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Ultra-high expression of a thermally responsive recombinant fusion protein in E. coli.

Publication ,  Journal Article
Chow, DC; Dreher, MR; Trabbic-Carlson, K; Chilkoti, A
Published in: Biotechnology progress
May 2006

Elastin-like polypeptides (ELPs) are recombinant peptide-based biopolymers that contain repetitive sequences enriched in glycine, valine, proline, and alanine. Because of the unusually large fraction of these amino acids in ELPs as compared to other cellular proteins, we hypothesized that intracellular pools of these amino acids can be selectively depleted and limit protein yields during expression. In this study, we examined how culture conditions and individual medium components affect protein yields by monitoring cell growth and protein expression kinetics of E. coli expressing an ELP tagged with a green fluorescent protein (GFP). By determining the underlying principles of superior fusion protein yields generated by the hyperexpression protocol, we further improved protein yields through the addition of glycerol and certain amino acids such as proline and alanine and found that amino acid concentrations and the type of basal medium used strongly influenced this beneficial effect. Surprisingly, amino acids other than those that are abundant in ELPs, for example, asparagine, aspartic acid, glutamine, and glutamic acid, also enhanced protein yields even in a nutrient-rich medium. Compared to commonly used Luria-Bertani medium, the protein yield was improved by 36-fold to the remarkable level of 1.6 g/L in shaker flask cultures with a modified medium and optimized culture conditions, which also led to a 8-fold reduction in the cost of the fusion protein. To our knowledge, this is the highest yield of an ELP-fusion protein purified from E. coli cultured in shaker flasks. This study also suggests a useful strategy to improve the yields of other ELP fusion proteins and repetitive polypeptides.

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Published In

Biotechnology progress

DOI

EISSN

1520-6033

ISSN

8756-7938

Publication Date

May 2006

Volume

22

Issue

3

Start / End Page

638 / 646

Related Subject Headings

  • Time Factors
  • Temperature
  • Recombinant Fusion Proteins
  • Oligopeptides
  • Nitrogen
  • Kinetics
  • Green Fluorescent Proteins
  • Escherichia coli
  • Elastin
  • Culture Media, Conditioned
 

Citation

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Chow, D. C., Dreher, M. R., Trabbic-Carlson, K., & Chilkoti, A. (2006). Ultra-high expression of a thermally responsive recombinant fusion protein in E. coli. Biotechnology Progress, 22(3), 638–646. https://doi.org/10.1021/bp0503742
Chow, Dominic C., Matthew R. Dreher, Kimberly Trabbic-Carlson, and Ashutosh Chilkoti. “Ultra-high expression of a thermally responsive recombinant fusion protein in E. coli.Biotechnology Progress 22, no. 3 (May 2006): 638–46. https://doi.org/10.1021/bp0503742.
Chow DC, Dreher MR, Trabbic-Carlson K, Chilkoti A. Ultra-high expression of a thermally responsive recombinant fusion protein in E. coli. Biotechnology progress. 2006 May;22(3):638–46.
Chow, Dominic C., et al. “Ultra-high expression of a thermally responsive recombinant fusion protein in E. coli.Biotechnology Progress, vol. 22, no. 3, May 2006, pp. 638–46. Epmc, doi:10.1021/bp0503742.
Chow DC, Dreher MR, Trabbic-Carlson K, Chilkoti A. Ultra-high expression of a thermally responsive recombinant fusion protein in E. coli. Biotechnology progress. 2006 May;22(3):638–646.
Journal cover image

Published In

Biotechnology progress

DOI

EISSN

1520-6033

ISSN

8756-7938

Publication Date

May 2006

Volume

22

Issue

3

Start / End Page

638 / 646

Related Subject Headings

  • Time Factors
  • Temperature
  • Recombinant Fusion Proteins
  • Oligopeptides
  • Nitrogen
  • Kinetics
  • Green Fluorescent Proteins
  • Escherichia coli
  • Elastin
  • Culture Media, Conditioned