Proteomics and its Application to the Human-Pathogenic Fungi Aspergillus fumigatus and Candida albicans
The number of opportunistic fungal infections has increased significantly during the past decades, at least in part as the result of a rising number of immunocompromised patients. Individuals at risk for the development of a serious fungal infection inclu
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CONTENTS I. Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . II. Methods of Proteome Analysis . . . . . . . . . . III. 2D Gel Electrophoresis . . . . . . . . . . . . . . . . . A. Evolution of the Technology . . . . . . . . . B. Sample Preparation . . . . . . . . . . . . . . . . . C. First Dimension: Isoelectric Focusing . . . . . . . . . . . . . . . . . . . . . . . . . . D. Second Dimension: SDS-PAGE . . . . . . . E. Detection of Proteins. . . . . . . . . . . . . . . . F. Scanning and Image Analysis . . . . . . . . G. Tryptic Digestion and Mass Spectrometry Identification . . . . . . . . . . H. Protein Identification by Mass Spectrometry . . . . . . . . . . . . . . . . . . . . . . J. Peptide Mass Fingerprint and Tandem-Mass Spectrometry . . . . . . . . . IV. Gel-Independent Techniques. . . . . . . . . . . . A. Liquid Chromatography-Based MS Techniques . . . . . . . . . . . . . . . . . . . . . B. Quantitative Proteomics Using LC-MS. . . . . . . . . . . . . . . . . . . . . . . V. Sample Fractionation to Identify Low-Abundant Proteins . . . . . . . . . . . . . . . . A. Subcellular Fractionation . . . . . . . . . . . . B. Membrane Proteomics . . . . . . . . . . . . . . C. Phosphoproteomics. . . . . . . . . . . . . . . . . 1. Detection of Phosphoproteins with 2D-PAGE . . . . . . . . . . . . . . . . . . . 2. Enrichment of Phosphoproteins. . . . 3. Chromatographic Methods for Phosphoproteomics . . . . . . . . . . . VI. Standard Development . . . . . . . . . . . . . . . . . VII. Proteomics of Human-Pathogenic Fungi . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . A. Sample Preparation . . . . . . . . . . . . . . . . . B. Proteomics Studies of Fungal Physiology. . . . . . . . . . . . . . . . . . . . . . . . . C. Drug-Induced Changes. . . . . . . . . . . . . . D. Biofilm Formation . . . . . . . . . . . . . . . . . . E. Proteomics of the Cell Wall . . . . . . . . . . F. Immunoproteomics. . . . . . . . . . . . . . . . . G. Secretome . . . . . . . . . . . . . . . . . . . . . . . . . H. Virulence Factors. . . . . . . . . . . . . . . . . . .
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1 Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology– Hans Knöll Institute, Beutenbergstrasse 11a, 07745 Jena; and Friedrich Schiller University, Jena, Germany; e-mail: Olaf.Kniemeyer@ hki-jena.de
J. Host–Pathogen Interaction . . . . . . . . . . VIII. Conclusion: Towards Integrative Analysis–Systems Biology . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . .
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I. Introduction The number of opportunistic fungal infections has increased significantly during the past decades, at least in part as the result of a rising number of immunocompromised patients. Individuals at risk for the development of a serious fungal infection include patients undergoing solid-organ, blood and bone marrow transplantation, cancer patient
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