DOCSLIB.ORG

Concerning the Biosynthesis of Sirohaem in Bacillus Mesaterium the Relationship with Cohalamin Metabolism

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Concerning the Biosynthesis of Sirohaem in Bacillus Mesaterium the Relationship with Cohalamin Metabolism Concerning the Biosynthesis of Sirohaem in Bacillus mesaterium The Relationship with Cohalamin Metabolism Presented by Mr. Richard Beck For the award of Doctorate (Ph.D.) in Biochemistry at the University of London (University College London). Department of Molecular Genetics, Institute of Ophthalmology, University College London, Bath Street, London EClV 9EL. United Kingdom. March 1997 ProQuest Number: 10044378 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. uest. ProQuest 10044378 Published by ProQuest LLC(2016). Copyright of the Dissertation is held by the Author. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code. Microform Edition © ProQuest LLC. ProQuest LLC 789 East Eisenhower Parkway P.O. Box 1346 Ann Arbor, Ml 48106-1346 Abstract Concerning the Biosynthesis of Sirohaem in Bacillus megaterium: The Relationship with Cobalamin Metabolism Sirohaem and vitamin B 12 (cyanocobalamin) share a significant part of their biosynthetic pathways, the last common intermediate being precorrin -2 (dihydrosirohydrochlorin). Dehydrogenation of precorrin-2 leads to sirohaem formation, whereas méthylation at C-20 directs production of vitamin B 12. A novel enzyme has been identified which may play a crucial role in regulating sirohaem and cobalamin biosynthesis at the precorrin-2 branch-point. In vivo, the CbiX protein from Bacillus megaterium has been shown to catalyse the final two steps of sirohaem synthesis, namely dehydrogenation and ferrochelation. CbiX is encoded within a cobalamin biosynthetic (cob) operon by cbiX. The protein consists of 301 amino acids with a Mr of 34,297. The C-terminus of the protein contains a novel poly-histidine motif and this has enabled the purification of CbiX to homogeneity by metal chelate chromatography. The protein was found to be soluble but relatively unstable, being broken down to give a 24kDa polypeptide. Although the native Mr of CbiX is approximately 62,000, suggesting a dimeric form, the purified protein was found to aggregate, forming multimers which could be visualised by PAGE. The poly-histidine motif is involved in metal binding: deletion from the protein inhibits sirohaem synthesis but truncation, retaining 7 of the 19 histidines, maintains function. In vivo assays have shown that cobalt (the metal associated with vitamin B 12) inhibits the function of CbiX in sirohaem synthesis. Other metals do not show this ability. Deletion of the cbiX gene from the cob operon greatly reduces cobalamin production. The position of the cbiX gene within a cob operon in B. megaterium, deletion experiments with the cbiX gene, and the apparent ability of the CbiX protein to bind cobalt, all suggest regulation of sirohaem and vitamin B 12 production by CbiX at the precorrin-2 branch-point of tetrapyrrole modification. Acknowledgements Throughout the time it has taken to compile this thesis, I have been fortunate to have worked with, and received assistance from, many people. I would particularly like to thank the following by name: Dr. Martin Warren, my supervisor, for his invaluable guidance at every stage. The members of Dr. Warren's research group, namely Sue, Sarah W., Treasa, Sarah A., Ed, Natalie, Evelyne, Seraphina, and lately Jenny, for their help and for putting up with me. Dr. Alain Rambach of CHROMagar and Dr. Claude Thermes of the Centre National de Recherche Scientifique, Paris, for their support