This book covers the broad microbiological applications ofproteomics and mass spectrometry. It is divided into six sectionsthat follow the general progression in which most microbiologylaboratories are approaching the subject –Transition, Tools,Preparation, Profiling by Patterns, Target Proteins, and DataAnalysis.
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About the Author
Professor Haroun N. Shah is Head of Molecular Identification Services Unit at the Centre for Infections, Health Protection Agency, London. The Centre for Infections provides infectious disease surveillance and microbial identification services, co-ordinating the investigation and cause of national and uncommon outbreaks of diseases. Haroun holds several chairs at various universities and spent 25 years in academic life at the University of London.
Professor Saheer E. Gharbia is Head of the Applied and Functional Genomics Unit at the HPA Centre for Infections. She has a PhD in molecular Genetics and Biochemistry, and has held various academic positions at the University of London and McGill University, Canada.
Table of Contents
PrefaceList of contributors Microbial Characterisation; the Transition from Conventional Methods to Proteomics.1) CHANGING CONCEPTS IN THE CHARACTERISATION OF MICROBES AND THE INFLUENCE OF MASS SPECTROMETRYHaroun Shah et al1.1 Background and early attempts to use mass spectrometry on microbes.1.2 Characterisation of microorganisms by MALDI-TOF mass spectrometry; from initial ideas to the development of the first comprehensive database.1.3 Characterisation of microorganisms from their intracellular/membrane bound protein profiles using affinity capture with particular reference to SELDI-TOF-MS.1.4 Comparative analysis of proteomes of diverse strains within a species; use of 2-d fluorescence difference gel electrophoresis (dige).1.5 Searching for low abundant and low molecular weight proteins and peptides using nanoparticles as a selective and concentration probes for MALDI-TOF-MS analysis.2) MICROBIAL PHYLOGENY AND EVOLUTION BASED ON PROTEIN SEQUENCES (THE CHANGE FROM TARGETED GENES TO PROTEINS)Radhey Gupta2.1 Abstract2.2 Microbial phylogeny: overview and key unresolved issues2.3 New protein-based molecular markers for systematic and evolutionary studies2.4 Molecular markers elucidating the evolutionary relationships among alpha (a)-proteobacteria2.5 Molecular markers for the bacteroidetes-chlorobi phyla2.6 Branching order and interrelationships among bacterial phyla2.7 Importance of protein markers for discovering unique properties for different groups of bacteria2.8 Concluding remarks2.9 Acknowledgements2.10 References2: PROTEOMICS TOOLS AND BIOMARKER DISCOVERY.3) OVERVIEW OF THE PROTEOMIC TOOLS AND IT LINKS TO GENOMICSRaju Misra.3.1 Protein identification3.2 Peptide Mass Fingerprint (PMF)3.3 Peptide Fragment Fingerprint (PFF)3.4 Peptide sequencing3.5 False discovery rates (FDR)3.6 Validating protein identifications3.7 Reference Database3.8 Data storage3.9 Biomarker discovery3.10 Integrating genomics with proteomics3.11 Reference List4) HIGH THROUGHPUT BIOMARKER DISCOVERY IN MICROORGANISMSMing Fang4.1 MALDI vs ESI4.2 Tandem Mass Spectrometry and Hybrid Mass Spectrometers4.3 Fragmentation in Tandem Mass SpectrometryProteomic Strategies for Protein Identification1. Bottom-up Proteomics2. Top-down ProteomicsMultidimensional Protein IdentificationMass Spectrometry Based Targeted Protein Quantification and Biomarker DiscoverySelected Reaction MonitoringConclusions5) MALDI MASS SPECTROMETRY IMAGING, A NEW FRONTIER IN BIOSTRUCTURAL TECHNIQUES: APPLICATIONS IN BIOMEDICINE Simona Francese and Malcolm R. Clench5.1 Introduction5.2 Practical Aspects of MALDI-MSI5.2 Applications5.3 Microbial molecular investigation by MALDI TOF MS5.4 Conclusions5.5 References3: PROTEIN SAMPLES PREPARATION TECHNIQUESCONVENTIONAL APPROACHES FOR SAMPLE PREPARATION FOR LIQUIDCHROMATOGRAPHY AND TWO-DIMENSIONAL GEL ELECTROPHORESISVesela Encheva and Robert Parker6.1 Introduction6.2 Cell lysis methods6.3 Sample preparation for 2D GE6.4 Fractionation strategies6.5 Sample preparation for Liquid Chromatography coupled to mass6.6 Conclusion6.7 Reference list7) ISOLATION AND PREPARATION OF SPORE PROTEINS AND SUBSEQUENT CHARACTERISATION BY ELECTROPHORESIS AND MASS SPECTROMETRY Nicola Thorne, Saheer Gharbia and Haroun Shah7.1 Introduction7.2 Experimental2.1 Sporulation media7.3 Conclusion8) CHARACTERIZATION OF BACTERIAL MEMBRANE PROTEINS USING A NOVEL COMBINATION OF A LIPID BASED PROTEIN IMMOBILIZATION TECHNIQUE WITH MASS SPECTROMETRYRoger Karlsson, Darren Chooneea, Elisabet Carlsohn, Vesela Encheva and Haroun Shah8.1 Introduction8.2 The surface proteome8.3 Proteomics of pathogenic bacteria8.4 Lipid-based protein immobilization technology8.5 Salmonella Typhimurium – disease mechanism and outer membrane proteins8.6 Outer membrane proteins of S. Typhimurium8.7 Helicobacter pylori – disease mechanism and outer membrane proteins8.8 Surface proteins of intact Helicobacter pylori9) Wider Protein Detection from Biological Extracts