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Yeast polypeptide fusion surface display levels predict thermal stability and soluble secretion efficiency
E V Shusta1, M C Kieke, E Parke
1Department of Chemical Engineering, University of Illinois, Urbana, IL 61801, USA.
Journal of Molecular Biology
|October 8, 1999
Summary
Yeast display efficiently screens for mutant proteins with improved thermal stability and secretion. This method identifies enhanced protein stability and secretion properties using directed evolution.
Area of Science:
- Biotechnology
- Protein Engineering
- Molecular Biology
Background:
- Yeast cell surface display is a powerful tool for protein engineering.
- Enhancing protein stability and secretion is crucial for therapeutic applications.
Purpose of the Study:
- To investigate yeast cell surface display as a proxy for screening mutant protein stability and secretion.
- To analyze single-chain T cell receptor (scTCR) mutants for improved properties.
Main Methods:
- Engineered single-chain T cell receptor (scTCR) mutants for yeast surface display.
- Cultured Saccharomyces cerevisiae to assess soluble secretion and surface display.
- Analyzed scTCR expression, secretion levels, and thermal stability.
Main Results:
- Wild-type scTCR was not secreted, but all single, double, and triple mutants showed secretion.
- Increased expression and secretion were observed with higher-order mutants.
- Soluble secretion correlated with surface display levels and thermal stability.
- Endoplasmic reticulum quality control influenced display efficiency.
Conclusions:
- Yeast display serves as an effective directed evolution scaffold for identifying mutant proteins with enhanced stability and secretion.
- Mutant scTCRs displayed improved characteristics compared to the wild-type.
- The study highlights the link between intracellular processing and protein display efficiency.