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Project Description

Content

This year we set out to characterize and submit a part capable of secretion based single-chain variable fragment antibody protein purification. Utilizing osmotically inducible protein Y (OsmY)in the RFC25 protein fusion standard, we have created a construct capable of secreting a protein of interest from E. coli. Once secreted, our construct allows for simple cleavage and purification using the His-tag and TEV protease sites found between OsmY and the gene of interest. To characterize the construct, we plan to use a number of different proteins. This will not only begin to assess what classes of proteins are capable of being purified and secreted by our construct, but will also allow us to assess the practicality of our construct in industry-level production of proteins. We plan to purify antibodies against Salmonella protein Dihydroxyacid Dehydratase with our construct and ultimately compare the efficacy of our purified proteins to those of more traditional purification methods.

We believe that our protein purification construct will be remarkably useful for purifying proteins and could significantly decrease the cost of antibody production, as proteins of interest are secreted from the cell, decreasing the number of steps required to purify the protein significantly. Future teams will be able to insert any protein of interest into the construct and secrete the protein into the cytoplasm of the cell, simplifying the purification process. OsmY is an exceptionally useful protein for secretion based purification and our construct allows the iGEM community a modular vector for expression and purification of proteins.

References

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1. Overall project summary
2. Project Details
3. Materials and Methods
4. The Experiments
5. Results
6. Data analysis
7. Conclusions

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