In Silico Structural Prediction and Production of a Chimeric Recombinant Dickkopf1 (DKK1) Antigen

Dickkopf (DKK) family of proteins are known as antagonists for the Wnt #x3B2;catenin signaling pathway. It is suggested that the Dickkopf1 (DKK1) has a role in several diseases such as hepatocellular carcinomas, hepatoblastomas, Wilms #x2019; tumors, lung cancer and Myeloma bone disease. The aim of the present study was to produce a chimericrecombinant DKK1 protein in order to induce immune response against the antigen. The recombinant Dickkopf1 (rDKK1) protein was designed using bioinformatics analysis. The standard methods were used for cloning, expression and purification. The structure of recombinant protein was analyzed by spectroscopy methods. Enzymelinked immunosorbent assay (ELISA) and Western blotting were performed to confirm the recombinant protein using a commercial antiDKK1 (whole protein) polyclonal antibody. The immunogenicity #xA0;of the recombinant DKK1 was assessed by immunizing, intraperitoneally, BALB/C mice four times with the 31kDa and 45kDa purified rDKK1 cloned in pET28a and pET32a vectors respectively. The antibody titer was measured in due course of time. Stronger immunogenic parts of the protein were selected based on insilico predictions and recombinant protein was successfully designed. The chimeric gene was subcloned, expressed, purified and refolded. The purified protein was confirmed by Western blotting and ELISA. The three dimensional structural was confirmed by CD spectrum and predicted structures by bioinformatics tools, revealed the stability of helix structures. rDKK1 protein was capable of inducing immune response with high titer antibody and #xA0; excessive humoral immune response. No significant difference was observed between immunization by 31kDa and 45kDa antigen. #xD; #xA0;