Korol N. Characteristics of the genome structural region of temperate bacteriophage ZF40 Pecobacterium carotovorum

The thesis is devoted to the study of peculiarities of virion morphogenetic development, identification of major structural proteins and analysis of the genome structural region organization of temperate bacteriophage ZF40 Pecobacterium carotovorum subsp. carotovorum (Рсс). In the course of the study it was determined that Рсс M2-4/50RI is the most adequate indicator strain for lytic growth of bacteriphage ZF40 and it virulent variants. It allowed obtaining stable phage particles resistant to osmotic shock. At the same time phage propagation on traditional bacterial culture Pcc RC5297 leads to an abortive infection, which is accompanied by accumulation of separate virion structural components. It was determined that the nature of phage populational heterogeneity lies in the violation of the key stages of virion morphogenesis and morphopoiesis. These peculiarities have a negative impact on stability of phage particles. An alternative methodological approach for obtaining highly concentrated samples of phage structural components was developed based on classic phage Т4 and its amber-mutant Т4am23(Н11). Concentrated preparations of capsids and tails of bacteriophage ZF40 were obtained using the mentioned technique. Protein content of native particles and their components was determined. A detailed analysis for all phage structural proteins was carried out by tandem mass-spectrometry technique. The major structural proteins and their coding genes were identified: gp46 – major capsid protein (33,95 kDa), gp52 – major sheath protein (39,2 kDa) and gp50 – hypothetical tail tube protein (18,3 kDa). It was proved that the major capsid protein is not exposed to considerable post-translational modification. A detailed map of the structural region of bacteriophage ZF40 was constructed. It is characterized by classic organization that is appropriate of most Myoviridae phages. It also correlates with the analogous regions in genomes of phages belonging to the PLPE group. Key words: Рectobacterium carotovorum, temperate bacteriophage ZF40, genome, morphogenesis, populational heterogeneity, structural region, structural proteins, major capsid protein, phage tail, chromatography, ESI-MS/MS.