Journal
PROCESS BIOCHEMISTRY
Volume 50, Issue 11, Pages 1767-1773Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.procbio.2015.08.005
Keywords
Maltose; Inducible vector; Antimicrobial activity; Bacillus subtilis; PR39
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Funding
- National Science and Technology Support Program [2013BAD10B03]
- National Natural Science Foundation of China [31472104]
- National Basic Research Program [2012CB124703]
- China Agriculture Research System [CARS-36]
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As a promising antibiotic candidate, it is necessary to find a safe method to produce the porcine cathelicidin antimicrobial peptide PR39. In this study, the antimicrobial peptide PR39 was successfully expressed with the maltose-inducible vector pGJ148 in the Bacillus subtilis WB800N. After induction with 3% maltose concentration, the fusion protein was successfully secreted onto the culture medium directly. After purification a Ni-NTA resin column, the fusion protein with a concentration of 17 mg/l was obtained from fermentation culture. Then, the purified fusion protein was cleaved using enterokinase, and tag-free PR39 was released. Approximately 2 mg of the recombinant PR39 with a purity of approximately 92.0% was obtained from a 1-1 culture medium after purification with cation exchange column. The evaluation of antimicrobial and haemolytic activity demonstrated that the recombinant PR39 exhibited a high antimicrobial activity against several Gram-negative strains and a low haemolytic activity (256 mu M) against human red blood cells. The synthetic PR39 produced similar results. This work expanded and provided a safe protocol for the large-scale production of antimicrobial peptides in B. subtilis. (C) 2015 Elsevier Ltd. All rights reserved.
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