Integrated genome and proteome analysis of therapeutically important bacteriophages by combination of electrophoresis and mass spectrometry

It is expected that selected bacteriophages (bacterial viruses), or even their specific proteins and/or peptides can be used as a new effective means to treat bacterial infections. For a rational development of phage therapy, without the adverse effects of current antibiotics, the relationship between the desired biological properties of suitable phages and their genome/proteome structure must be found. To achieve this goal it is necessary to find new technologies for the rapid analysis of the phage molecular structure. In this project, we will develop a three-dimensional separation system for analysis and typing of the phage proteome. First, micropreparative capillary electrophoresis with laser-induced fluorescence detection (LIF) and noncovalent labeling will be used for the separation of the phage proteins. This fingerprint will be used to trace differences in the protein content of phages and to control the automated fraction collection. The resulting fractions will be digested by trypsin immobilized directly on the walls of the collection micro wells. In the final step, the tryptic peptides will be analyzed by capillary electrophoresis coupled on-line to mass spectrometry. To provide the peptide sequence and/or information on post-translational modifications both the electrospray with ion trap (ESI/IT) and matrix assisted laser desorption/ionization with time-of-flight (MALDI/TOF) mass spectrometers will be used as complementary tools. Phage 812 and its host-range mutants, which exhibit strong lytic activity and different host-specificity, have been chosen for this study. Most important results or innovations The development of a novel interface for the coupling of separation techniques with MALDI mass spectrometry Cooperation * Department of Genetics and Molecular Biology, MU, Brno * Laboratory of Mass Spectrometry of Biomolecules, MU, Brno * Institute of Analytical Chemistry of the Czech Academy of Science, Brno Research team * Jan Preisler preisler@chemi.muni.cz * Josef Havel * Pavel Krásenský Project duration 2003-2005 "Zdroj":[ http://bart.chemi.muni.cz/research.htm]

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