pFastBac4x: A Novel Vector for the Simultaneous Expression of Four Proteins-Expression and Purification of a G-Protein Complex as a Case Study.

Baculovirus-mediated expression in insect cells has become one of the most widely used expression systems for the production of eukaryotic proteins. Since the initial discovery that baculoviruses can be employed for the heterologous expression of proteins, the system has evolved and improved in various ways. In this study, we describe the design of a novel vector, termed pFastBac4x, and present a case study involving the co-expression of four proteins in Sf9 insect cells: Gqi alpha, G beta, G gamma, and Ric8A. Protein expression was assessed by Flow Cytometry and Western Blot techniques. The Gqi protein complex was purified via Immobilized Metal Affinity Chromatography, followed by a polishing Size Exclusion Chromatography step. The data demonstrate that the pFastBac4x vector can successfully be used for the co-expression of up to four proteins or protein subunits.