b ABSTRACTBoth prokaryotic and eukaryotic organisms possess mechanisms for the detoxification of heavy metals, and these mechanisms are found among distantly related species. We investigated the role of intracellular glutathione (GSH), which, in a large number of taxa, plays a role in protection against the toxicity of common heavy metals. Anaerobically grown Lactococcus lactis containing an inducible GSH synthesis pathway was used as a model organism. Its physiological condition allowed study of putative GSH-dependent uranyl detoxification mechanisms without interference from additional reactive oxygen species. By microcalorimetric measurements of metabolic heat during cultivation, it was shown that intracellular GSH attenuates the toxicity of uranium at a concentration in the range of 10 to 150 M. In this concentration range, no effect was observed with copper, which was used as a reference for redox metal toxicity. At higher copper concentrations, GSH aggravated metal toxicity. Isothermal titration calorimetry revealed the endothermic binding of U(VI) to the carboxyl group(s) of GSH rather than to the reducing thiol group involved in copper interactions. The data indicate that the primary detoxifying mechanism is the intracellular sequestration of carboxyl-coordinated U(VI) into an insoluble complex with GSH. The opposite effects on uranyl and on copper toxicity can be related to the difference in coordination chemistry of the respective metal-GSH complexes, which cause distinct growth phase-specific effects on enzyme-metal interactions. IMPORTANCEUnderstanding microbial metal resistance is of particular importance for bioremediation, where microorganisms are employed for the removal of heavy metals from the environment. This strategy is increasingly being considered for uranium. However, little is known about the molecular mechanisms of uranyl detoxification. Existing studies of different taxa show little systematics but hint at a role of glutathione (GSH). Previous work could not unequivocally demonstrate a GSH function in decreasing the presumed uranyl-induced oxidative stress, nor could a redox-independent detoxifying action of GSH be identified. Combining metabolic calorimetry with cell number-based assays and genetics analysis enables a novel and general approach to quantify toxicity and relate it to molecular mechanisms. The results show that GSH-expressing microorganisms appear advantageous for uranyl bioremediation.T he natural abundance of uranium and the increasing health risks that originate from its anthropogenic release to the environment have generated a high demand for understanding the molecular mechanisms of uranium toxicity. Its radiotoxicity is of minor ecological relevance compared to its toxicity due to chemical interference with metabolism (1, 2). The latter is caused mostly by the displacement of Ca 2ϩ and Mg 2ϩ from functionally important binding sites in enzymes (3). The water-soluble uranyl cation UO 2 2ϩ is the most commonly found contaminant, which contains uranium in its highest ...

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The performance of a PIN silicon photodiode as a microdosimetric detector was explored. Microdosimetric spectra of a 137Cs gamma source and that of an americium-beryllium neutron source were determined with a PIN photodiode as well as with a tissue-equivalent proportional counter of 1 micron simulated diameter. The spectra were compared.

Studying the toxicity of chemical compounds using isothermal microcalorimetry (IMC), which monitors the metabolic heat from living microorganisms, is a rapidly expanding field. The unprecedented sensitivity of IMC is particularly attractive for studies at low levels of stressors, where lethality-based data are inadequate. We have revealed via IMC the effect of low dose rates from radioactive β−-decay on bacterial metabolism. The low dose rate regime (<400 µGyh−1) is typical of radioactively contaminated environmental sites, where chemical toxicity and radioactivity-mediated effects coexist without a predominance or specific characteristic of either of them. We found that IMC allows distinguishing the two sources of metabolic interference on the basis of “isotope-editing” and advanced thermogram analyses. The stable and radioactive europium isotopes 153Eu and 152Eu, respectively, were employed in monitoring Lactococcus lactis cultures via IMC. β−-emission (electrons) was found to increase initial culture growth by increased nutrient uptake efficiency, which compensates for a reduced maximal cell division rate. Direct adsorption of the radionuclide to the biomass, revealed by mass spectrometry, is critical for both the initial stress response and the “dilution” of radioactivity-mediated damage at later culture stages, which are dominated by the chemical toxicity of Eu.