1. "Top-down" overexpression optimization of butelase-1 in Escherichia coli and its application in anti-tumor peptides.
- Author
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Zhao J, Song W, Huang Z, Yuan X, Huang Y, Hou Y, Liu K, Jin P, and Hu SQ
- Subjects
- Humans, Gene Expression, Peptides chemistry, Peptides metabolism, Peptides genetics, Recombinant Proteins genetics, Recombinant Proteins metabolism, Recombinant Proteins chemistry, Antineoplastic Agents pharmacology, Antineoplastic Agents chemistry, Escherichia coli genetics, Escherichia coli metabolism
- Abstract
Butelase-1, the fastest known Asn/Asp-specific peptide ligase capable of catalyzing peptide ligation and cyclization, holds promising application prospects in the fields of food and biology. However, limited research exists on its recombinant expression and potential applications in peptide drugs. In this study, the activity of recombinantly-produced butelase-1 was enhanced by co-expressing it with a molecular chaperone in the SHuffle T7 strain. By introducing single or multiple synonymous rare codons at the beginning of the coding regions of beta-strand or alpha-helix, in combination with ribosomal binding site engineering, the activity of butelase-1 could be further improved. Consequently, the butelase-1 with a specific activity of 386.93 U/mg and a catalytic efficiency of 11,048 M
-1 s-1 was successfully prepared in E. coli, resulting in a total activity of 8183.54 U/L and a yield of about 100 mg/L. This optimized butelase-1 was then used to efficiently cyclize the redesigned anti-cancer peptide lunasin, leading to enhanced bioavailability and anti-cancer effects. Overall, this study not only provided valuable biotechnology strategies for improving the recombinant expression of butelase-1 but also demonstrated a successful application for enhancing the biological efficacy of anti-cancer peptides., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)- Published
- 2024
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