and assistance in the early stages, and for providing inspiration. Dr. Lynn Sibanda and Dr. Armando Albert of Birkbeck College, London, for their insights into the world of x-ray crystallography. Dr. J.A. Cole, University of Birmingham, for the gift of 302Aa and 712cyjC::Tn5 Escherichia coli strains. Dr. Lawrence Hunt, University of Southampton, for the protein sequencing work. Dr. Kanwaljit Dulai, Institute of Ophthalmology, for help with computers and associated black boxes. Thanks also to everyone I encountered both at Queen Mary & Westfield College and at the Institute of Ophthalmology, especially members of the football and cricket teams. I am indebted to my parents and to those of my fiancée for their ceaseless support. Dedication To Sonia 7 never felt magic crazy as this, I never saw moons knew the meaning of the sea I never held emotion in the palm of my hand Or felt sweet breezes in the top of a tree But now you're here Brighten my Northern sky' Nick Drake 'Northern Sky' © Warlock Music Ltd, 1970 Contents Abstract 2 Acknowledgements 3 Dedication 4 Contents 5 Glossary 17 Abbreviations Used 17 Sources of Materials 20 Amino Acid Codes 21 The Genetic Code 22 Chapter One The Biosynthesis of Uroporphyrinogen III and its Transformation into Sirohaem and Vitamin B12 23 1 .1 Tetrapyrroles in Biology 24 1.1.1 The Versatility of Tetrapyrroles 24 1 .1.2 Tetrapyrrole-Derived Prosthetic Groups 25 1 . 1 .2.1 Haem 25 1 . 1 .2 .2 Chlorophyll 25 1.1.2.3 Co-enzyme (Factor) F 430 25 1.1.2.4 Vitamin B 12 27 1.1.2.5 Sirohaem 29 1.1.2.6 Siroamide 30 1.2 Biosynthesis and Modification of Uroporphyrinogen III 32 1.2.1 Biosynthesis of Uroporphyrinogen III 32 1.2.1.1 Biosynthesis of 5-Aminolaevulinic Acid 32 1.2.1.2 Biosynthesis of Uroporphyrinogen III from ALA 33 1.2.2 Biosynthesis of Haem and Chlorophyll 36 1.2.3 Biosynthesis of Precorrin-2 38 1.2.4 Biosynthesis of Co-enzyme F4 30 from Precorrin-2 3 8 1.2.5 Biosynthesis of Vitamin B12 from Precorrin-2 3 8 1.2.6 Biosynthesis of Sirohaem from Precorrin-2 39 1.3 Molecular Genetics and Biochemistry of Sirohaem Biosynthesis 40 1.3.1 Sirohaem Synthesis in Escherichia coli 40 1.3.1.1 The Multi-Functional Sirohaem Synthase Enzyme 40 1.3.1.2 The Regulation of Sirohaem Synthase Expression in E. coli 4 1 1.3.1.3 Sirohaem Synthase in Salmonella typhimurium and Neisseria meningitidis 43 1.3.2 Sirohaem Synthesis in Bacillus megaterium and Pseudomonas denitrificans 43 1.3.2.1 A Discrete Uro'gen III Methylase 43 1.3.3 Sirohaem Biosynthesis in Other Species 44 1.3.3.1 Putative Multifunctional SUMT Enzymes 44 1.3.3.2 Other Discrete SUMT Enzymes 45 1.3.4 Classes of SUMT Enzymes 49 1.3.4.1 The Precorrin-2 Branch-Point of Sirohaem and Vitamin B 12 Biosynthesis 50 1.4 Molecular Genetics and Biochemistry of Cobalamin Biosynthesis 51 1.4.1 Bacterial Cobalamin Synthesis 51 1.4.1.1 Early work in P. denitrificans and S. typhimurium 51 1.4.1.2 CysG and Vitamin B 12 Biosynthesis 52 1.4.1.3 A cob Operon in Bacillus megaterium 53 1.4.1.4 Other Potential cob Opérons 57 1.4.2 Evolution of a Vitamin B 12 Biosynthetic Pathway 59 1.4.2.1 Similarities and Differences Bet wen the cob Opérons 59 1.4.2.2 A Case for Re-acquisition of the cob Operon 61 1.5 The Aims of this Thesis 64 Chapter Two Materials and Methods 65 2.1 General Techniques 66 2.1.1 Growth Media 66 2.1.1.1 LB Rich Broth and Agar 66 2.1.1.2 Minimal Medium (M9) Broth and Agar 67 2.1.1.3 Supplements to the Growth Media 67 2.1.1.4 Sterilisation 68 2.1.1.5 Buffers and Solutions 69 2.1.2 Bacterial Strains 71 2.1.3 Bacterial Manipulation 72 2.1.3.1 Storage of Baeterial Strains 72 2.1.3.2 Preparation of Competent Bacteria 72 2.1.3.3 Genomic DNA Extraction 72 2.1.3.4 