Expression, purification and characterization of a recombinant antimicrobial peptide Hispidalin in Pichia pastoris
| Autor: | Demei Meng, Xue‐Qing Sun, Jin-Cheng Hu, Lin-Yue Shi, Zhen-Chuan Fan, Yu-Jie Lv, Li Wenjuan |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
0106 biological sciences
Gene Expression Microbial Sensitivity Tests medicine.disease_cause 01 natural sciences Pichia Microbiology law.invention Pichia pastoris 03 medical and health sciences chemistry.chemical_compound Drug Stability law 010608 biotechnology medicine Escherichia coli 030304 developmental biology Antibacterial agent 0303 health sciences biology Bacteria Chemistry Hydrogen-Ion Concentration Antimicrobial biology.organism_classification Proteinase K Recombinant Proteins Anti-Bacterial Agents Papain Cucurbitaceae Recombinant DNA biology.protein Antibacterial activity Biotechnology Antimicrobial Cationic Peptides |
| Zdroj: | Protein expression and purification. 160 |
| ISSN: | 1096-0279 |
| Popis: | Hispidalin is a novel antimicrobial peptide isolated from the seeds of Benincasa hispida and is reported to have broad antimicrobial activity against various bacterial and fungal pathogens. To produce significant amounts of Hispidalin, a recombinant Hispidalin with an N-terminal 6 × His tag and an enterokinase sequence, for the first time, was successfully expressed in Escherichia coli or Pichia pastoris cell factory. Results showed that the E. coli-derived recombinant Hispidalin did not show any antimicrobial activity against all the tested strains, whereas the P. pastoris-derived recombinant Hispidalin (rHispidalin) showed a broad antibacterial spectrum against five pathogenic bacteria of both Gram-negative and Gram-positive. rHispidalin also has bactericidal activity and completely killed all of the Staphylococcus aureus within 40 min. Additionally, rHispidalin showed a broad range of thermostability and pH stability, and a hemolytic activity of less than 2% even at a concentration of 300 μg/ml; it was resistant to trypsin and proteinase K, but was moderately sensitive to pepsin and papain. Moreover, rHispidalin effectively permeabilized the cytoplasmic membrane and disrupted the morphology of targeted bacterial cells. After an initial optimization was performed, the amount of rHispidalin accumulation could reach as high as 98.6 μg/ml. These results indicate that Hispidalin could be produced on a large scale by P. pastoris and has a great potential to be utilized as a new antibacterial agent for further development. |
| Databáze: | OpenAIRE |
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