Investigation of Biological Systems Using Synchrotron X-ray Light Sources.

Abstract

In this thesis metals were studied in several biological systems using XRF microscopy and XAS spectroscopy. In chapter two, I examined the budding yeast Sacchromyces cervisiae using XRF microscopy. Using fluorescent stains to identify the vacuole and the nucleus, we were able to determine which elements localized where. We found that phosphorus fluorescence was a reasonable surrogate for the nucleus of the cell. Zinc and iron localized to both the vacuole and the nucleus. Sulfur and potassium typically have smooth distributions, and calcium and manganese localized in the vacuole. These results establish the groundwork for the subcellular elemental distributions of yeast grown under normal conditions, allowing for the later exploration of the subcellular localizations of elements under other conditions. In chapter three, we examined the effects of cadmium treatment on yeast. The cadmium-treated yeast showed dramatic localizations of copper in the bud and bud neck. The other elements did not show any significant changes in localization compared to the yeast that had not been exposed to cadmium. While iron and zinc did not change their localizations, they did show a significant increase in concentration. X-ray absorption spectroscopy of zinc in yeast cell paste treated with cadmium shows a shift towards sulfur ligation compared to cell paste that was not treated with cadmium. These results suggest that the cellular damage caused by cadmium is a result of an increase in the cell’s free copper, leading to increased oxidative stress. Chapter 4 uses XRF microscopy to examine human erythrocytes that have been infected with the malaria parasite. Infected erythrocytes were found to accumulate zinc 3-6 fold in excess when compared to uninfected cells. This accumulated zinc was found to be localized close to, but not overlapping with, the iron localization of the hemozoin crystal. In addition to the increase in zinc, phosphorus, sulfur and potassium were found to increase substantially, while iron showed a small but likely significant increase upon infection.