title: Each peptide of the two-component lantibiotic lacticin 3147 requires a separate modification enzyme for activity.
abstract: The genetic determinants for production and immunity to the two-component lantibiotic lacticin 3147 are encoded by a 12.6 kb region of the plasmid pMRC01. This region contains ten genes arranged in two divergent clusters; these include the structural genes and a number of genes whose products show significant similarity to proteins involved in the biosynthesis of other lantibiotics. Using a strategy of deletion and mutational analysis, the effect of disruption of a number of these genes was investigated. Inactivation of either of the structural genes, ltnA1 or ltnA2, resulted in mutants that were incapable of producing active lacticin 3147; however, the combination of the cell-free supernatant from both mutants resulted in a restoration of bacteriocin activity, confirming that processing and export of the structural peptides can occur independently. An unusual feature of the lacticin 3147 gene cluster is the presence of two lanM homologues, whose gene products are proposed to be involved in the dehydration and thioether-forming reactions which result in lanthionine bridge formation. Mutants created in the ltnM1 and ltnM2 genes were also incapable of lantibiotic production, confirming an essential role for these enzymes in the lacticin 3147 biosynthetic pathway and supporting the assertion that these proteins are modification enzymes. Interestingly, addition of purified LtnA1, but not purified LtnA2, to the cell-free supernatant of the ltnM1 mutant restored bacteriocin activity; in contrast, only purified LtnA2 could complement the cell-free supernatant of the ltnM2 mutant. Creation of a number of double mutants supported these findings, and confirmed that LtnM1 is required to produce mature LtnA1, while LtnM2 is required to produce mature LtnA2.
authors.affiliation: Department of Microbiology and National Food Biotechnology Centre, University College Cork, Ireland1.
authors.forename: Olivia
authors.initials: O
authors.lastname: McAuliffe
authors.affiliation: Department of Microbiology and National Food Biotechnology Centre, University College Cork, Ireland1.
authors.forename: Colin
authors.initials: C
authors.lastname: Hill
authors.affiliation: National Dairy Products Research Centre, Moorepark, Fermoy, Co. Cork, Ireland2.
authors.forename: R Paul
authors.initials: RP
authors.lastname: Ross
mesh_terms: D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D001430:Bacteriocins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D004352:Drug Resistance, Microbial / Q000235:genetics; D004798:Enzymes / Q000235:genetics* / Q000378:metabolism*; D017353:Gene Deletion; D005798:Genes, Bacterial; D013294:Lactococcus lactis / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011498:Protein Precursors / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D001430:Bacteriocins; D004798:Enzymes; D010455:Peptides; D011498:Protein Precursors; C082508:lacticin 481