Isolation and bioactivity of relaxin from the tammar wallaby (Macropus eugenii)

Abstract

Recent cloning of a prepro-relaxin cDNA from the tammar wallaby (Macropus eugenii)[1] indicated that the derived amino acid sequence retains the structural elements common to relaxin peptides from all species, including the three residues that make up the putative receptor binding motif (Fig.1). Although neither marsupial prorelaxin nor relaxin have been extracted and purified from any tissue, early studies demonstrated that crude extracts of corpora lutea (CL) from the pregnant tammar wallaby contain bioactive relaxin [2]. Both the CL and placenta are sites of relaxin mRNA expression during pregnancy in the tammar but transcripts are considerably more abundant in the CL [1]. Within the CL, relaxin gene expression is highest in early and mid-pregnancy, reduced at term and absent postpartum [1]. Antisera to porcine relaxin was used to show immunoreactive relaxin localized to membrane-bound, electron-dense granules in the luteal cell cytoplasm [3]. A large increase in immunoreactive relaxin in the tammar CL is observed from Day 22 of the 26-day gestation but does not coincide with the increase in relaxin gene expression [1]. Moreover, electron-dense granules, the known storage site for relaxin, are not present in the luteal cell cytoplasm in large numbers until Day 10 of gestation [1]. It was suggested that prorelaxin is stored in cytoplasmic granules in the CL during the early stages of gestation and that further processing to form mature relaxin peptide occurs towards the end of gestation when the hormone is released. The objective of this study was to isolate and purify tammar prorelaxin or processed relaxin peptides from the tammar wallaby CL and determine their structure and bioactivity.