REQUIREMENTS FOR OPTIMAL EXPRESSION OF SECRETED AND NONSECRETED RECOMBINANT PROTEINS IN VACCINIA VIRUS SYSTEMS

Selection of an optimal promoter is necessary for efficient expression of foreign genes with vaccinia virus. Since a variety of powerful (ho mologous) vaccinia virus promoters and foreign (heterologous) promoter systems have been described for use in vaccinia, we have addressed th e question of whether a general rule exists that allows the prediction of the optimal promoter/gene combination. We have compared the expres sion properties of four secreted proteins, the human blood clotting fa ctor IX (FM), the human blood glycoprotein Protein S (ProtS), the huma n von Willebrand factor (vWF), and the Hepatitis B virus (HBV) middle surface glycoprotein preS2, with proteins that were reported not to be secreted, the HBV large surface glycoprotein preS1 and the murine leu kemia virus (MuLV) BM-5 Eco gag protein. In addition, we have included in our study an internal control protein, the vaccinia virus pll prot ein, to monitor possible side effects of the promoter system used. Gen es encoding the foreign proteins were placed either under control of a synthetic vaccinia virus early/late promoter (selP) or under control of the bacteriophage T7 promoter (T7/emc system). The secreted protein s were more efficiently expressed when fused to the homologous promote r. Direct comparison of the two promoters indicated that the expressio n level ranged between 1.4 (ProtS) and 3.9 (FM)-fold higher with the s elP than with the T7 promoter. In contrast, the cell-associated HBV pr eS1 was more efficiently expressed under the T7 promoter and the MuLV BM-5 Eco gag polypeptide was expressed equally well from both promoter s. These data indicate that a careful prediction of optimal promoter/f oreign gene combinations for the vaccinia virus expression system is p ossible. The choice of the optimal promoter/expression system is based on a simple classification scheme, discriminating secreted and nonsec reted proteins. (C) 1995 Academic Press, Inc.