A protein overexpression protocol refers to any method to get an organism to make a desired protein in sufficient quantity for further study. Scientists often use bacteria and yeast to make their specific protein of interest, but in theory any organism could work.
In order to study the structure or function of a particular protein, you need to have a significant quantity available for your desired tests. Some proteins naturally occur in large quantities and purify easily from their host organism. Most proteins, however, occur in very small quantities or occur in organisms from which proteins cannot be easily purified. Protein overexpression protocols generate large quantities of desired proteins for further study, allowing scientists to study low-quantity, rare, toxic and even mutated proteins.
Common organisms used in overexpression protocols include bacteria or yeast. Scientists engineer these organisms to carry a gene that codes for a desired protein. They place the gene under specific control so that the organism does not express or make the desired protein until specifically induced to do so. For example, scientists can engineer an organism to carry a gene under direct control of a specific sugar. In the absence of that specific sugar, the organism grows but cannot make the protein. In the presence of the sugar, the organism will make a lot of the protein.
Many different overexpression protocols work, but must be optimized to a specific protein and organism. Optimization usually requires trial and error and often depends upon the structure and function of the protein. Overexpression of some proteins can kill the organism making the protein. In this case, you may need to wait until the population of the organism grows large enough before inducing the production of that protein. Changing organisms or changing types of the same organism may also help.
- Zymo Research: Dual Media Set for Protein Expression
- PNAS: Tuning Escherichia Coli for Membrane Protein Overexpression
- Baker Laboratory: Protocol for the Expression and Purification of Protein
- PubMed.gov: Consequences of Membrane Protein Overexpression in Escherichia coli.
- PubMed.gov: RNAII Transcribed by IPTG-induced T7 RNA Polymerase Is Non-functional as a Replication Primer for ColE1-type Plasmids in Escherichia coli.
About the Author
Robin Wasserman has been writing and prosecuting biochemical patents since 1998. She has served as a biochemical patent agent and a research scientist for a gene-therapy company. Wasserman earned her Doctor of Philosophy in biochemistry and molecular biology, graduating from Harvard University in 1995.
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