ABSTRACT
The baculovirus-insect cell system has become a widely used tool for protein expression. Because of its ease to use in combination with the processing potential of eukaryotic cells (Table 9.1 and Table 9.2), this system is especially useful for expression of integral membrane proteins in a structurally relevant, functional conformation. In addition, the baculovirus genome tolerates large insertions (30 kb) enabling coexpression of multiple domains or constituents of multimeric complexes. Finally, baculovirus allows selection of medium to strong promoters that are active during various periods after infection, can be obtained in high titers, and — last but not least — has a very low pathogenicity. Nevertheless, a PubMed search indicates that after a strong increase between 1987 and 1995, the number of publications in which the baculovirus-insect cell system was used for expression of membrane
proteins has remained constant or may have started to decline since 2001 (Figure 9.1). We interpret this trend as correlated with the difficulties to obtain high-resolution data on the structure of membrane proteins. The availability of functional membrane proteins is the bottleneck for obtaining two-dimensional (2D) and threedimensional (3D) crystals. The increase in knowledge on the processes that influence folding and trafficking of membrane proteins in the baculovirus system has lagged far behind the development of molecular biology methodology. Also, standardized methods for solubilization, purification, and reconstitution of membrane proteins with preservation of function are only now beginning to emerge. Against this background, we present here our view of the state of the baculovirus-insect cell expression system, with emphasis on the new developments required to meet the need for detailed knowledge of structure and function of membrane proteins in the current genomic and proteomic landslide.
