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Recombinant Production of Interleukin-1 Receptor Antagonist in Fusion to Albumin Binding Domain With Potential Affinity to Human Serum Albumin Publisher



Shafiee F1, 2 ; Yazdani A1
Authors
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Authors Affiliations
  1. 1. Department of Pharmaceutical Biotechnology, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, Iran
  2. 2. Bioinformatics Research Center, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, Iran

Source: Research in Pharmaceutical Sciences Published:2024


Abstract

Background and purpose: Anakinra must be injected daily due to its short half-life and this leads to lower patient compliance. Therefore, the aim of this study was to produce an interleukin-1 receptor antagonist (IL-1Ra) with albumin binding domain (ABD) as a novel fusion protein and evaluate its binding ability to albumin and its biological effects. Experimental approach: The three-dimensional structure of IL-1Ra-ABD was predicted by MODELLER software and its interaction with IL-1R was evaluated by the HADDOCK server. The expression of IL-1Ra-ABD was performed in E. coli in fusion with intein 1 of pTWIN1 in soluble form and then purified. The affinity of IL-1Ra-ABD to human serum albumin (HSA) was determined on native-PAGE, and its release percent toward time was evaluated. Moreover, an MTT assay was used to determine the antagonizing properties of recombinant IL-1Ra-ABD against IL-1β in A375 and HEK293 cell lines. Findings/Results: The stable complex of IL-1Ra-ABD with IL-1R established the absence of steric hindrance due to the addition of ABD to IL-1Ra. The expression induction of intein 1-IL-1Ra-ABD using 0.1 mM IPTG at 15 °C, and its cleavage represented bands approximately in 50 and 23 kDa. Furthermore, about 78% of IL-1Ra-ABD was attached to the HSA after 2 h of incubation, and the MTT assay showed no significant differences between the effects of IL-1Ra-ABD and native IL-1Ra in cell survival. Conclusions and implications: The production of soluble IL-1Ra-ABD with no significant differences in IL-1Ra antagonizing effects was successfully performed. IL-1Ra-ABD showed suitable interaction with HSA and was released over time. However, the half-life of IL-1Ra-ABD in vivo must be determined in the subsequent investigations. © 2024 Research in Pharmaceutical Sciences.