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Molecules, Vol. 23, Pages 1491: Expression, Purification, and Characterization of a Novel Hybrid Peptide with Potent Antibacterial Activity

Molecules, Vol. 23, Pages 1491: Expression, Purification, and Characterization of a Novel Hybrid Peptide with Potent Antibacterial Activity

Molecules doi: 10.3390/molecules23061491

Authors: Xubiao Wei Rujuan Wu Lulu Zhang Baseer Ahmad Dayong Si Rijun Zhang

The hybrid peptide cecropin A (1–8)–LL37 (17–30) (C–L), derived from the sequence of cecropin A (C) and LL-37 (L), showed significantly increased antibacterial activity and minimized hemolytic activity than C and L alone. To obtain high-level production of C–L, the deoxyribonucleic acid sequence encoding C–L with preferred codons was cloned into pET-SUMO to construct a fusion expression vector, and overexpressed in Escherichia coli (E. coli) BL21 (DE3). The maximum fusion protein (92% purity) was obtained with the yield of 89.14 mg/L fermentation culture after purification with Ni-NTA Sepharose column. The hybrid C–L was cleaved from the fusion protein by SUMO-protease, and 17.54 mg/L pure active C–L was obtained. Furthermore, the purified C–L showed identical antibacterial and hemolytic activity to synthesized C–L. Stability analysis results exhibited that the activity of C–L changed little below 80 °C for 20 min, but when the temperature exceeded 80 °C, a significant decrease was observed. Varying the pH from 5.0 to 10.0 did not appear to influence the activity of C–L, however, pH below 4.0 decreased the antibacterial activity of C–L rapidly. Under the challenge of several proteases (pepsin, trypsin, and proteinase K), the functional activity of C–L was maintained over 50%. In summary, this study not only supplied an effective approach for high-level production of hybrid peptide C–L, but paved the way for its further exploration in controlling infectious diseases of farm animals or even humans.

Authors:   Wei, Xubiao ; Wu, Rujuan ; Zhang, Lulu ; Ahmad, Baseer ; Si, Dayong ; Zhang, Rijun
Journal:   Molecules
Volume:   23
edition:   6
Year:   2018
Pages:   1491
DOI:   10.3390/molecules23061491
Publication date:   20-Jun-2018
Facts, background information, dossiers
  • production
  • molecules
  • Expression
  • fermentation
  • deoxyribonucleic acid
  • animals
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