Design, Expression, and Binding Interactions Study of the Recombinant Engineered IL-6R

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Soluble interleukin-6 receptor (IL-6R) trans-signaling is implicated in chronic inflammatory diseases, autoimmune disorders, and cancers.
• Targeting the IL-6R-gp130 interaction is a key strategy for managing these IL-6-mediated pathologies.

Purpose of the Study:

• To computationally model and engineer a fragment of IL-6R (residues 121-300) to reduce its binding affinity to glycoprotein 130 (gp130).
• To express and characterize the engineered IL-6R fragment (seIL-6R) for potential therapeutic applications.

Main Methods:

• Protein engineering and molecular modeling using AlphaFold and ClusPro for docking simulations.
• Recombinant expression of seIL-6R in Chinese hamster ovary (CHO)-K1 cells.
• Biochemical and biophysical characterization including Western blotting and Fourier transform infrared (FTIR) spectroscopy.

Main Results:

• The engineered seIL-6R fragment exhibited improved stability, solubility, and a reduced molecular weight (20.6 kDa).
• Molecular docking predicted a diminished binding interaction between seIL-6R and gp130.
• Western blotting confirmed successful recombinant protein expression, and FTIR analysis verified the preservation of secondary structure.

Conclusions:

• The engineered seIL-6R protein demonstrates favorable physicochemical properties and reduced binding to gp130.
• This engineered protein represents a promising candidate for further development as a therapeutic agent against IL-6-driven diseases.