Heterologous expression and purification of NisA, the precursor peptide of lantibiotic nisin from Lactococcus lactis

Karakas-Sen, Asuman and Narbad, A. (2012) Heterologous expression and purification of NisA, the precursor peptide of lantibiotic nisin from Lactococcus lactis. Acta Biologica Hungarica, 63 (2). pp. 301-310. ISSN 0236-5383

Text abiol.63.2012.2.11.pdf Restricted to Repository staff only until 30 June 2032. Download (315kB)

Abstract

The lantibiotic nisin is a ribosomally synthesised and post-translationally modified antimicrobial peptide produced by strains of Lactococcus lactis, and used as safe and natural preservative in food industry. The nisA structural gene encodes ribosomally synthesised and biologically inactive a 57 amino acid precursor peptide (NisA) which undergoes several post-translational modifications. In this study, we report the expression of precursor nisin as a His₆-tagged peptide in Escherichia coli and its purification using a nickel affinity column. The technique of spliced-overlap extension PCR was used to amplify the nisA gene and the T7 promoter region of pET-15b vector. This approach was used to introduce six histidine residues at the C-terminus of prenisin. The identity of the expressed peptide was confirmed by N-terminal sequencing. The expressed His-tagged prenisin was purified under denaturing conditions, and named as prenisin-His₆. The purified prenisin-His6 was analyzed by SDS-PAGE, Western blotting and mass spectroscopy. These results showed that the nisin precursor peptide can be successfully produced using an E. coli expression system.

Actions (login required)

Edit Item