Distribution and HPLC study of chromium-51 binding sites in Chinese hamster ovary cells

Chinese hamster ovary cells were cultured with chromium-51 chromate to study the site of chromium interaction with cell biomolecules. After incubation, cells were homogenized and separated into nuclear, mitochondrial, microsomal, and cytosolic fractions. Greater than 75% of the radioactivity was found in the cytosolic fraction. The supernatant from the centrifuged cell homogenate, which contained greater than 90% of the chromium radioactivity, was subjected to chromatographic investigation. The combination of anion exchange and ion-pair high-performance liquid chromatography (HPLC) showed that at least three different molecular species interact with chromate or its reduced derivative, Cr(III). These species are glutathione, the nucleotides cytosine triphosphate, uridine triphosphate, guanine triphosphate, adenosine triphosphate, and adenosine diphosphate, plus an as yet unknown species of protein or peptide. Preliminary data for the specific activity of nucleoside triphosphates range from 6000 to 18,000 cpm/micrograms ribonucleoside triphosphate. The glutathione accounted for 50% of the observed radioactivity, the nucleotides for 30%, and the metalloprotein accounted for the remainder.