Chemical synthesis and biological activity of ovine Insulin 3, a relaxin structural homologue

Abstract

Relaxin, a two-chain, three disulfide-bonded peptide, is produced primarily by the ovary during pregnancy. Its principle roles in many mammalian species include softening the cervix and remodelling the interpubic ligaments in preparation for birth. No relaxin peptides have been identified in ruminants, despite target tissues of these animals being responsive to exogenous relaxin from other species [Bathgate et al., this volume]. A peptide isolated from the Leydig cells in pig testis was shown to possess striking structural similarity to relaxin and which is also expressed in the ovary [1]. The primary structure of sheep Leydig cell insulin-like peptide, or INSL3, was determined from isolated mRNA from sheep ovary and shown to consist of a 26-residue A-chain, a 32-residue B-chain and three disulfide bonds in the same disposition as for relaxin and insulin [2]. We undertook the chemical synthesis of this native peptide and two INSL3 analogues, each analogue containing modifications to their B-chains to more closely mimic the human relaxin Gene 2 (H2) B-chain (Fig 1). After comprehensive chemical characterization, we tested all three peptides for any relaxin-like biological activity.