Invariant NKT (iNKT) Cell Mice Reveal a Role for Both Tissue of Origin and the TCR in Development of iNKT Functional Subsets Eleanor Clancy-Thompson,* Gui Zhen Chen,* Paul M.
    [Show full text]
  • Discovery and Characterization of an Anti-Inflammatory Lipid Derived from Mycobacterium Vaccae
    Discovery and characterization of an anti-inflammatory lipid derived from Mycobacterium vaccae By David G. Smith B.S., The Ohio State University, 2010 A thesis submitted to the Faculty of the Graduate School of the University of Colorado in partial fulfillment of the requirement for the degree of Doctor of Philosophy Department of Chemistry and Biochemistry 2017 i This thesis entitled: Discovery and characterization of an anti-inflammatory lipid derived from Mycobacterium vaccae written by David G. Smith has been approved by the Department of Chemistry and Biochemistry ________________________________________________ Christopher A. Lowry, PhD ________________________________________________ Dylan J. Taatjes, PhD Date______________________ The final copy of this thesis has been examined by the signatories, and we find that both the content and the form meet acceptable presentation standards of scholarly work in the above-mentioned discipline. ii Smith, Gregory David (PhD, Chemistry and Biochemistry) Discovery and characterization of an anti-inflammatory lipid derived from Mycobacterium vaccae Thesis directed by Christopher A. Lowry, PhD. In modern urban environments there is an increased prevalence of allergies, asthma, inflammatory bowel diseases (IBD), and anxiety disorders. The underlying cause for these disorders, as postulated by the Hygiene Hypothesis, is chronic inflammation, and the imbalance in the immune system is caused by a lack of interaction with microbes that have been removed from the urban space and lifestyle. One such microbe is the environmental saprophyte, Mycobacterium vaccae. In animal models, immunization with M. vaccae protects against the development of these prevalent inflammatory disorders. This immunoregulatory effect has been shown to occur due to the expansion of regulatory T cells, but it is still unclear how M.
    [Show full text]
  • Identification and Validation of Mode of Action of Chalcones for Their Anti-Mycobacterial Activity
    Identification and validation of mode of action of chalcones for their anti-mycobacterial activity Bhavani Anagani School of Life and Medical Sciences University of Hertfordshire, College Lane, Hatfield, UK. AL10 9AB 1 “The world is headed for a post-antibiotic era, in which common infections and minor injuries which have been treatable for decades can once again kill.” (WHO, 2015) 2 Tuberculosis • Top infectious killer disease – 1.6 Million people • Mycobacterium tuberculosis • Rifampicin, Isoniazid, Ethambutol, and Pyrazinamide • Multi drug treatment – 6 Months Rod shaped Mycobacterium tuberculosis 3 RR-TB; MDR-TB; XDR-TB; TDR-TB --- Treatment limited • New drugs • New classes • New targets • Understanding the resistance 4 Existing Antibiotics Streptomycin - Streptomyces griseus Rifampicin - Amycolatopsis rifamycinia 5 Chalcones Galenia africana Helichrysum melanacme Dalbergia odorifera 6 Synthesis of chalcones Scheme 1: Synthesis of chalcones 7 Physical data for the chalcones (1a-1o) Table1: In vitro preliminary screening data of Chalcones Substituents MIC (µg/ml) Compound M.wt Clogp m.smegmati J-774 (IC50) SI=IC50/MIC R R R m.bovis 1 2 3 s 1a H H H 270.33 3.35 50 6 50 8.4 1b H Br H 349.22 4.18 - - 25 - 1c Br H H 349.22 4.18 100 25 25 1 1d H H Br 349.22 4.18 200 12 25 2.08 1e H Cl H 304.77 3.91 - - 12 - 1f Cl H H 304.77 3.91 100 12 12 1 1g H H Cl 304.77 3.91 - 200 12 0.06 1h H H F 288.32 3.51 200 25 100 4 1i H H OCH3 300.35 3.23 200 50 100 2 1j H H CH3 284.35 3.84 100 25 25 1 1k H H C6 H5 346.43 5.03 200 100 25 0.25 1l H NO2 H 315.32 3.48 - 100 25 0.25 1m CH3 H H 284.35 3.84 200 50 25 0.5 1n H H NO2 315.32 3.48 - 200 50 0.25 1o Cl H Cl 339.21 4.47 - 200 6 0.03 8 MIC Determination against Mycobacteria • Colorimetric broth micro-dilution method according to CLSI guidelines • M.
