Skip to Main content Skip to Navigation
Journal articles

Engineering an efficient secretion of leech carboxypeptidase inhibitor in Escherichia coli.

Abstract : BACKGROUND: Despite advances in expression technologies, the efficient production of heterologous secreted proteins in Escherichia coli remains a challenge. One frequent limitation relies on their inability to be exported to the E. coli periplasm. However, recent studies have suggested that translational kinetics and signal sequences act in concert to modulate the export process. RESULTS: In order to produce leech carboxypeptidase inhibitor (LCI) in the bacterial periplasm, we compared expression of the natural and optimized gene sequences, and evaluated export efficiency of LCI fused to different signal sequences. The best combination of these factors acting on translation and export was obtained when the signal sequence of DsbA was fused to an E. coli codon-optimized mature LCI sequence. When tested in high cell density cultures, the protein was primarily found in the growth medium. Under these conditions, the engineered expression system yields over 470 mg.l-1 of purified active LCI. CONCLUSION: These results support the hypothesis that heterologous secreted proteins require proper coupling between translation and translocation for optimal high-level production in E. coli.
Document type :
Journal articles
Complete list of metadatas

Cited literature [27 references]  Display  Hide  Download

https://hal-pasteur.archives-ouvertes.fr/pasteur-00670636
Contributor : Ed. Bmc <>
Submitted on : Wednesday, February 15, 2012 - 5:17:09 PM
Last modification on : Monday, January 13, 2020 - 5:08:06 PM
Long-term archiving on: : Thursday, November 22, 2012 - 12:51:03 PM

Files

1475-2859-8-57.pdf
Publisher files allowed on an open archive

Identifiers

Collections

Citation

Juan-Miguel Puertas, Jean-Michel Betton. Engineering an efficient secretion of leech carboxypeptidase inhibitor in Escherichia coli.. Microbial Cell Factories, BioMed Central, 2009, 8 (1), pp.57. ⟨10.1186/1475-2859-8-57⟩. ⟨pasteur-00670636⟩

Share

Metrics

Record views

391

Files downloads

674