Identification of a new plasmid-encoded sec-dependent bacteriocin producedby Listeria innocua 743

Listeria innocua 743 produces an inhibitory activity demonstrating broad-sp ectrum inhibition of Listeria monocytogenes isolates. Gel-electrophoretic a nalysis of culture supernatants indicated that two inhibitors with differen t molecular weights were produced by this strain. Insertion of Tn917 into a 2.9 Kb plasmid (pHC743) generated mutants with either an impaired ability or a loss in ability to produce one of the inhibitors. Sequence analysis of the transposon insertion regions revealed the presence of two continuous o pen reading frames, the first encoding a new pediocin-like bacteriocin (lis A) and the second encoding a protein homologous with genes involved in immu nity toward other bacteriocins (lisB). Translation of the bacteriocin gene (lisA) initiates from a noncanonical start codon and encodes a 71-amino-aci d prebacteriocin which lacked the double glycine leader peptidase processin g site common in other type II bacteriocins. Alignment of the sequence with the processed N termini of related bacteriocins suggests that the mature b acteriocin consists of 43 amino acids, with a predicted molecular mass of 4 ,484 Da. Mutants containing insertions into lisA were sensitive to the inhi bitor, indicating that lisAB forms a single operon and that lisB represents the immunity protein. Cloning of an amplicon containing the lisAB operon i nto Escherichia coli resulted in expression and export of the bacteriocin. This finding confirms that the phenotype is dependent on the structural and immunity gene only and that export of this bacteriocin is sec dependent. T his is the first confirmation of bacteriocin production in a Listeria spp., and it is of interest that this bacteriocin is closely related to the pedi ocin family of bacteriocins produced by lactic acid bacteria.