Protein Expression system

Proteins are synthesized and regulated depending upon the functional need in the cell. The blueprints for proteins are stored in DNA and decoded by highly regulated transcriptional processes to produce messenger RNA (mRNA). The message coded by an mRNA is then translated into a protein. Transcription is the transfer of information from DNA to mRNA, and translation is the synthesis of protein based on a sequence specified by mRNA.

The selection of the system depends on the type of protein, the requirements for functional activity and the desired yield. These expression systems include mammalian, bacterial, yeast and insect. Each system has advantages and challenges, and choosing the right system is important for successful recombinant protein expression.

Mammalian cell system

Mammalian expression systems can be used to produce mammalian proteins that have the most native structure and activity due to its physiologically relevant environment. This results in high levels of post-translational processing and functional activity.

Mammalian expression systems can be used to produce proteins transiently or through stable cell lines, transient production can generate large amounts of protein in one to two weeks. These transient, high-yield mammalian cell expression systems utilize suspension cultures and can produce gram-per-liter yields. Furthermore, these proteins have more native folding and post-translational modifications, such as glycosylation, as compared to other expression systems.

Bacterial system

The genetics and biochemistry of Escherichia coli are probably the best understood of any known organism. The knowledge gained in the study of E. coli biology has been applied to the development of many of today¡¯s molecular cloning techniques. Most cloning vectors and methods utilize E. coli or its phages as a preferred host, primarily because of the ease with which the bacterium can be grown and genetically manipulated. These same characteristics made E. coli an attractive early choice as a host for the production of large quantities of protein encoded by cloned genes. Aside from its well-studied biology, E. coli is suitable as the basis of an expression system because of its rapid doubling time and its ability to grow in inexpensive media. Years of study devoted to gene expression in E. coli have provided numerous choices for transcriptional and translational control elements that can be applied to the expression of foreign genes.

Insect system

Insect cells can be used for high level protein expression with modifications similar to mammalian systems. There are several systems that can be used to produce recombinant baculovirus, which can then be utilized to express the protein of interest in insect cells. These systems can be easily scaled up and adapted to high-density suspension culture for large-scale expression of protein that is more functionally similar to native mammalian protein. Recombinant baculovirus production can be time consuming and culture conditions more challenging than prokaryotic systems.

Yeast system

The highly developed genetic system, ease of use, reduced time input and costs have made S. cerevisiae an attractive organism for the expression and production of recombinant proteins. Yeasts are able to carry specifically designed plasmids and this ability is valuable in a recombinant protein expression system. The plasmid used consists of restriction sites that can be used to insert the gene sequence of interest. Transformation of yeasts with the plasmid produces the desired protein and can be appropriately scaled up