Plasmid Preparation 73 2.1.3.5 T ransformation 7 4 2.1.3.6 Complementation 7 5 2.1.3.7 Growth Curves 75 2.1.4 DNA Analysis 7 6 2.1.4.1 Restriction Enzyme Digestion 76 2.1.4.2 Agarose Gel Electrophoresis 76 2.1.4.3 DNA Size Markers 77 2.1.5 Isolation and Recovery of DNA from Agarose Gels 7 8 2.1.5.1 GeneClean 78 2.1.5.2 Creation of Blunt Ends' 78 2.1.5.3 Ligation 79 2.1.6 Polymerase Chain Reaction 80 2.1.6.1 Reaction Protocol 80 2.1.6.2 Expression-Cassette PCR 80 2.1.6.3 Thermal Cycling 81 2.1.6.4 PCR Primers 82 2.1.7 Plasmid Library 8 3 2.1.8 DNA Sequencing 85 2.1.8.1 Reaction Protocol 85 2.1.8.2 Gel Preparation 86 2.1.8.3 Eleetrophoresis 87 2.2 Protein Purification 88 2.2.1 Gene Expression 88 2.2.1.1 pKK223.3 and pCIQ Plasmids 88 2.2.1.2 Growth of E. coli and Over-Expression of Protein 88 2.2.1.3 Large-scale Protein Preparation 89 2.2.2 Polyacrylamide Gel Electrophoresis 89 2.2.2.1 Gel Assembly 89 2.2.2.2 Sample Preparation and Gel Loading 90 2.2.2.3 Protein Standard Markers 91 2.2.2.4 Staining and De-staining 91 2.2.2.5 Native (non-SDS) PAGE 91 2.2.3 Purification Techniques 92 2.2.3.1 Buffers 92 2.2.3.2 Sonication 93 2.2.3.3 Centrifugation 93 2.2.3.4 Ammonium Sulphate Differential Precipitation 93 2.2.4 Chromatography 95 2.2.4.1 Ion Exchange 95 2.2.4.2 Gel Filtration 95 2.2.4.3 FPLC 96 2.2.4.4 Metal Binding 97 2.3 Protein Assessment 99 2.3.1 Protein Sequencing 99 2.3.1.1 SDS-PAGE 99 2.3.1.2 Protein Blotting 99 2.3.1.3 PVDF Membrane Staining and Drying 100 2.3.2 Spectroscopic Analysis of Tetrapyrrole Biosynthesis 101 2.3.2.1 Propogation and Harvesting of Bacteria 101 2.3.2.2 Isolation of Non-Protein Material from Bacteria 101 2.3.2.3 Spectroscopy 101 Chapter Three Identification of the Bacillus megaterium Genes Required for the Complementation of the Escherichia coli cysG Deletion Strain, 302Aa 102 3.1 Introduction 103 3.1.1 The CysG Protein and Definition of Cysteine Auxotrophy 103 3.1.2 Sirohaem Synthesis in Bacillus megaterium 104 3.1.3 Verification of the B.
Load more

Recommended publications
  • An Investigation of the Chlorophylls of Selected Prasinophyte Algae
    University of Nebraska at Omaha DigitalCommons@UNO Student Work 5-1-1986 An investigation of the chlorophylls of selected prasinophyte algae. Leslie Carlat Kwasnieski Follow this and additional works at: https://digitalcommons.unomaha.edu/studentwork Recommended Citation Kwasnieski, Leslie Carlat, "An investigation of the chlorophylls of selected prasinophyte algae." (1986). Student Work. 3376. https://digitalcommons.unomaha.edu/studentwork/3376 This Thesis is brought to you for free and open access by DigitalCommons@UNO. It has been accepted for inclusion in Student Work by an authorized administrator of DigitalCommons@UNO. For more information, please contact [email protected]. AN INVESTIGATION OF THE CHLOROPHYLLS OF SELECTED PRASINOPHYTE ALGAE A Thesis Presented to the Department of Biology and the Faculty of the Graduate College U n iv e rs ity o f Nebraska In Partial Fulfillment o f the Requirements fo r the Degree Master of Arts University of Nebraska at Omaha by Leslie Carl at Kwasnieski May, 1986 UMI Number: EP74978 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. UMI EP74978 Published by ProQuest LLC (2015). Copyright in the Dissertation held by the Author. Microform Edition © ProQuest LLC. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code ProQuest ProQuest LLC. 789 East Eisenhower Parkway P.O. Box 1345 Ann Arbor, Ml 48105-1346 THESIS ACCEPTANCE Accepted for the faculty of the Graduate College, University of Nebraska, in partial fulfillm ent of the requirements for the degree Master of Arts, University of Nebraska at Omaha.
