Published June 18, 2021
| public
Book Section - Chapter
Incorporation of proline analogs into recombinant proteins expressed in Escherichia coli
- Creators
- Breunig, Stephanie L.
- Tirrell, David A.
- Other:
- Petersson, E. James
Abstract
Proline residues are unique in the extent to which they constrain the conformational space available to the protein backbone. Because the conformational preferences of proline cannot be recapitulated by any of the other proteinogenic amino acids, standard mutagenesis approaches that seek to introduce new chemical functionality at proline positions unavoidably perturb backbone flexibility. Here, we detail the incorporation of proline analogs into recombinant proteins in Escherichia coli via a residue-specific mutagenesis strategy. This approach results in global replacement of proline residues with high yields of the recombinant protein of interest, minimal genetic manipulation, and maintenance of backbone conformational constraints.
Additional Information
© 2021 Elsevier Inc. Available online 18 June 2021. This work was supported by grants from the National Institutes of Health (1R01GM134013-01) and the Joseph J. Jacobs Institute for Molecular Engineering for Medicine at the California Institute of Technology. S.L.B. was supported by a National Science Foundation Graduate Research Fellowship under grant number 1745301. We thank our colleagues Alex Chapman, Katharine Fang, and Seth Lieblich for their contributions to our work on proline analogs.Additional details
- Eprint ID
- 110347
- DOI
- 10.1016/bs.mie.2021.05.008
- Resolver ID
- CaltechAUTHORS:20210821-142002730
- 1R01GM134013-01
- NIH
- Joseph J. Jacobs Institute for Molecular Engineering for Medicine
- DGE-1745301
- NSF Graduate Research Fellowship
- Created
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2021-08-21Created from EPrint's datestamp field
- Updated
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2021-08-21Created from EPrint's last_modified field
- Caltech groups
- Jacobs Institute for Molecular Engineering for Medicine
- Series Name
- Methods in Enzymology
- Series Volume or Issue Number
- 656