Bioinformatics prediction and experimental validation of VH antibody fragment interacting with factor H binding protein.

OBJECTIVES

We previously conducted an research on the interactions between the ribosome display-selected single chain variable fragment (scFv) and factor H binding protein (fHbp) of . We found that heavy chain variable (VH) fragment of this scFv had considerable affinity to fHbp. These results led us to evaluate the ability of this small antibody fragment in binding and detection of fHbp antigen.

MATERIALS AND METHODS

In this study, at first, the three-dimensional structure of VH fragment was simulated by Kotai Antibody Builder web server. By using ClusPro 2.0 web server, the 3D structure of the soluble form of fHbp (PDB: 2KC0) was docked to the modeled VH fragment to extract the structure of the complex's binding. Molecular dynamics (MD) simulation was carried out using GROMACS 4.5.3 package for 65 ns. Secondly, coding sequence of VH fragment was cloned separately and expressed in . After purification of the VH fragment, its binding activity to fHbp protein was analyzed by enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR) method.

RESULTS

Important amino acids involved in antigen- antibody interaction were identified by analyzing the fHbp-VH complex. The ability of the VH antibody fragment to bind and detect fHbp antigen has been confirmed by the results of analysis, ELISA and SPR methods.

CONCLUSION

These results showed that this small fragment of antibody could be used for designing diagnostic kits.