Purification and antibacterial activity of recombinant warnericin RK expressed in Escherichia coli. - Archive ouverte HAL Accéder directement au contenu
Article Dans Une Revue Applied Microbiology and Biotechnology Année : 2012

Purification and antibacterial activity of recombinant warnericin RK expressed in Escherichia coli.

Résumé

Warnericin RK is a small cationic peptide produced by Staphylococcus warneri RK. This peptide has an antimicrobial spectrum of activity almost restricted to the Legionella genus. It is a membrane-active peptide with a proposed detergent-like mechanism of action at high concentration. Moreover, the fatty acids content of Legionella was shown to modulate the peptide activity. In order to decipher the mode of action in details using solid-state NMR spectroscopy, large amount of an isotopic labeled peptide is required. Since it is less expensive to obtain such a peptide biologically, we report here methods to express warnericin RK in Escherichia coli with or without a fusion partner and to purify resulting recombinant peptides. The cDNA fragment encoding warnericin RK was synthesized and ligated into three expression vectors. Two fusion peptides, carrying polyhistidine tag in N- or C-terminal and a native peptide, without tag, were expressed in E. coli cells. Fusion peptides were purified, with a yield of 3 mg/l, by affinity chromatography and reverse-phase HPLC. The recombinant native peptide was purified using a two-step purification method consisting of a hydrophobic chromatography followed by a reverse-phase HPLC step with a yield of 1.4 mg/l. However, the anti-Legionella activity was lower for both tagged peptide probably because of structural modifications. So, the native recombinant peptide was preferentially chosen for (15)N-labeling experiments. Our results suggest that the developed production and purification procedures will be useful in obtaining a large quantity of recombinant isotope-labeled warnericin RK for further studies.

Dates et versions

hal-00763552 , version 1 (11-12-2012)

Identifiants

Citer

Julien Verdon, Nicolas Girardin, Adrienne Marchand, Yann Héchard, Jean-Marc Berjeaud. Purification and antibacterial activity of recombinant warnericin RK expressed in Escherichia coli.. Applied Microbiology and Biotechnology, 2012, epub ahead of print. ⟨10.1007/s00253-012-4417-1⟩. ⟨hal-00763552⟩
91 Consultations
0 Téléchargements

Altmetric

Partager

Gmail Facebook X LinkedIn More