Reconstitution of Recombinant Viral Envelope Proteins

Abstract

Liposomes comprising lipid bilayer membranes enclosing aqueous internal compartments serve as attractive models for reconstitution of a variety of membrane proteins such as viral envelope proteins. The lipid membranes consist of amphipathic lipids that are similar to those found in authentic cell membranes. The preparation of liposomes is a routine procedure and the possibility of engineering liposomes of varying size, as well as incorporating different membrane proteins on their surface, has made them useful for many biotechnological applications including vaccine development, drug delivery, and gene delivery. This chapter describes production, purification, and reconstitution of the envelope glycoproteins E1 and E2 of rubella virus (RV). The corresponding viral envelope proteins are engineered to display a FLAG epitope, which is a polypeptide protein tag, and to also display a polyhistidine tag at their N and C termini for specific and sensitive immunological identification, as well as for gentle and easy purification. The genes encoding the modified viral proteins are transferred into the baculovirus genome under the transcriptional regulation of the polyhedrin gene promoter and produced in infected S f 9 insect cell suspension cultures, using the resultant baculovirus expression vectors. The recombinant proteins are purified by immobilized metal ion affinity chromatography and reconstituted into liposomes via their hydrophobic transmembrane anchors.