Seleno-relaxin analogues: effect of internal and external diselenide bonds on the foldability and a fibrosis-related factor of endometriotic stromal cells

Human relaxin-2 (H2 relaxin) is a peptide hormone of about 6 kDa, first identified as a reproductive hormone involved in vasoregulation during pregnancy. It has recently attracted strong interest because of its diverse functions, including anti-inflammatory, anti-fibrotic, and vasodilatory, and has been suggested as a potential peptide-based drug candidate for a variety of diseases. Mature H2 relaxin is constituted by the A- and B-chains stabilized by two interchain disulfide (SS) bridges and one intrachain SS linkage. In this study, seleno-relaxins, SeRlx-α and SeRlx-β, which are [C11UA,C11UB] and [C10UA,C15UA] variants of H2 relaxin, respectively, were synthesized via a one-pot oxidative chain assembly (folding) from the component A- and B-chains. The substitution of SS bonds in a protein with their analogue, diselenide (SeSe) bonds, has been shown to alter the physical, chemical, and physiological properties of the protein. The surface SeSe bond (U11A–U11B) enhanced the yield of chain assembly while the internal SeSe bond (U10A–U15A) improved the reaction rate of the folding, indicating that these bridges play a major role in controlling the thermodynamics and kinetics, respectively, of the folding mechanism. Furthermore, SeRlx-α and SeRlx-β effectively reduced the expression of a tissue fibrosis-related factor in human endometriotic stromal cells. Thus, the findings of this study indicate that the S-to-Se substitution strategy not only enhances the foldability of relaxin, but also provides new guidance for the development of novel relaxin formulations for endometriosis treatment.