IDENTIFICATION OF BIOTINYLATED MOLECULES USING A BACULOVIRUS-EXPRESSED LUCIFERASE-STREPTAVIDIN FUSION PROTEIN

A genetic fusion between streptavidin of Streptomyces avidinii and luc iferase of Pyrophorus plagiophthalamus was constructed The fusion prot ein was produced in the Sf9 insect cell line using the baculovirus exp ression vector system (BEVS). Sodium dodecyl sulfate polyacrylamide ge l electrophoresis of the proteins from cells infected with the recombi nant virus, VL1393-LucGR-StreptAv, revealed that the fusion protein mi grated with an apparent molecular weight of 75 kDa. Light emission mea surements showed that the infected cells produced about 255 mg of the chimeric protein per liter of cell culture (127.5 mu g/l x 10(6) cells ). precipitation of the LucGR-StreptAv fusion protein with biotinylate d acrylic beads as well as immunoblot analyses using biotinylated immu noglobulins indicated that both fusion moieties of the chimeric protei n product were functional with respect to their physical and enzymatic activities.