Applying the Bioinformatics Methods to Design and Evaluate the SapM-M13 pIX Fusion Protein and Its Theoretical Role in the Phage ELISA System

Phage ELISA is a common method used to confirm binding of obtained phages from phage display technique to related antigens. Enzyme-conjugated antibody directed against the major capsid protein (pVIII) or enzyme-conjugated secondary antibody against the primary antibody is used as a detection system in phage ELISA. We suggested expression of the secreted acid phosphatase (SapM) enzyme on M13 pIX minor coat protein directly, and evaluated this hypothesis using In Silico techniques. 3D structure model of the fusion protein (SapM+M13 pIX) was generated and evaluated by related software. MD simulation and TMHMM program results showed a stable fusion protein which is anchored to the inner membrane of E. coli by membrane spanning region suggesting a proper assembling on M13 phage. In theory, SapM enzyme on the phage surface can catalyze the p-nitrophenyl phosphate as substrate and creates yellow color which can be measured at OD=405 nm by microtiter plate reader. We believe that decreasing the antibody layers in phage ELISA will significantly increase the reliability and reproducibility of the test and reduce its time.