Protein Expression Systems

Protein Production Summary - Protein Expression Systems

To produce recombinant proteins, the gene is isolated and cloned into an expression vector. Most recombinant proteins are from humans but are expressed in other organisms such as bacteria, yeast, or animal cells in culture.

Many issues affect the expression of recombinant proteins because of organisms difference. The gene for a recombinant protein can also be altered to make the protein more stable. Usage of regulated expression vectors can control the rate at which recombinant protein is produced. Another factor is protein export. It is easier to isolate and purify recombinant proteins if they are exported outside the cell.

Many recombinant proteins require protein modifications, such as glycosylation, that are available only in eukaryotic cells; this sometimes leads to the use of yeast, insect cells, and mammalian cell culture systems.

A major consideration in the production of recombinant proteins is the choice of expression system. The three major main systems for the expression of recombinant proteins are bacterial expression systems (e.g. Lactococcus lactis and Escherichia coli), yeast and baculovirus and mammalian cell mediated protein expression. 

Bacterial Expression Systems

What is Bacterial Protein Expression System

The first choice of an expression system for the production of recombinant proteins for many investigators is E. coli expression system. In other words, E. coli protein expression system is now the most wide-used and economical expression system. Many proteins require special modifications that are available only in eukaryotic cells. Disulfide bonds are not usually formed inside bacterial cells and are rarely formed properly even when bacterial hosts secrete eukaryotic proteins.

Advantages of Bacterial Expression System

The E. coli expression system has many advantages compared with other expression systems such as easy growth conditions and rapid biomass accumulation. Expressing eukaryotic proteins in eukaryotic cells that contamination with bacterial components can be avoided. Despite purification, bacterial components that are toxic or promote immune reactions may be present in traces if bacteria are used for production. Furthermore, some eukaryotic proteins are unstable or inactive after being made by bacterial cells.

Protein Modifications in Bacterial Protein Expression

1. Chemical modifications that form novel amino acids in the polypeptide chain.
2. Formation of disulfide bonds between correct cysteine partners.
3. Glycosylation.
4. Addition of a variety of extra groups, such as fatty acid chains or acetyl groups.
5. Cleavage of precursor proteins may be needed for secretion, correct folding, or activation of proteins.

Sino Biological can provide you high-quality bacterial protein expression service. 

Protein Expression in Mammalian Cells

What is Mammalian Cells Expression System

Some cloned animal genes may need to be expressed in cultured mammalian cells. Mammalian shuttle vectors contain an origin of replication and an antibiotic resistance gene for growth in bacteria. In other words, mammalian shuttle vectors have features for producing recombinant proteins in mammalian cell cultures. Because most antibiotics do not harm mammalian cells, a different method of selecting the mammalian shuttle vector must be used.

Mammalian Cell Lines Yield High Levels of Recombinant Protein

A major issue with protein expression in mammalian cells is finding a cell with high levels of the recombinant protein. This single chosen cell is then used to generate large cultures. This ensures that the recombinant protein is identical throughout the culture. Many techniques are available to isolate single cells with high levels of recombinant protein such as limiting dilution cloning, cell sorting, gel microdrop technology and automated cell selection.

Mammalian transient expression systems enable flexible and rapid production of proteins. They are ideal for generating large amounts of protein within 1 to 2 weeks. Sino Biological offers mammalian transient expression recombinant protein services in CHO and HEK 293 cells. Click the button to get your mammalian expression recombinant proteins below. 

Protein Expression in Baculovirus Expression System

What is Baculovirus Expression System

Mammalian cells are relatively delicate and have complex nutritional requirements. In contrast, cultured insect cells are relatively robust and can be grown in simpler media than mammalian cells. Insect-based expression systems have the additional advantage of providing post-translational modifications that are similar to mammalian cells.

The baculovirus-mediated insect protein expression system is based on the ability of recombinant baculovirus to infect insect cells, and the foreign protein that is encoded by the baculovirus is expressed by the cellular machinery. Insect cells are easy to grow for making recombinant protein. Insect cells are infected with a virus genome that has the recombinant protein gene. Rather than making viral particles, the infected insect cells make recombinant protein.

Insect Cell Expression Advantages

Baculovirus-meditated expression systems possess a number of advantages:
1. High expression levels driven by the strong polyhedrin promoter.
2. Ability of the insect cells to perform a variety of post-translational modifications.
3. High capacity for multiple genes or large inserts.
4. Bio-safe and easy to handle.

Click the button below to inquire Sino Biological's high-efficiency expression of intracellular and extracellular proteins by baculovirus expression system.

Advantages and Disadvantages of Different Expression Systems

Differences of Recombinant Protein Expression Systems

In conclusion, which expression system is used is usually dictated by two things: the ultimate use of the product, and the nature of the individual protein to be expressed. Each expression system has its own unique strengths and weaknesses when it comes to
characteristics such as cost, speed, and ability to glycosylate or fold proteins.

How to Select a Proper Expression System

No expression system is perfect, and each recombinant protein must be evaluated for which system will work the best. Each protein expression system falls on a continuum of worst to best for characteristics such as speed, cost, glycosylation, folding, and government regulations.

  WORST     BEST
SPEED
Transgenics

Plants

Mammalian

Insect

Yeast

Bacteria
COST
Transgenics

Mammalian

Plants

Insect

Yeast

Bacteria
GLYCO-SYLATION
Bacteria

Yeast

Plants

Insect
 
Transgenics

Mammalian
FOLDING
Bacteria

Yeast
 
 
Plants

Insect

Transgenics

Mammalian
GOVERNMENT REGULATION
Transgenics

Plants

Insect
 
Yeast

Mammalian

References

Zhu J (2012) Mammalian cell protein expression for biopharmaceutical production. Biotechnol Adv 30(5): 1158-1170.
Yilmaz H, et al. (2019) Production of recombinant n protein of infectious bronchitis virus using the baculovirus expression system and its assessment as a diagnostic antigen. Appl Biochem Biotechnol 187(2): 506-517.
Murphy CI, et al. (2018) Expression and purification of recombinant proteins using the baculovirus system. Current protocols in molecular biology 123(1): e61.
Rosano GL, et al. (2014) Recombinant protein expression in escherichia coli: Advances and challenges. Front Microbiol 5: 172.
Chiou HC, et al. (2014) Scalable transient protein expression. Methods in molecular biology (Clifton, N.J.) 1104: 35-55.

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