In-cell production of a genetically-encoded library based on the θ-defensin RTD-1 using a bacterial expression system
| Autor: | Alexander Shekhtman, Tao Bi, Yilong Li, Julio A. Camarero |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
medicine.medical_treatment Bacterial Toxins Clinical Biochemistry Pharmaceutical Science Peptide medicine.disease_cause Peptides Cyclic Biochemistry Article Defensins 03 medical and health sciences Peptide Library Drug Discovery Escherichia coli medicine Protein Splicing Amino Acid Sequence Peptide library Molecular Biology Peptide sequence Defensin chemistry.chemical_classification Antigens Bacterial Protease 030102 biochemistry & molecular biology biology Organic Chemistry biology.organism_classification Molecular biology Bacillus anthracis 030104 developmental biology chemistry Molecular Medicine Heterologous expression |
| Zdroj: | Bioorganic & Medicinal Chemistry. 26:1212-1219 |
| ISSN: | 0968-0896 |
| DOI: | 10.1016/j.bmc.2017.09.002 |
| Popis: | We report the high-yield heterologous expression of bioactive θ-defensin RTD-1 inside Escherichia coli cells by making use of intracellular protein trans-splicing in combination with a high efficient split-intein. RTD-1 is a small backbone-cyclized polypeptide with three disulfide bridges and a natural inhibitor of anthrax lethal factor protease. Recombinant RTD-1 was natively folded and able to inhibit anthrax lethal factor protease. In-cell expression of RTD-1 was very efficient and yielded ≈ 0.7 mg of folded RTD-1 per gram of wet E. coli cells. This approach was used to generate of a genetically-encoded RTD-1-based peptide library in live E. coli cells. These results clearly demonstrate the possibility of using genetically-encoded RTD-1-based peptide libraries in live E. coli cells, which is a critical first step for developing in-cell screening and directed evolution technologies using the cyclic peptide RTD-1 as a molecular scaffold. |
| Databáze: | OpenAIRE |
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