    [Show full text]
  • Erythropoietic Protoporphyrias: Studies of the Natural History, Genotype-Phenotype Correlations, and Psychosocial Impact
    Erythropoietic Protoporphyrias: Studies of the Natural History, Genotype-Phenotype Correlations, and Psychosocial Impact PI: Dr. Manisha Balwani NCT01688895 Document Date: Jun 26, 2015 RDCRN PC #7207 EPP Natural History Version Date: V2 26Jun2015 Rare Diseases Clinical Research Network Porphyrias Consortium Erythropoietic Protoporphyrias: Studies of the Natural History, Genotype-Phenotype Correlations, and Psychosocial Impact This protocol is for research purposes only and should not be copied, redistributed, or used for any other purpose. The procedures in this protocol are intended only for use by the Porphyrias Consortium investigators in carefully controlled settings. The Study Chair of this study should be consulted before using this protocol. Study Chair: Manisha Balwani, MD, MS Department of Genetics & Genomic Sciences Icahn School of Medicine at Mount Sinai One Gustave L. Levy Place, Box 1497 New York, NY 10029 Phone: 212-241-0915 Fax: 212-426-9065 Email: [email protected] Page 1 of 31 RDCRN PC #7207 EPP Natural History Version Date: V2 26Jun2015 Table of Contents Contents Participating Institutions/Investigators Table (contact information)............................................................... 4 1. Synopsis .................................................................................................................................................... 6 1.A. Protocol Overview .............................................................................................................................. 8 2. Objective
    [Show full text]
  • Hyperbilirubinemia
    Porphyrins Porphyrins (Porphins) are cyclic tetrapyrol compounds formed by the linkage )). of four pyrrole rings through methenyl bridges (( HC In the reduced porphyrins (Porphyrinogens) the linkage of four pyrrole rings (tetrapyrol) through methylene bridges (( CH2 )) The characteristic property of porphyrins is the formation of complexes with the metal ion bound to nitrogen atoms of the pyrrole rings. e.g. Heme (iron porphyrin). Proteins which contain heme ((hemoproteins)) are widely distributed e.g. Hemoglobin, Myoglobin, Cytochromes, Catalase & Tryptophan pyrrolase. Natural porphyrins have substituent side chains on the eight hydrogen atoms numbered on the pyrrole rings. These side chains are: CH 1-Methyl-group (M)… (( 3 )) 2-Acetate-group (A)… (( CH2COOH )) 3-Propionate-group (P)… (( CH2CH2COOH )) 4-Vinyl-group (V)… (( CH CH2 )) Porphyrins with asymmetric arrangement of the side chains are classified as type III porphyrins while those with symmetric arrangement of the side chains are classified as type I porphyrins. Only types I & III are present in nature & type III series is more important because it includes heme. 1 Heme Biosynthesis Heme biosynthesis occurs through the following steps: 1-The starting reaction is the condensation between succinyl-CoA ((derived from citric acid cycle in the mitochondria)) & glycine, this reaction is a rate limiting reaction in the hepatic heme synthesis, it occurs in the mitochondria & is catalyzed by ALA synthase (Aminolevulinate synthase) enzyme in the presence of pyridoxal phosphate as a cofactor. The product of this reaction is α-amino-β-ketoadipate which is rapidly decarboxylated to form δ-aminolevulinate (ALA). 2-In the cytoplasm condensation reaction between two molecules of ALA is catalyzed by ALA dehydratase enzyme to form two molecules of water & one 2 molecule of porphobilinogen (PBG) which is a precursor of pyrrole.
