Rapid catabolism of tyrosine-O-sulphated proteins and the formation of free tyrosine O-sulphate as an end product in rat embryo fibroblasts

Abstract

Rat embryo fibroblasts, line 3Y1, were prelabelled for 24 h with [35S]sulphate and incubated in fresh medium without [35S]sulphate. A rapid efflux of the overall 35S-labelled compounds from the cells into the medium was observed. After 9 h of incubation, about 50% of the total 35S radioactivity appeared in the medium and up to 84.3% did so at the end of a 48 h incubation. Determination of [35S]sulphated macromolecules present in both the cell-associated and the incubation-medium fractions at different time points during incubation indicated that the majority of the 35S-labelled compounds released from the cells were low-Mr products derived from digestion of the [35S]sulphated macromolecules. Further analysis for tyrosine-O-[35S]sulphated proteins, which constituted only a small fraction of the overall [35S]sulphated macromolecules, showed that, after 9 h of incubation, there was a 65% decrease in the cell-associated fraction, and only 16.4% remained after 48 h. During that time, an amount equivalent to 20.7% of the cell-associated tyrosine-O-[35S]sulphated proteins originally present was released into the medium. Free tyrosine O-[35S]sulphate was generated in the cells and excreted into the incubation medium. Its rate of increase with time, however, was slow, and could account for only 12.4% of the tyrosine-O-[35S]sulphated proteins catabolized at the end of the 48 h incubation.