Biosynthesis of lutropin in ovine pituitary slices: Incorporation of [ 35S]sulfate in carbohydrate units

Abstract

Sulfate incorporation into carbohydrate of lutropin (LH) has been studied in sheep pituitary slices using H 2 35SO 4. Labeled ovine LH was purified to homogeneity by Sephadex G-100 and carboxymethyl-Sephadex chromatography from both the incubation medium and tissue extract. Autoradiography of the gel showed only two protein bands which comigrated with the α and β subunits of ovine LH in both the purified ovine LH and the immunoprecipitate obtained with LH-specific rabbit antiserum. Furthermore, [ 35S]sulfate was also incorporated into several other proteins in addition to LH. The location of 35SO 4 2− in the oligosaccharides of ovine LH was evidenced by its presence in the glycopeptides obtained by exhaustive Pronase digestion. The location and the point of attachment of sulfate in the carbohydrate unit were established by the isolation of 4- O-[ 35S]sulfo- N-acetylhexosaminyl-glycerols and 4- O-[ 35S]sulfo- N-acetylglucosaminitol from the Smith degradation products and by the release of 35SO 4 2− by chondro-4-sulfatase. Thus, the present line of experimentation indicates the presence of sulfate on both the terminal N-acetylglucosamine and N-acetylgalactosamine in the oligosaccharide chains of the labeled ovine LH.