    [Show full text]
  • Levels of 6-Aminolevulinate Dehydratase, Uroporphyrinogen-I Synthase, and Protoporphyrin IX in Erythrocytes from Anemic Mutant M
    Proc. Natl. Acad. Sci. USA Vol. 74, No. 3, pp. 1181-1184, March 1977 Genetics Levels of 6-aminolevulinate dehydratase, uroporphyrinogen-I synthase, and protoporphyrin IX in erythrocytes from anemic mutant mice (hemolytic disease/reticulocytosis/heme biosynthesis/development) SHIGERU SASSA* AND SELDON E. BERNSTEINt * The Rockefeller University, New York, N.Y. 10021; and t The Jackson Laboratory, Bar Harbor, Maine 04609 Communicated by Elizabeth S. Russell, December 23, 1976 ABSTRACT Levels of erythrocyte 6-aminolevulinate MATERIALS AND METHODS dehydratase [ALA-dehydratase; porphobilinogen synthase; 5- aminolevulinate hydro-lyase (adding 5-aminolevulinate and Animals. Genetically anemic mutant mice, their littermate cyclizing), EC 4.2.1.241, uroporphyrinogen-I synthase [Uro- controls, and the C57BL/6J and DBA/2J inbred strains were synthase; porphobilinogen ammonia-lyase (polymerizing), EC reared in research colonies of the Jackson Laboratory. The or- 4.3.1.8], and protoporphyrin IX (Proto) were measured by sen- igins and hematological, as well as general, characteristics of sitive semimicroassays using 2-5 IAl of whole blood obtained the various types of anemic mice have been reviewed elsewhere from normal and anemic mutant mice. The levels of erythrocyte (1-3). The mutants employed in these studies of ALA-dehydratase and Uro-synthase showed marked develop- hemolytic mental changes and ALA-dehydratase was influenced by the disease included: jaundice (ja) (4), spherocytosis (sph) (5), he- L v gene. molytic anemia (ha) (1, 6), and normoblastosis (nb) (1, 7). Other Mice with overt hemolytic diseases (ja/ja, sph/sph, nb/nb, anemias used were: microcytosis (mk) (8), Steel (SI) (9), W series ha/ha) had 10- to 20-fold increases in ALA-dehydratase, Uro- (W) (10), and Hertwig's anemia (an) (11).
    [Show full text]
  • RLIN1, Encoding a Putative Coproporphyrinogen III Oxidase, Is Involved in Lesion Initiation in Rice
    Available online at www.sciencedirect.com Journal of Genetics and Genomics 38 (2011) 29e37 www.jgenetgenomics.org RLIN1, encoding a putative coproporphyrinogen III oxidase, is involved in lesion initiation in rice Changhui Sun a,b,d,1, Linchuan Liu b,c,1, Jiuyou Tang b, Aihong Lin b,c, Fantao Zhang b,d, Jun Fang b, Genfa Zhang a,*, Chengcai Chu a,b,c,* a Key Laboratory for Cell Proliferation and Regulation Biology of Ministry of Education, College of Life Science, Beijing Normal University, Beijing 100875, China b The State Key Laboratory of Plant Genomics and National Plant Gene Research Center (Beijing), Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Datun Road, Chaoyang District, Beijing 100101, China c Graduate School of the Chinese Academy of Sciences, Yuquan Road, Beijing 100039, China d Rice Research Institute, Sichuan Agricultural University, Chengdu 611130, China Received 11 October 2010; revised 17 October 2010; accepted 20 October 2010 Abstract Lesion mimic is necrotic lesions on plant leaf or stem in the absence of pathogenic infection, and its exact biological mechanism is varied. By a large-scale screening of our T-DNA mutant population, we identified a mutant rice lesion initiation 1 (rlin1), which was controlled by a single nuclear recessive gene. Map-based cloning revealed that RLIN1 encoded a putative coproporphyrinogen III oxidase in tetrapyrrole biosynthesis pathway. Sequencing results showed that a G to T substitution occurred in the second exon of RLIN1 and led to a missense mutation from Asp to Tyr. Ectopic expression of RLIN1 could rescue rlin1 lesion mimic phenotype.
