Master of Science (MS)
The mechanisms that contribute to bacterial ionizing radiation resistance are not completely understood. The Daly model uses Deinococcus radiodurans to suggest that bacterial ionizing radiation resistance is primarily determined by the amount of protein oxidation produced in response to ionizing radiation, so protein oxidation will be limited in radioresistant microorganisms due to a high intracellular level of manganese (Mn). This thesis investigates the Daly model by first introducing oxidative damage to Beta-Galactosdiase using increasing doses of gamma irradiation. Beta-Galactosidase activity was measured by the production of o-nitrophenol (ONP) which is produced from the hydrolysis of o-nitrophenyl-Beta-D-galactoside (ONPG). Beta-Galactosidase activity decreased with increasing doses of gamma irradiation. Beta-Galactosidase activity did not decrease when the protein wasirradiated in the presence of 50 mM Beta-mercaptoethanol or 50 mM mannitol and 50 mM benzoic acid, Beta-Galactosidase. This demonstrated that free radicals present in the cell could protect proteins. To determine if proteins could also shield proteins from free radicals, Beta-Galactosidase was irradiated with 150 mg/ml BSA which was about half the concentration of proteins in E. coli cell. Surprisingly, we found that BSA protected Beta-Galactosidase from ionizing radiation induced damage. This suggested that proteins could protect other proteins in the cell and would less likely to be targets of free radicals, so this could occur in all microorganisms. Therefore, protein oxidation would not be the primary determinant of ionizing radiation resistance. To determine if Mn was essential for ionizing radiation resistance, we irradiated stationary phase D. radiodurans R1 in a rich medium and a minimal salts medium in which other compounds were added. When stationary phase R1 cells were irradiated in M9 minimal media containing Mn, cell survival did not increase resistance and instead the cells showed an increase in sensitivity. When a carbon source was added to the M9 minimal media containing Mn, cell survival increased but not to the level of survival in rich medium. We concluded that Mn(II) was not essential for ionizing radiation resistance and appeared to be toxic to the cells.
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Robins, Jana Ronnette, "Manganese (II) and Protein Oxidation as Determinants of Bacterial Ionizing Radiation Resistance" (2011). LSU Master's Theses. 1331.