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Updated: Jan 19, 2026
Super-Resolution Imaging of Bacterial Secreted Proteins Using Genetic Code Expansion
Published on: February 10, 2023
Soluble overexpression of a flagellin derivative from Salmonella enterica using synonymous codon substitutions of
Dae-Eun Cheong1, Jihye Lee1, Hye-Ji Choi1
1Department of Biological Sciences and Research Center of Ecomimetics, College of Natural Sciences, Chonnam National University, Yongbong-ro, Buk-gu, Gwangju, 61186, South Korea.
Objective:
To obtain a recombinant flagellin derivative CBLB502, expressed in functionally soluble form, the technology of library construction and screening of synonymous codon variants was employed, and its expression, solubility, and activity were assessed.
Results:
We screened several synonymous codon variants scvCBLB502s with the enhanced solubility from the constructed library, harboring the random substitutions of the first ten amino acid residues of the parental CBLB502 with synonymous codons. Among them, scvCBLB502-5 was purified (> 8.4 mg/l) by single step procedure using an affinity chromatography without any ancillary treatment with protease inhibitor cocktail solution and/or boiling at 90 °C. Subsequent study showed that the recombinant protein scvCBLB502-5 distinctly induced the TLR5 (Toll-Like Receptor 5)-mediated NF-κB activation and also IL-8 production in HEK293-hTLR5 cells.
Conclusion:
Results showed that scvCBLB502-5, engineered through the synonymous codon substitutions, was easily expressed in functionally soluble form and maintained the proper folding to be recognized by TLR5, as an inducer for pathogen-associated molecular pattern (PAMP).