    [Show full text]
  • Porphyrins & Bile Pigments
    Bio. 2. ASPU. Lectu.6. Prof. Dr. F. ALQuobaili Porphyrins & Bile Pigments • Biomedical Importance These topics are closely related, because heme is synthesized from porphyrins and iron, and the products of degradation of heme are the bile pigments and iron. Knowledge of the biochemistry of the porphyrins and of heme is basic to understanding the varied functions of hemoproteins in the body. The porphyrias are a group of diseases caused by abnormalities in the pathway of biosynthesis of the various porphyrins. A much more prevalent clinical condition is jaundice, due to elevation of bilirubin in the plasma, due to overproduction of bilirubin or to failure of its excretion and is seen in numerous diseases ranging from hemolytic anemias to viral hepatitis and to cancer of the pancreas. • Metalloporphyrins & Hemoproteins Are Important in Nature Porphyrins are cyclic compounds formed by the linkage of four pyrrole rings through methyne (==HC—) bridges. A characteristic property of the porphyrins is the formation of complexes with metal ions bound to the nitrogen atom of the pyrrole rings. Examples are the iron porphyrins such as heme of hemoglobin and the magnesium‐containing porphyrin chlorophyll, the photosynthetic pigment of plants. • Natural Porphyrins Have Substituent Side Chains on the Porphin Nucleus The porphyrins found in nature are compounds in which various side chains are substituted for the eight hydrogen atoms numbered in the porphyrin nucleus. As a simple means of showing these substitutions, Fischer proposed a shorthand formula in which the methyne bridges are omitted and a porphyrin with this type of asymmetric substitution is classified as a type III porphyrin.
    [Show full text]
  • UROPORPHYRINOGEN HII COSYNTHETASE in HUMAN Hemolysates from Five Patientswith Congenital Erythropoietic Porphyriawas Much Lower
    UROPORPHYRINOGEN HII COSYNTHETASE IN HUMAN CONGENITAL ERYTHROPOIETIC PORPHYRIA * BY GIOVANNI ROMEO AND EPHRAIM Y. LEVIN DEPARTMENT OF PEDIATRICS, THE JOHNS HOPKINS UNIVERSITY SCHOOL OF MEDICINE Communicated by William L. Straus, Jr., April 24, 1969 Abstract.-Activity of the enzyme uroporphyrinogen III cosynthetase in hemolysates from five patients with congenital erythropoietic porphyria was much lower than the activity in control samples. The low cosynthetase activity in patients was not due to the presence of a free inhibitor or some competing en- zymatic activity, because hemolysates from porphyric subjects did not interfere either with the cosynthetase activity of hemolysates from normal subjects or with cosynthetase prepared from hematopoietic mouse spleen. This partial deficiency of cosynthetase in congenital erythropoietic porphyria corresponds to that shown previously in the clinically similar erythropoietic porphyria of cattle and explains the overproduction of uroporphyrin I in the human disease. Erythropoietic porphyria is a rare congenital disorder of man and cattle, characterized by photosensitivity, erythrodontia, hemolytic anemia, and por- phyrinuria.1 Many of the clinical manifestations of the disease can be explained by the production in marrow, deposition in tissues, and excretion in the urine and feces, of large amounts of uroporphyrin I and coproporphyrin I, which are products of the spontaneous oxidation of uroporphyrinogen I and its decarboxyl- ated derivative, coproporphyrinogen I. In cattle, the condition is inherited
    [Show full text]
  • Noncanonical Coproporphyrin-Dependent Bacterial Heme Biosynthesis Pathway That Does Not Use Protoporphyrin
    Noncanonical coproporphyrin-dependent bacterial heme biosynthesis pathway that does not use protoporphyrin Harry A. Daileya,b,c,1, Svetlana Gerdesd, Tamara A. Daileya,b,c, Joseph S. Burcha, and John D. Phillipse aBiomedical and Health Sciences Institute and Departments of bMicrobiology and cBiochemistry and Molecular Biology, University of Georgia, Athens, GA 30602; dMathematics and Computer Science Division, Argonne National Laboratory, Argonne, IL 60439; and eDivision of Hematology, Department of Medicine, University of Utah School of Medicine, Salt Lake City, UT 84132 Edited by J. Clark Lagarias, University of California, Davis, CA, and approved January 12, 2015 (received for review August 25, 2014) It has been generally accepted that biosynthesis of protoheme of a “primitive” pathway in Desulfovibrio vulgaris (13). This path- (heme) uses a common set of core metabolic intermediates that way, named the “alternative heme biosynthesis” path (or ahb), has includes protoporphyrin. Herein, we show that the Actinobacteria now been characterized by Warren and coworkers (15) in sulfate- and Firmicutes (high-GC and low-GC Gram-positive bacteria) are reducing bacteria. In the ahb pathway, siroheme, synthesized unable to synthesize protoporphyrin. Instead, they oxidize copro- from uroporphyrinogen III, can be further metabolized by suc- porphyrinogen to coproporphyrin, insert ferrous iron to make Fe- cessive demethylation and decarboxylation to yield protoheme (14, coproporphyrin (coproheme), and then decarboxylate coproheme 15) (Fig. 1 and Fig. S1). A similar pathway exists for protoheme- to generate protoheme. This pathway is specified by three genes containing archaea (15, 16). named hemY, hemH, and hemQ. The analysis of 982 representa- Current gene annotations suggest that all enzymes for pro- tive prokaryotic genomes is consistent with this pathway being karyotic heme synthetic pathways are now identified.
