Using bacteria, especially E. coli, as production hosts is the cheapest and fastest way to produce recombinant proteins. Many proteins however cannot be functionally produced in bacterial cells but require eukaryotic expression hosts. This includes complex mammalian proteins of biomedical interest, such as those with posttranslational modifications, most membrane proteins and large protein complexes. The most effective eukaryotic expression systems are currently:
- the Baculovirus Expression Vector System (BEVS) that uses insect cells as expression hosts
- mammalian cells (mainly CHO and HEK293 cells and variants thereof)
- the yeast Pichia pastoris
- plant cells, especially root and leave cells from A. thaliana and N. tabacum.
Insect cells are particularly successful in production of large proteins with complex folding pathways and protein complexes and can handle many post-translational modifications. Mammalian cells are the natural host of many biomedically relevant proteins and thus able to produce the most native-like proteins, including mammalian types of glycosylation. P. pastoris is especially good to produce eukaryotic membrane proteins in high yields. Plant cells provide a low-cost production platform for active mammalian proteins and complex plant proteins and their assemblies. All eukaryotic systems are endotoxin-free which is important in several downstream applications. The use of eukaryotic cell expression systems often requires a considerable investment in time, expensive media, equipment and expertise and is often non-trivial compared to bacterial expression. Cell-free expression of proteins fulfills an important niche function, for example for producing specifically labeled proteins for NMR investigations.