High-yield production of a chimeric glycoprotein based on permuted E1 and E2 HCV envelope ectodomains

Abstract In this report it is described for the first time the expression and purification of large quantities of a soluble and correctly folded chimeric recombinant protein, E2 661 E1 340 , containing the permuted Hepatitis C virus (HCV) glycoprotein ectodomains E1 (amino acids 192–340) and E2 (amino acids 384–661). Using the baculovirus/insect cell expression system, 8 mg of secreted protein were purified from 1 L of culture media, a yield 4 times higher than the described for its counterpart E1 341 E2 661 . This permuted chimeric protein is glycosylated and possesses a high tendency to self-associate. The fluorescence emission spectrum indicates that Trp residues occupy a relatively low hydrophobic environment. The secondary structure was determined by deconvolution of the far-UV circular dichroism spectrum yielding 13% α-helix structure, 49% extended structure and 38% non-ordered structure. E2 661 E1 340 binds to antibodies present in human sera from HCV-positive patients, a binding that is blocked at different levels by a rabbit anti-E2 661 antibody. All these structural and antigenic features of E2 661 E1 340 are very similar to those described for E1 340 E2 661 , Thus, this high-yield isolated chimeric protein may be a valuable tool to study the first steps of the HCV infection.