    [Show full text]
  • Published Version (PDF 2MB)
    This may be the author’s version of a work that was submitted/accepted for publication in the following source: Harris, Tegan M., Price, Erin P., Sarovich, Derek S., Nørskov-Lauritsen, Niels, Beissbarth, Jemima, Chang, Anne B., & Smith-Vaughan, Heidi C. (2020) Comparative genomic analysis identifies x-factor (Haemin)-independent haemophilus haemolyticus:A formal re-classification of ’haemophilus in- termedius’. Microbial Genomics, 6(1), Article number: 000303. This file was downloaded from: https://eprints.qut.edu.au/211802/ c 2020 The Authors This work is covered by copyright. Unless the document is being made available under a Creative Commons Licence, you must assume that re-use is limited to personal use and that permission from the copyright owner must be obtained for all other uses. If the docu- ment is available under a Creative Commons License (or other specified license) then refer to the Licence for details of permitted re-use. It is a condition of access that users recog- nise and abide by the legal requirements associated with these rights. If you believe that this work infringes copyright please provide details by email to [email protected] License: Creative Commons: Attribution 4.0 Notice: Please note that this document may not be the Version of Record (i.e. published version) of the work. Author manuscript versions (as Sub- mitted for peer review or as Accepted for publication after peer review) can be identified by an absence of publisher branding and/or typeset appear- ance. If there is any doubt, please refer to the published source. https://doi.org/10.1099/mgen.0.000303 RESEARCH ARTICLE Harris et al., Microbial Genomics 2020;6 DOI 10.1099/mgen.0.000303 Comparative genomic analysis identifies X- factor (haemin)- independent Haemophilus haemolyticus: a formal re-classification of 'Haemophilus intermedius' Tegan M.
    [Show full text]
  • <I>Lactobacillus Reuteri</I>
    University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln Faculty Publications in Food Science and Food Science and Technology Department Technology 2014 From prediction to function using evolutionary genomics: Human-specific ecotypes of Lactobacillus reuteri have diverse probiotic functions Jennifer K. Spinler Texas Children’s Hospital, [email protected] Amrita Sontakke Baylor College of Medicine Emily B. Hollister Baylor College of Medicine Susan F. Venable Baylor College of Medicine Phaik Lyn Oh University of Nebraska, Lincoln See next page for additional authors Follow this and additional works at: http://digitalcommons.unl.edu/foodsciefacpub Spinler, Jennifer K.; Sontakke, Amrita; Hollister, Emily B.; Venable, Susan F.; Oh, Phaik Lyn; Balderas, Miriam A.; Saulnier, Delphine M.A.; Mistretta, Toni-Ann; Devaraj, Sridevi; Walter, Jens; Versalovic, James; and Highlander, Sarah K., "From prediction to function using evolutionary genomics: Human-specific ce otypes of Lactobacillus reuteri have diverse probiotic functions" (2014). Faculty Publications in Food Science and Technology. 132. http://digitalcommons.unl.edu/foodsciefacpub/132 This Article is brought to you for free and open access by the Food Science and Technology Department at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Faculty Publications in Food Science and Technology by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. Authors Jennifer K. Spinler, Amrita Sontakke, Emily B. Hollister,
    [Show full text]
  • Open Matthew R Moreau Ph.D. Dissertation Finalfinal.Pdf
