Genetic and biochemical studies of the lipid-containing bacteriophage PR4

Bacteriophage PR4 is a lipid-containing bacterial virus able to infect Escherichia coli and Salmonella typhimurium. The icosahedral virion consists of an external protein capsid layer which surrounds a membrane vesicle enclosed ds DNA genome.I have analyzed the time course of phage PR4 protein synthesis and have identified at least 34 proteins present in phage infected cells not detected in uninfected control cultures. In addition, I have isolated a more extensive set of conditional-lethal nonsense mutants of this virus. This collection of mutants permitted the identification of seven additional phage PR4 gene products, including the terminal genome protein and an accessory lytic factor. The present collection of phage PR4 mutants has been assigned to 19 distinct genetic groups on the basis of genetic complementation tests and sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of the proteins produced in mutant infected UV irradiated cells.A restriction endonuclease map of the phage PR4 genome was constructed which includes 59 sites for ten restriction endonucleases. In addition, I have constructed a collection of recombinant plasmids containing subgenomic DNA fragments of bacteriophage PR4. I have used this collection of plasmids to generate a physical-genetic map of the PR4 genome. The physical-genetic map localizes mutations in 13 phage PR4 genetic groups on the viral DNA molecule.To investigate the role of phosphatidylglycerol (PG) in phage assembly and infectivity, I propagated PR4 on an E. coli mutant defective in PG synthesis. The PG content of phage PR4 grown on the mutant host accounted for 0.4% of the total viral phospholipids, representing a 90-fold decrease in PG relative to the PG content of phage grown on a wild type host. This drastic alteration of the phage phospholipid composition had little or no effect on either the stability or infectivity of the phage.U of I OnlyETDs are only available to UIUC Users without author permissio