Thalassiosira pseudonana Husedt (Hasle and Heimdal) clone 3H was grown in axenic culture in artificial seawater medium containing 10(-8) molar Na(2) (75)SeO(3). Biochemical distribution of radiolabeled Se was determined by solvent extraction techniques, gel filtration, and polyacrylamide gel electrophoresis. Of the total cellular Se, 51% was protein bound. Two soluble macromolecules of 21 and 29 kilodaltons contained (75)Se. These results are the first to provide evidence of specific Se-containing compounds in a photosynthetic organism. Glutathione peroxidase (GSH-Px) activity was measured in cell-free extracts and on nondenaturing polyacrylamide gels by a glutathione-reductase coupled assay. Two enzymes showing GSH-Px activity were present. One enzyme was active with H(2)O(2) and tert-butyl hydroperoxide (tBOOH); consistent with known Sedependent GSH-Pxs, but the other enzyme was only active with tBOOH. Co-migration of the H(2)O(2)-active GSH-Px and (75)Se on nondenaturing polyacrylamide gels provides evidence that T. pseudonana contains a Sedependent GSH-Px. The molecular weight of one of the (75)Se-labeled macromolecules is identical with the weight of previously characterized GSH-PX subunits. We conclude that the obligate requirement for Se in Thalassiosira pseudonana is due in part to the presence of the selenoenzyme glutathione peroxidase.
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