29] Technique for the identification of a biosynthetic precursor to parathyroid hormone

Abstract

Publisher Summary A biosynthetic precursor, proparathyroid hormone (proPTH), to parathyroid hormone (PTH) has been identified recently in bovine human, and, tentatively, in chick and rat parathyroid tissue. The prohormone from the bovine contains an additional hexapeptide sequence at the amino terminus of PTH. Four of the six additional amino acids are positively charged (1 arginine and 3 lysine residues), providing a ready means for the physical separation of the prohormone from PTH by gel electrophoresis or ion-exchange chromatography. The initial identification of the precursor was accomplished by electrophoretic analysis of the radioactive tissue proteins synthesized during short incubations of the tissue in vitro with radioactive amino acids (pulse labeling). This chapter describes the techniques that have been used to identify the prohormone in bovine parathyroid glands. Similar techniques have been used for the identification of biosynthetic precursors for other polypeptide hormones, for example, insulin and glueagon and for identification of precursors from human, chick, and rat parathyroids. The procedures described here include methods (1) for location of parathyroid glands in the bovine, (2) for demonstration of the precursor to product relationship between proPTH and PTH by in vitro pulse labeling of the parathyroid proteins, and (3) for analyses of the chemical properties of PTH and proPTH by use of selective labeling with radioactive amino acids followed by limited cleavage and eleetrophoretie separation of the labeled peptides.