Expression of plectasin in Pichia pastoris and its characterization as a new antimicrobial peptide against Staphyloccocus and Streptococcus.

Abstract

Recombinant plectasin, the first fungus defensin, was expressed in Pichia pastoris and purified, and its physical, chemical and antimicrobial characteristics were studied. Following a 120 h induction of recombinant yeast, the amount of total secreted protein reached 748.63 μg/ml. The percentage of recombinant plectasin was estimated to be 71.79% of the total protein. After purification with a Sephadex G-25 column and RP-HPLC, the identity of plectasin was verified by MALDI-TOF MS. Plectasin exhibited strong antimicrobial activity against the Gram-positive bacteria Staphyloccocusaureus, Staphylococcus epidermidis, Streptococcus pneumoniae, and Streptococcus suis. At a concentration of 2560 μg/ml, this peptide showed approximately equal activity against S. aureus, S. epidermidis, S. suis, and S. pneumoniae, when compared to 320 μg/ml vancomycin, 640 μg/ml penicillin, 320 μg/ml vancomycin and 160 μg/ml vancomycin, respectively. In addition, plectasin showed anti-S. aureus activity over a wide pH range of 2.0 and 10.0, a high thermal stability at 100 °C for 1h and remarkable resistance to papain and pepsin. The expression and characterization of recombinant plectasin in P. pastoris has potential to treat Streptococcus and Staphyloccocus infections when most traditional antibiotics show no effect on them. Our results indicate that plectasin can be produced in large quantities, and that it has pharmaceutical importance for the prevention and clinical treatment of Staphyloccocus and Streptococcus infections.

• Authors: Jun Zhang, Yalin Yang, Da Teng, Zigang Tian, Shaoran Wang, Jianhua Wang
• Journal: Protein expression and purification (Protein Expr Purif) Vol. 78 Issue 2 Pg. 189-96 (Aug 2011) ISSN: 1096-0279 [Electronic] United States
• PMID: 21558006 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2011 Elsevier Inc. All rights reserved.

Chemical References

• Anti-Bacterial Agents
• Antimicrobial Cationic Peptides
• Peptides
• Recombinant Proteins
• plectasin

Topics

• Amino Acid Sequence
• Animals
• Anti-Bacterial Agents (chemistry, isolation & purification, metabolism)
• Antimicrobial Cationic Peptides (biosynthesis, chemistry, genetics, isolation & purification)
• Ascomycota (genetics)
• Base Sequence
• Chromatography, Gel
• Cloning, Molecular
• Electrophoresis, Polyacrylamide Gel
• Hemolysis
• Hydrogen-Ion Concentration
• Industrial Microbiology
• Microbial Sensitivity Tests
• Molecular Sequence Data
• Peptides (chemistry, genetics, isolation & purification)
• Pichia (chemistry, genetics, metabolism)
• Rabbits
• Recombinant Proteins (biosynthesis, chemistry, genetics, isolation & purification)
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
• Staphylococcus (drug effects)
• Streptococcus (drug effects)
• Temperature