Efficient Protein Purification Pipeline at SSGCID
Author Information
Author(s): Bryan Cassie M., Bhandari Janhavi, Napuli Alberto J., Leibly David J., Choi Ryan, Kelley Angela, Van Voorhis Wesley C., Edwards Thomas E., Stewart Lance J.
Primary Institution: Seattle Structural Genomics Center for Infectious Disease
Hypothesis
Can a standardized protein purification protocol improve the yield of crystal-quality proteins?
Conclusion
The SSGCID's protein purification protocol significantly increases the success rate of obtaining protein structures.
Supporting Evidence
- The protocol allows for the purification of 400 proteins per year.
- 94% of proteins treated with 3C protease successfully had their tags cleaved.
- The success rate of obtaining structures increased from 13% to 15.9% with the new protocol.
- An average of 53 mg of highly purified protein was obtained per purification.
- 315 purification attempts were made with an overall success rate of 87.6%.
- Using the new method, the projected yield of solved structures increased from 260 to 318.
Takeaway
The researchers created a method to clean proteins that helps scientists see their shapes better, which is important for making new medicines.
Methodology
The study used a semi-automated protein purification pipeline involving immobilized metal-affinity chromatography and size-exclusion chromatography.
Limitations
Some proteins had low solubility and were prone to aggregation during purification.
Statistical Information
P-Value
p<0.05
Statistical Significance
p<0.05
Digital Object Identifier (DOI)
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