ANALYSIS OF A RECOMBINANT DENGUE-2 VIRUS DENGUE-3 VIRUS HYBRID ENVELOPE PROTEIN EXPRESSED IN A SECRETORY BACULOVIRUS SYSTEM

In a step towards a tetravalent dengue virus subunit vaccine which is economical to produce, highly immunogenic and stable, a hybrid dengue virus envelope (E) protein molecule has been constructed, It consists of 36 amino acids from the membrane protein, the N-terminal 288 amino acids of the dengue-2 virus E protein plus amino acids 289-424 of the dengue-3 virus E protein, It has been engineered for secretory express ion by fusion to a mellitin secretory signal sequence and truncation o f the hydrophobic transmembrane segment, Using the baculovirus express ion system and serum-free conditions, more than 95% of recombinant den gue-2 virus-dengue-3 virus hybrid E protein (rD2D3E) was secreted into the cell culture supernatant in a stable form with multiple features indicative of preserved conformation, The hybrid molecule reacted with a panel of dengue virus-and flavivirus-specific MAbs which recognize linear or conformational epitopes on dengue virions, Human dengue viru s-specific antisera also reacted with the protein, The hybrid rD2D3E p rotein was able to inhibit the in vitro binding of dengue-2 and dengue -3 viruses to human myelomonocytic cells, suggesting that the receptor -binding epitope(s) was preserved, Adjuvant-free immunization with the hybrid protein induced an antibody response to both dengue-2 and deng ue-3 virus in outbred mice, comparable in strength to that of individu al rD2E and rD3E proteins. Notably, these antibody responses were prim arily of the IgG2a and IgG2b isotype. A strong dengue virus cross-reac tive T cell response was also induced in the mice, suggesting that den gue virus hybrid E proteins could form the basis of an efficacious mul tivalent dengue virus vaccine.