Abstract

The murine double minute 2 (MDM2) protein is a major negative regulator of the tumour suppressor protein p53. Under normal conditions, MDM2 constantly binds to p53 transactivation domain and/or ubiquinates p53 via its role as E3 ubiquitin ligase to promote p53 degradation as well as nuclear export to maintain p53 levels in cells. Meanwhile, amplification of MDM2 and appearance of MDM2 spliced variants occur in many tumours and normal tissues making it a prognostic indicator for human cancers. The mutation or deletion of p53 protein in half of human cancers inactivates its tumour suppressor activity. However, cancers with wild type p53 have its function effectively inhibited through direct interaction with MDM2 oncoprotein. Here, we described the construction of a MDM2 spliced variant (rMDM215kDa) consisting of SWIB/MDM2 domain and its central region for antibody generation. Biopanning with a human naïve scFv library generated four scFv clones specific to rMDM215kDa. Additionally, the selected scFv clones were able to bind to the recombinant full length MDM2 (rMDM2-FL). Computational prediction showed that the selected scFv clones potentially bind to exon 7–8 of MDM2 while leaving the MDM2/SWIB domain free for p53 interaction. The developed antibodies exhibit good specificity can be further investigated for downstream biomedical and research applications.

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