    The Pennsylvania State University The Graduate School Department of Veterinary and Biomedical Sciences Pathobiology Program PATHOGENOMICS AND SOURCE DYNAMICS OF SALMONELLA ENTERICA SEROVAR ENTERITIDIS A Dissertation in Pathobiology by Matthew Raymond Moreau 2015 Matthew R. Moreau Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy May 2015 The Dissertation of Matthew R. Moreau was reviewed and approved* by the following: Subhashinie Kariyawasam Associate Professor, Veterinary and Biomedical Sciences Dissertation Adviser Co-Chair of Committee Bhushan M. Jayarao Professor, Veterinary and Biomedical Sciences Dissertation Adviser Co-Chair of Committee Mary J. Kennett Professor, Veterinary and Biomedical Sciences Vijay Kumar Assistant Professor, Department of Nutritional Sciences Anthony Schmitt Associate Professor, Veterinary and Biomedical Sciences Head of the Pathobiology Graduate Program *Signatures are on file in the Graduate School iii ABSTRACT Salmonella enterica serovar Enteritidis (SE) is one of the most frequent common causes of morbidity and mortality in humans due to consumption of contaminated eggs and egg products. The association between egg contamination and foodborne outbreaks of SE suggests egg derived SE might be more adept to cause human illness than SE from other sources. Therefore, there is a need to understand the molecular mechanisms underlying the ability of egg- derived SE to colonize the chicken intestinal and reproductive tracts and cause disease in the human host. To this end, the present study was carried out in three objectives. The first objective was to sequence two egg-derived SE isolates belonging to the PFGE type JEGX01.0004 to identify the genes that might be involved in SE colonization and/or pathogenesis.
    [Show full text]
  • Mapping Mass Spectral Databases in Genome-Scale Metabolic Networks Reveals Poorly Covered Areas
    H OH metabolites OH Article Mind the Gap: Mapping Mass Spectral Databases in Genome-Scale Metabolic Networks Reveals Poorly Covered Areas Clément Frainay 1 , Emma L. Schymanski 2,3, Steffen Neumann 4,5 , Benjamin Merlet 1, Reza M. Salek 6 , Fabien Jourdan 1,* and Oscar Yanes 7,8,* 1 Toxalim (Research Centre in Food Toxicology), Université de Toulouse, INRA, ENVT, INP-Purpan, UPS, 31555 Toulouse, France; [email protected] (C.F.); [email protected] (B.M.) 2 Eawag: Swiss Federal Institute for Aquatic Science and Technology, Überlandstrasse 133, 8600 Dübendorf, Switzerland; [email protected] 3 Luxembourg Centre for Systems Biomedicine (LCSB), University of Luxembourg, 7, avenue des Hauts-Fourneaux, L-4362 Esch-sur-Alzette, Luxembourg 4 Leibniz Institute of Plant Biochemistry, Department of Stress and Developmental Biology, Weinberg 3, 06120 Halle, Germany; [email protected] 5 German Centre for Integrative Biodiversity Research (iDiv), Halle-Jena-Leipzig Deutscher Platz 5e, 04103 Leipzig, Germany 6 The International Agency for Research on Cancer (IARC), 150 Cours Albert Thomas, 69372 Lyon CEDEX 08, France; [email protected] 7 Metabolomics Platform, IISPV, Department of Electronic Engineering, Universitat Rovira i Virgili, Avinguda Paisos Catalans 26, 43007 Tarragona, Spain 8 Spanish Biomedical Research Center in Diabetes and Associated Metabolic Disorders (CIBERDEM), Monforte de Lemos 3-5, 28029 Madrid, Spain * Correspondence: [email protected] (F.J.); [email protected] (O.Y.); Tel.: +33-582-066-395 (F.J.); +34-977-776-617 (O.Y.) Received: 18 July 2018; Accepted: 7 September 2018; Published: 15 September 2018 Abstract: The use of mass spectrometry-based metabolomics to study human, plant and microbial biochemistry and their interactions with the environment largely depends on the ability to annotate metabolite structures by matching mass spectral features of the measured metabolites to curated spectra of reference standards.
    [Show full text]