Journal
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
Volume 97, Issue 8, Pages 3535-3543Publisher
SPRINGER
DOI: 10.1007/s00253-012-4441-1
Keywords
Ranalexin; Fusion protein; Antimicrobial peptide; Escherichia coli; Recombinant protein
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Funding
- Syrian Ministry of Higher Education and Tishreen University, Syria
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The coding sequence, which corresponds to the mature antimicrobial peptide ranalexin from the frog Rana catesbeiana, was chemically synthesized with preferred codons for expression in Escherichia coli. It was cloned into the vector pET32c (+) to express a thioredoxin-ranalexin fusion protein which was produced in soluble form in E. coli BL21 (DE3) induced under optimized conditions. After two purification steps through affinity chromatography, about 1 mg of the recombinant ranalexin was obtained from 1 L of culture. Mass spectrometrical analysis of the purified recombinant ranalexin demonstrated its identity with ranalexin. The purified recombinant ranalexin is biologically active. It showed antibacterial activities similar to those of the native peptide against Staphylococcus aureus, Streptococcus pyogenes, E. coli, and multidrug-resistant strains of S. aureus with minimum inhibitory concentration values between 8 and 128 mu g/ml. The recombinant ranalexin is also cytotoxic in HeLa and COS7 human cancer cells (IC50 = 13-15 mu g/ml).
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