Two Dimensional Structural Analysis and Expression of a New Staphylococcus aureus Adhesin Based Fusion Protein

Document Type: Original Article

Authors

1 Department of Bacteriology and Virology, Faculty of Medicine , Isfahan University of Medical Sciences, Isfahan, Iran

2 Biotechnology Research Centre, Pasteur Institute of Iran, Tehran, Iran

3 Department of Biotechnology, Faculty of pharmacy, Isfahan University of Medical Sciences, Isfahan, Iran

Abstract

Objective(s)
Staphylococcus aureus is a foremost source of numerous nosocomial and community acquired infections. Antibiotic therapy for vancomycin resistant S. aureus (VRSA) can not promise the eradication of infections. Since adhesion is the major route of infections, adhesin based vaccine could suppress S. aureus infections. Fibronectin binding protein A (FnBPA) and clumping factor A (ClfA) are major responsible adhesions involved in S. aureus infections, so they could be candidate vaccine molecules against an extensive range of infections. This project intended to express a new fusion protein construct and analysis of biological activity regarding binding activity.
Materials and Methods
pfnbA- ClfA construct was transformed to Escherichia coli BL21 (DE3). Transformant E. coli were grown in LB broth and induced with IPTG and cellular extracts were separated on SDS–PAGE. RT-PCR was performed to verify expression. Binding activity of fusion protein was studied using human gingival fibroblast (HGF) cell line. D1-D3 protein from unpublished study was used as control.
Results
The expected fusion protein fragment showed by SDS-PAGE. RT-PCR verified the existence of mRNA relating to expressed fusion protein. Binding activity of S. aureus decreased after treatment of HGF cells with fusion protein.
Conclusion
In total,binding activity of fusion protein was approximately two fold lesser than D1-D3 protein. It is supposed that the fusion protein could not be attached to its ligand easily and would be more accessible to antigen presenting cells and consequentlyprotective antibodies will be produced. This project is pending for in vivo infection study in animal model. 

Keywords


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