Functional expression of an antimicrobial peptide, belonging to halocin C8 family, from Natrinema sp. RNS21 in Escherichia coli

Halocins, the proteinaceous antimicrobial agents produced by haloarchaea, may be used for the preservation of salted foods and the treatment of diseases. For their application and function explanation, it is necessary to produce the active recombinants. In this work, a haloarchaeal strain producing halocin was isolated from the salt-fermented shrimp and identified as Natrinema sp. RNS21 by 16S rRNA gene sequence analysis. From 1 L of RNS21 culture, about 0.32 mg of halocin with 96% purity was obtained. Based on the molecular weight, stability and amino acid sequence alignment, the antimicrobial peptide belonged to the halocin C8 (HalC8) family. HalC8 was expressed by fusion with glutathione-S-transferase (GST) in E. coli, followed by affinity purification and enterokinase (EK) cleavage. About 6.2 mg of recombinant HalC8 with 95% purity was obtained from 1 L of E. coli culture. MALDI-TOF-MS and RP-HPLC analysis indicated that the molecular weight and folding pattern of purified recombinant HalC8 were the same as those of native HalC8. Recombinant HalC8 showed obvious inhibitory activity against Haloferax volcanii. Contrast to native HalC8, the active recombinant HalC8 could be easily produced in a short time with a high yield.

Automated summary

From a haloarchaeal strain isolated from salt-fermented shrimp, a pure 0.32 mg halocin with 96% purity was obtained. Through fusion with glutathione-S-transferase (GST) in E. coli and affinity purification followed by enterokinase (EK) cleavage, a 6.2 mg recombinant HalC8 with 95% purity was obtained. The recombinant HalC8 showed the same molecular weight, folding pattern, and inhibitory activity against Haloferax volcanii as the native HalC8.