Note: Intro -- Proteomics of Microbial Pathogens -- Table of Contents -- Preface Proteomics of microbial pathogens -- 1 Genome and proteome analysis of Chlamydia -- 1.1 Introduction -- 1.1.1 Chlamydia biology -- 1.1.1.1 Diseases -- 1.1.1.2 The developmental cycle -- 1.2 Chlamydia genomes -- 1.2.1 Sequenced Chlamydia genomes -- 1.2.2 Chlamydial genes -- 1.2.3 Genome comparison -- 1.3 Proteome analysis of Chlamydia -- 1.3.1 Early Chlamydia proteome studies -- 1.3.2 C. trachomatis proteome studies -- 1.3.3 C. pneumoniae proteome studies -- 1.3.4 Identification of secreted proteins by comparative proteomics -- 1.3.5 Proteome studies of comc -- 1.3.6 Proteome comparison of S. trachomatis serovars -- 1.3.7 Proteome analysis of growth conditions -- 1.3.8 Considerations in proteomics -- 1.4 Concluding remarks -- 2 Helicobacter pylori vaccine development based on combined subproteome analysis -- 2.1 Introduction -- 2.2 Classical whole-cell inactivated Helicobacter vaccines -- 2.3 Subunit Helicobacter vaccines: Conventional antigen selection -- 2.4 Subunit Helicobacter vaccines: Global antigen selection based on proteomics -- 2.4.1 Proteomics as a tool for antigen characterization -- 2.4.2 The Helicobacter proteome -- 2.4.3 Criteria for promising antigen candidates -- 2.4.4 Identification of protective antigens based on multiple criteria -- 2.5 Concluding remarks -- 3 Towards a comprehensive understanding of Bacillus subtilis cell physiology by physiological proteomics -- 3.1 Introduction -- 3.2 Subproteomes vs. the total theoretical proteome -- 3.3 The vegetative proteome of growing cells -- 3.4 Proteomes of nongrowing cells - the adaptational network -- 3.5 Proteomic signatures - tools for microbial physiology and their practical application -- 3.6 Transcriptomics vs. proteomics - towards a second generation of proteomics -- 3.7 The interactome. , 3.8 The secretome -- 3.9 Post-translational modifications -- 3.10 Protein quality control/protein degradation at a proteomic scale -- 3.11 Gene expression network - horizontal and vertical approach -- 3.12 Concluding remarks -- 4 Web-accessible proteome databases for microbial research -- 4.1 Introduction -- 4.2 Materials and methods -- 4.2.1 Data generation and data storage -- 4.2.2 Software tools -- 4.3 Results and discussion -- 4.3.1 Data management, analysis and presentation -- 4.3.2 2D-PAGE database -- 4.3.3 ICAT-LC/MS database -- 4.3.4 FUNC_CLASS database -- 4.3.5 Data analysis and visualization -- 4.4 Concluding remarks -- 5 A targeted proteomics approach to the rapid identification of bacterial cell mixtures by matrix-assisted laser desorption/ionization mass spectrometry -- 5.1 Introduction -- 5.2 Materials and methods -- 5.2.1 Chemicals -- 5.2.2 Bacillus strains -- 5.2.3 Vegetative cell digestion -- 5.2.4 MALDI-TOF MS and unimolecular decomposition product ion analysis -- 5.2.5 Database searches and identification of Bacillus species -- 5.3 Results and discussion -- 5.3.1 On-probe tryptic digestion of bacterial cells -- 5.3.1.1 Bacillus subtilis 168 -- 5.3.1.2 Bacillus globigii and sphaericus 14577 -- 5.3.1.3 Bacillus cereus Tand anthracis Sterne -- 5.3.2 Partial sequencing for rapid identification of bacterial cells -- 5.3.2.1 Unimolecular decomposition product ion analysis (UDPIA) -- 5.3.2.2 Identification of bacteria -- 5.3.3 Identification of proteins and protein-families in bacterial cell digests -- 5.3.3.1 Flagellin and surface layer protein precursor -- 5.3.3.2 Cold shock and cold shock-like proteins -- 5.3.3.3 Ribosomal proteins -- 5.3.3.4 DNA-binding proteins -- 5.3.3.5 Heat shock proteins -- 5.3.3.6 Other stress related proteins and the prosthetic group of an acyl-carrier protein. , 5.3.4 Analysis of a 1:1 mixture of B. globigii and B. sphaericus 14577 -- 5.4 Concluding remarks -- 6 Protein identification and tracking in two-dimensional electrophoretic gels by minimal protein identifiers -- 6.1 Introduction -- 6.2 Materials and methods -- 6.2.1 Materials -- 6.2.2 Two-dimensional electrophoresis -- 6.2.3 Mass spectrometry -- 6.2.4 Compilation of a theoretical dataset comprising all tryptic peptides of M. tuberculosis H37Rv -- 6.2.5 Comparison of MALDI spectra by the program MS-Screener -- 6.2.6 Determination of exogenous contaminant masses -- 6.2.7 Generation of template spectra -- 6.3 Results and discussion -- 6.3.1 Proteome analysis of M. tuberculosis H37Rv CSN -- 6.3.2 Determination of exogenous contaminant masses by MS-Screener -- 6.3.3 The MPI approach revealed HspX-specific peptide masses in multiple spectra -- 6.3.4 The MS-Screener analysis revealed truncated variants of Tuf previously not identified by PMF -- 6.3.5 