    [Show full text]
  • N-Methylation of a Bactericidal Compound As a Resistance Mechanism in Mycobacterium Tuberculosis
    N-methylation of a bactericidal compound as a PNAS PLUS resistance mechanism in Mycobacterium tuberculosis Thulasi Warriera, Kanishk Kapilashramib, Argyrides Argyrouc, Thomas R. Ioergerd, David Littlea, Kenan C. Murphye, Madhumitha Nandakumara, Suna Parka, Ben Golda, Jianjie Mia, Tuo Zhanga, Eugenia Meilerf, Mike Reesc, Selin Somersan-Karakayag, Esther Porras-De Franciscof, Maria Martinez-Hoyosf, Kristin Burns-Huanga, Julia Robertsa, Yan Linga, Kyu Y. Rheea,g, Alfonso Mendoza-Losanaf, Minkui Luob,h, and Carl F. Nathana,1 aDepartment of Microbiology and Immunology, Weill Cornell Medicine, New York, NY 10021; bChemical Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065; cPlatform Technology and Science, GlaxoSmithKline, Stevenage SG1 2NY, United Kingdom; dDepartment of Computer Science and Engineering, Texas A&M University, College Station, TX 77843-3474; eDepartment of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, MA 01655; fDiseases of the Developing World, GlaxoSmithKline (GSK), 28760 Madrid, Spain; gDepartment of Medicine, Weill Cornell Medicine, New York, NY 10021; and hDepartment of Pharmacology, Weill Cornell Medicine, New York, NY 10021 Contributed by Carl F. Nathan, June 8, 2016 (sent for review April 26, 2016; reviewed by Thomas Dick, Babak Javid, and Richard E. Lee) The rising incidence of antimicrobial resistance (AMR) makes it im- accumulation of spontaneous mutations allows selection for drug perative to understand the underlying mechanisms. Mycobacterium resistance. Clinically, resistance to most frontline TB drugs is caused tuberculosis (Mtb) is the single leading cause of death from a bacterial by mutations in the activation pathways of prodrugs (e.g., katG pathogen and estimated to be the leading cause of death from AMR.
    [Show full text]
  • Annual Review 2007/08.Indd
    07/08 Annual review Medical research: benefiting people “RESEARCH IS ABOUT PEOPLE – NOT JUST THE SCIENTISTS BEHIND DISCOVERY, The Medical Research Council is the UK’s leading publicly funded biomedical BUT ALSO THE research organisation. MEMBERS OF Our mission is to: THE PUBLIC • Encourage and support high-quality research WHO HELP with the aim of improving human health. RESEARCHERS IN • Produce skilled researchers, and to advance and disseminate knowledge and technology THEIR QUEST TO to improve the quality of life and economic competitiveness in the UK. IMPROVE HEALTH • Promote dialogue with the public about AND THE MANY medical research. OTHERS WHO ARE AFFECTED BY THE findings.” SIR LESZEK BORYSIEWICZ, MRC CHIEF EXECUTIVE Medical research: benefiting people Molecules, genes & cancer 04 12 Lifelong health & ageing Improving global health 18 26 Nutrition, obesity & diabetes Heart & lung health 32 38 Research into the brain Mental health & addiction 44 50 Developing medicines & technologies Introduction from the MRC Chief Executive, Sir Leszek Borysiewicz SINCE 1913 THE MEDICAL RESEARCH COUNCIL HAS MADE DISCOVERIES THAT HAVE IMPROVED THE HEALTH OF MILLIONS OF PEOPLE IN THE UK AND WORLDWIDE. First and foremost in the MRC’s mission had two heart attacks before she reached is supporting research with the aim of middle age. Her heart specialist is also an MRC improving human health. Our scientists scientist, and what he sees in the clinic with are behind some of the most important patients like Amanda feeds directly back into his medical advances of the past century. research in the lab. David Ward, who recently retired as a journalist, was enrolled in a study Our early work included tackling tuberculosis by his mother before he was born and has now and rickets and developing antibiotics to treat been tracked for 62 years by scientists.
    [Show full text]