Cloning and sequencing of curA encoding curvacin A, the bacteriocin produced by Lactobacillus curvatus LTH1174

Abstract

Curvacin A is a bacteriocin produced by Lactobacillus curvatus LTH1174 which is a potential starter organism for the production of fermented dry sausages. This peptide inhibits the growth of the opportunistic food pathogens Listeria monocytogenes and Enterococcus faecalis and thus, curvacin A may enable better performance of a starter and improvement of the hygienic status of meat products. Oligonucleotides were constructed deduced from the peptide sequence and used for the identification of the curvacin A structural gene curA on a 60 kb plasmid of L. curvatus LTH1174. Plasmid-cured derivatives of this strain were unable to produce curvacin A but were still resistant to the bacteriocin. CurA was cloned into Escherichia coli NM554 and its nucleotide sequence was determined. Sequencing revealed the presence of an additional open reading frame of 51 amino acids with unknown function. A promoter was detected upstream of curA by primer extension. Both reading frames form a single transcript. Curvacin A is synthesised as a prepeptide of 59 amino acids which is proteolytically processed to the mature bacteriocin of 41 amino acids.