Frequency of tryptic peptides with similar m/z ratios in M. tuberculosis H37Rv -- 6.3.6 In mass spectrometry it is important to consider detection probabilities of proteins and peptides -- 6.4 Concluding remarks -- 7 Continued proteomic analysis of Mycobacterium leprae subcellular fractions -- 7.1 Introduction -- 7.2 Materials and methods -- 7.2.1 Preparation and separation of M. leprae proteins -- 7.2.2 MS -- 7.2.3 Isolation of basic proteins of M. leprae -- 7.3 Results and discussion -- 7.4 Concluding remarks -- 8 CFP10 discriminates between nonacetylated and acetylated ESAT-6 of Mycobacterium tuberculosis by differential interaction -- 8.1 Introduction -- 8.2 Materials and methods -- 8.2.1 Protein samples -- 8.2.2 2-DE blot overlay -- 8.2.3 Mass spectrometry -- 8.3 Results -- 8.3.1 High resolution separation of acidic ST-CF proteins -- 8.3.2 Mass analysis of ESAT-6 spots. , 8.3.3 Interaction of recombinant CFP10 with ESAT-6 spots -- 8.4 Discussion -- 9 The cell wall subproteome of Listeria monocytogenes -- 9.1 Introduction -- 9.2 Material and methods -- 9.2.1 Bacterial strain and growth conditions -- 9.2.2 Serial extraction of cell wall proteins -- 9.2.3 Aminopeptidase C assay -- 9.2.4 SDS-PAGE Western blotting, and N-terminal sequencing -- 9.2.5 2-D-PAGE -- 9.2.6 Gel staining and protein identification by mass spectrometry -- 9.2.7 Immunoelectron microscopy -- 9.2.7.1 Postembedding labeling studies -- 9.2.7.2 Field emission scanning electron microscopic immunolabeling -- 9.2.8 Overlay blot -- 9.2.9 Ligand fishing -- 9.2.10 Cloning and purification of proteins -- 9.2.11 Kinetic analyses using SPR detection -- 9.2.12 Bioinformatic analysis -- 9.3 Results -- 9.3.1 Prediction and validation of cell wall-associated proteins -- 9.3.1.1 Prediction of exported proteins -- 9.3.1.2 Validation of the cell wall subproteome -- 9.3.2 Analysis of protein processing and localization -- 9.3.3 Analysis of plasminogen-binding proteins -- 9.4 Discussion -- 9.4.1 Analysis of proteins identified from surface extracts -- 9.4.2 Analysis of protein processing and localization in surface extracts -- 9.4.3 Possible function of plasminogen binding in virulence -- 9.5 Concluding remarks -- 10 Low virulent strains of Candida albicans: Unravelling the antigens for a future vaccine -- 10.1 Introduction -- 10.2 Materials and methods -- 10.2.1 Microorganism and culture conditions -- 10.2.2 Mice -- 10.2.3 Systemic infection conditions and generation of immune sera -- 10.2.4 Tcell purification and passive immunization -- 10.2.5 2-DE -- 10.2.5.1 Protoplast lysate preparation -- 10.2.5.2 Analytical and micropreparative 2-DE -- 10.2.6 Immunoblot analyses -- 10.2.7 MALDI-TOF and MALDI-TOF MS analyses of spots -- 10.2.8 Database search -- 10.3 Results. , 10.3.1 Vaccination assays with different mutant strains and generation of immune sera -- 10.3.2 Importance of cellular immunity in vaccination with C. albicans CNC13 -- 10.3.2.1 Role of Th1/Th2 cytokines in CNC13 immune response -- 10.3.2.2 Protection induced by passive transfer of sensitized CNC13 lymphocytes -- 10.3.3 Profile of C. albicans immunoreactive proteins in the different mutant strains -- 10.3.3.1 Detection and identification of the immunoreactive proteins -- 10.4 Discussion -- 10.4.1 Low virulent C. albicans strains as a tool to study the host immune response -- 10.4.2 C. albicans hog1 mutant induces protection in a vaccination assay -- 10.4.3 C. albicans new antigenic proteins -- 10.4.4 Antibody profile linked to successful vaccination against systemic candidiasis -- 10.5 Concluding remarks -- 11 Proteomic analysis of the sarcosine-insoluble outer membrane fraction of the bacterial pathogen Bartonella henselae -- 11.1 Introduction -- 11.2 Materials and methods -- 11.2.1 Strains and culture conditions -- 11.2.2 Enrichment of B. henselae OMPs -- 11.2.3 Protease exposure -- 11.2.4 1-D SDS-PAGE -- 11.2.5 Protein solubilization and protein quantitation -- 11.2.6 2-D NEPHGE -- 11.2.7 MALDI-TOF-MS -- 11.2.8 Database query -- 11.2.9 In silico analysis -- 11.3 Results -- 11.3.1 Enrichment of B. henselae OMPs -- 11.3.2 1-D SDS-PAGE of B. henselae OMPs and protein assignment by PMF -- 11.3.3 2-D NEPHGE of B. henselae OMPs and protein assignment by PMF -- 11.4 Discussion -- 12 The influence of agr and σ(B) in growth phase dependent regulation of virulence factors in Staphylococcus aureus -- 12.1 Introduction -- 12.2 Material and methods -- 12.2.1 Bacterial strains and culture conditions -- 12.2.2 Preparation of the extracellular protein fraction -- 12.2.3 Analytical and preparative PAGE -- 12.2.4 Quantitation of protein spots. , 12.2.5 Transcriptional analyses.