Purification and characterization of rat relaxin.

Two highly purified forms of relaxin, designated CM1 and CM2, were obtained from rat ovaries collected on day 20 of gestation. The isolation procedure consisted of aqueous extraction, followed by fractionation with Sephadex G-50 and ion exchange chromatography. The yields of CM1 and CM2 were approximately 140 microgram/geq ovarian fresh tissue. No difference in biological potency between CM1 and CM2 was found when they were bioassayed with mouse pubic symphysis bioassays. Physicochemical analyses indicated that CM1 and CM2 were similar but not identical. The molecular weights of CM1 and CM2 were approximately 6000, as determined by ultracentrifugation. Analytical acrylamide disc gel electrophoresis at pH 4.3 demonstrated that CM1 and CM2 had different electrophoretic mobilities. Electrofocusing indicated the isoelectric points of CM1 and CM2 were pH 7.6 and pH 9.4, respectively. The amino acid compositions of CM1 and CM2 were similar but not identical. Slab gel electrophoresis in polyacrylamide gel with sodium dodecyl sulfate showed that both reduced rat relaxin and reduced porcine relaxin migrated farther than their unreduced forms. This observation supports the view that rat relaxin, like porcine relaxin, consists of two chains linked by disulfide bonds.