Expression, purification, and biological characterization of recombinant human interleukin-31 protein.

Interleukin-31 (IL-31), belonging to the IL-6 cytokine family, is involved in skin inflammation and pruritus, as well as some tumors' progression. Here, we reported the expression and purification of recombinant human IL-31 (rhIL-31) using a prokaryotic system. This recombinant protein was expressed in the form of inclusion bodies, refolded and purified by size-exclusion chromatography. Circular dichroism analysis revealed that the secondary structure of rhIL-31 was mainly composed of alpha-helix, which is in consistence with the 3D model structure built by AlphaFold server. In vitro studies showed that rhIL-31 exhibited a good binding ability to the recombinant hIL-31 receptor alpha fused with human Fc fragment (rhIL-31RA-hFc) with EC 50 value of 16.36 µg/mL in ELISA assay. Meanwhile, flow cytometry demonstrated that rhIL-31 was able to bind to hIL-31RA or hOSMRβ expressed on the cell surface, independently. Furthermore, rhIL-31 could induce the phosphorylation of STAT3 in A549 cells. In conclusion, the prepared rhIL-31 in this study possesses the binding ability to its receptors, and can activate the signal pathway of JAK/STAT. Thus, it can be applied in further studies, including investigation of hIL-31-related diseases, structural analysis, and development of therapeutic drugs, and monoclonal antibodies targeting hIL-31.