by the Reduction of Dynamic Concentration Range.Luc Guerrier, Egisto Boschetti and Piergiorgi Roghetti9.1 Introduction9.2 Dealing with low-abundance protein discovery9.3 Conclusions and future prospects9.4 References10) 3D-gel electrophoresis - a new development in protein analysis.Robert Ventzki and Josef Stegemann10.1. Introduction10.2. Methods10.3 Results and discussion10.4 ReferencesSECTION 4: CHARACTERISATION OF MICROORGANISMS BY PATTERN MATCHING OF MASS SPECTRAL PROFILES AND BIOMARKER APPROACHES REQUIRING MINIMAL SAMPLE PREPARATION.11) Microbial Disease Biomarkers using ProteinChip ArraysShea Hamilton, Michael Levin, J. Simon Kroll, Paul R. Langford11.1 Introduction11.2 Biomarker studies involving patients infected with viruses11.3 Biomarker studies involving patients infected with parasites11.4 Biomarker studies involving patients infected with bacteria11.5 Other diseases of possible infectious origin11.6 Conclusions11.7 References12) MALDI-TOF MS and microbial identification: years of experimentaldevelopment to an established protocol.Wibke Kallow, Marcel Erhard,Haroun N. Shah, Emmanuel Raptakis, Martin Welker.12.1 Identification of Microorganisms in Clinical Routine12.2 Mass Spectrometry and Microbiology12.3 Mass Spectral ‘Fingerprints’ of Whole Cells12.4 Reproducibility of Mass Spectral Fingerprints12.5 Species and Strain Discrimination by Mass Spectrometry12.6 Pattern Matching Approaches for automated Identification12.7 Mass Spectral Identification of Microorganism – Requirements for Routine Diagnostics12.8 Automated Mass Spectral Analysis of Microorganisms in Clinical Routine Diagnostics12.9 Acknowledgements and references5: Targeted Molecules and Analysis of Specific Microorganisms.13) Whole Cell MALDI Mass Spectrometry for the Rapid Characterisation ofBacteria; A Survey of Applications to Major Phyletic Lines in MicrobialKingdom.Ben van Baar13.1 Introduction13.2Scope13.3 Reproducibility13.3.1 Factors concerning the sample13.4 Factors concerning the MALDI MS process13.5 Sample application and ionisation13.5 Data analysis13.6 Spectrum libraries13.6Whole cell MALDI MS of particular bacteria genera and speciesBacillus spp.Staphylococcus spp.Streptococcus spp.Mycobacterium spp.Other Gram-positive bacteriaEscherichia coliGram-negative food- and waterborne pathogen proteobacteria, other than E. ColiTypical sexually transmitted pathogens: Neisseria spp. and Haemophilus spp.Gram-negative biothreat agent bacteriaOther Gram-negative bacteriaPathogenic CyanobacteriaStrategies for the identification of biomarkers in whole cell MALDI MS spectraProtein database considerationOn-target treatment and analysisOff-target’ Analysis and correlation with proteomics studiesGeneral consideration of biomarker identification strategiesConclusions and outlook14) The power of Gel-based proteomics to understandphysiology in Bacillus subtilisHaike Antelmann and Michael HeckerIntroductionResults1 Proteomics of protein secretion mechanisms in Bacillus subtilis1.1. Protein export machineries of B. subtilis1.1 The extracellular proteome of B. subtilis1.2 The cell wall proteome of B. subtilis1.3. The membrane attached lipoproteome of B. subtilis1.3 The proteome analysis of protein secretion mechanisms in B. subtilis2 Definition of proteomic signatures to study cell physiology2.1. Proteomic signatures of B. subtilis in response to stress and starvation2.2. Proteomic signatures of B. subtilis in response to thiol-reactive electrophiles uncovered novel regulatory mechanisms2.3. The MarR/DUF24-family YodB repressor is directly sensing thiol- reactive electrophiles via the conserved Cys6 residue3 Proteomics as tool to visualize reversible and irreversible thiol- modifications3.1. The thiol-redox proteome of B. subtilis in response to diamide and quinones3.2. Depletion of thiol-containing proteins by quinones due to thiol-(S)- alkylation4 Proteomics as tool to define regulon structures and targets for non- coding RNAs5 Acknowledgment15) Mass Spectrometry in the study of Tularemia Pathogenesis.Jiri Stulik, Juraj Lenco, Jiri Dresler, Jana Klimentova, Lenka Hernychova, Lucie Balonova and Alena Fucikova.15.1 Introduction to molecular pathogenesis of Francisella tularensis infection15.2 Francisella tularensis LVS proteome alterations induced by different temperatures and stationary phase of growth15.3Analysis of membrane protein complexes of Francisella tularensis15.4 Analysis of Francisella tularensis glycoproteins and phosphoproteins15.5Identification of Francisella tularensis transcription factors potentially involved in its virulence15.6 AcknowledgementsReferences16) Bacterial Post-Genomics for Vaccine developmentGiulia Bernardini, Daniela Braconi and Annalisa SantucciSummarycomparative genomicstranscriptomicsproteomics and immmunoproteomicsother high-throughput technologiesmeningococcal vaccines and reverse vaccinologyhelicobacter pylori vaccinesconclusionsreferences6 Statistical Analysis of 2D Gels and Analysis of Mass Spectral Data
- Machine Learning Techniques for the Analysis of Mass spectrometry Data.