Engineering an improved bioconjugation enzyme : a case study in yeast surface display
Abstract/Contents
- Abstract
- Enzyme engineering is a valuable tool for generating powerful and robust biocatalysts for a broad range of applications. Directed evolution using cell surface display has been previously applied for enzyme engineering, and involves iterative optimization of a protein sequence to yield a desired phenotype. However, previous applications have been carefully tailored for specific enzymes, and this process of developing a good screen is a significant bottleneck in engineering. Co-display of an enzyme and its substrate on the yeast surface is a promising general strategy for evolving bond-forming enzymes, but has only been used to engineer one enzyme. We extended yeast surface co-display to engineer Sfp, a bioconjugation enzyme, for improved activity with a non-native substrate. Sfp library screening identified mutational hotspots in the enzyme and led to the design of Sfp variants with orders of magnitude enhancements in activity. However, these activity improvements were only apparent in contexts similar to yeast surface co-display, reflecting artifacts in the use of this approach for enzyme engineering. Based on our results, we offer suggestions for improving the success of engineering strategies based on yeast surface display.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2019; ©2019 |
| Publication date | 2019; 2019 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Ye, Anne Zijing |
|---|---|
| Degree supervisor | Cochran, Jennifer R |
| Thesis advisor | Cochran, Jennifer R |
| Thesis advisor | Bertozzi, Carolyn R, 1966- |
| Thesis advisor | Fordyce, Polly |
| Degree committee member | Bertozzi, Carolyn R, 1966- |
| Degree committee member | Fordyce, Polly |
| Associated with | Stanford University, Department of Bioengineering. |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Anne Zijing Ye. |
|---|---|
| Note | Submitted to the Department of Bioengineering. |
| Thesis | Thesis Ph.D. Stanford University 2019. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2019 by Anne Zijing Ye
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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