Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Engineering HlyA hypersecretion in Escherichia coli based on proteomic and microarray analyses.

Pat S Lee1, Kelvin H Lee

  • 1School of Chemical and Biomolecular Engineering, Cornell University, 120 Olin Hall, Ithaca, New York 14853-5201, USA.

Biotechnology and Bioengineering
|December 8, 2004
PubMed
Summary

Engineered Escherichia coli shows enhanced protein secretion by optimizing translation speed. This metabolic engineering approach improves recombinant protein production for biotechnology applications.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Harvest Process and Affinity Resin Selection Impacts on Adeno-Associated Virus Residual Host Cell Protein Retention.

Human gene therapy·2026
Same author

Characterization of difficult-to-remove host cell proteins in adeno-associated virus downstream processing.

Molecular therapy. Methods & clinical development·2025
Same author

NIIMBL-facilitated active listening meeting between industry and FDA identifies common challenges for adoption of alternative and rapid microbiological methods.

PDA journal of pharmaceutical science and technology·2025
Same author

Evaluation of "Difficult-to-Express" Monoclonal Antibodies in a CHO-Based Hybrid Site-Specific Integration System Under Industrially Relevant Conditions.

Biotechnology journal·2025
Same author

Defining a Generic Technology Transfer Process for use Across the Biopharmaceutical Manufacturing Industry.

PDA journal of pharmaceutical science and technology·2025
Same author

A comparison of SWATH-MS methods for measurement of residual host cell proteins in adeno-associated virus preparations.

Frontiers in bioengineering and biotechnology·2025

Area of Science:

  • Microbiology
  • Biotechnology
  • Metabolic Engineering

Background:

  • Escherichia coli is a key host for recombinant protein production.
  • Extracellular secretion simplifies purification and reduces protein aggregation.
  • Engineering enhanced secretion in E. coli has been challenging due to complex mechanisms.

Purpose of the Study:

  • To engineer a hypersecreting Escherichia coli strain for enhanced HlyA (alpha-hemolysin) secretion.
  • To utilize transcriptomic and proteomic data for metabolic engineering insights.
  • To develop a novel strategy for improving recombinant protein secretion.

Main Methods:

  • Random mutagenesis of E. coli W3110 cells secreting HlyA.
  • Activity assays to identify hypersecreting mutants.

Related Experiment Videos

  • High-density microarrays for mRNA profiling and proteomics for protein expression analysis.
  • Mathematical modeling of prokaryotic translation to design a modified hlyA gene.

    • Main Results:

    • A mutant strain secreting fourfold more active HlyA was identified.
    • Decreased expression of tRNA-synthetases observed in the mutant.
    • A variant hlyA gene using rare codons to slow translation enhanced HlyA secretion.
    • The engineered strain demonstrated improved hypersecretion phenotype.

    Conclusions:

    • Successful metabolic engineering of E. coli for enhanced protein secretion using transcriptomic and proteomic data.
    • Slowing translation via rare codon usage can improve protein secretion.
    • The developed strategy offers a valuable tool for the biotechnology community.