Bacterial expression of a subfragment of human factor VIII heavy-chain in Escherichia coli

Human factor VIII (hFVIII) plays a major role in the intrinsic pathway of blood coagulation. The absence or malfunction of FVIII is associated with the X-linked recessive bleeding disorder; known as haemophilia A. A number of these patients who are routinely treated with the hFVIII infusion therapy, develop antibodies that inhibit hFVIII activity. Bacterially produced FVIII-epitopes are capable to neutralize the alloantibodies. The purpose of present study was to express a subfragmment of FVIII heavy chain in E.coli. with this goal, A 1640 bp DNA fragment from heavy-chain of hFVIII was inserted in a T7-based expression vector followed by the transformation of BL21(DE3) strain of E. coli. The use of His6-tagged tail was also considered for purification purpose. Among the total proteins of the isolated clones, a protein was expressed that was detectable with anti-hFVIII-heavy chain monoclonal antibody. Due to low expression level of the A1A2-related protein, its efficient purification was not possible. Further attempts are required for optimization of the expression of sub fragments from hFVIII-heavy chain.

Keywords:

hFVIII , Heavy chain , A1A2 domain , E.coli