High-Level Periplasmic Expression in Escherichia coli Using a Eukaryotic Signal Peptide: Importance of Codon Usage at the 5′ End of the Coding Sequence

• Published in: Protein expression and purification Vol. 20; no. 2; pp. 252 - 264
• Main Authors: Humphreys, David P., Sehdev, Mukesh, Chapman, Andrew P., Ganesh, Ravindra, Smith, Bryan J., King, Lloyd M., Glover, David J., Reeks, Dominic G., Stephens, Paul E.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.11.2000
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Cloning, Molecular; Codon - genetics; codon usage; Databases, Factual; E. coli; Escherichia coli - genetics; Escherichia coli - metabolism; eukaryotic signal peptide; Fab; Gene Expression Regulation, Bacterial; Genes, Bacterial - genetics; Genetic Code; Humans; Immunoglobulin Fab Fragments - genetics; Mice; Molecular Sequence Data; mRNA structure; Mutagenesis - genetics; Nucleic Acid Conformation; Periplasm - metabolism; Plasmids - genetics; Protein Sorting Signals - genetics; Protein Transport; RNA Stability; RNA, Messenger - chemistry; RNA, Messenger - genetics; RNA, Messenger - metabolism; Sequence Analysis, Protein; Yeasts - genetics; eukaryotic signal peptide; Fab; codon usage; E. coli; mRNA structure
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1006/prep.2000.1286

We investigated the ability of signal peptides of eukaryotic origin (human, mouse, and yeast) to efficiently direct model proteins to the Escherichia coli periplasm. These were compared against a well-characterized prokaryotic signal peptide—OmpA. Surprisingly, eukaryotic signal peptides can work very efficiently in E. coli, but require optimization of codon usage by codon-based mutagenesis of the signal peptide coding region. Analysis of the 5′ of periplasmic and cytoplasmic E. coli genes shows some codon usage differences.