Data mining and characterization of a novel pediocin-like bacteriocin system from the genome of Pediococcus pentosaceus ATCC 25745.
The genome of Pediococcus pentosaceus ATCC 25745 contains a gene cluster that resembles a regulated bacteriocin system. The gene cluster has an operon-like structure consisting of a putative pediocin-like bacteriocin gene (termed penA) and a potential immunity gene (termed peiA). Genetic determinants involved in bacteriocin transport and regulation are also found in proximity to penA and peiA but the so-called accessory gene involved in transport and the inducer gene involved in regulation are missing. Consequently, this bacterium is a poor bacteriocin producer. To analyse the potency of the putative bacteriocin operon, the two genes penA-peiA were heterologously expressed in a Lactobacillus sakei host that contains the complete apparatus for gene activation, maturation and externalization of bacteriocins. It was demonstrated that the heterologous host expressing penA and peiA produced a strong bacteriocin activity; in addition, the host became immune to its own bacteriocin, identifying the gene pair penA-peiA as a potent bacteriocin system. The novel pediocin-like bacteriocin, termed penocin A, has an isotopic mass [M+H]+ of 4684.6 Da as determined by mass spectrometry; this value corresponds well to the expected size of the mature 42 aa peptide containing a disulfide bridge. The bacteriocin is heat-stable but protease-sensitive and has a calculated pI of 9.45. Penocin A has a relatively broad inhibition spectrum, including pathogenic Listeria and Clostridium species. Immediately upstream of the regulatory genes reside some features that resemble remnants of a disrupted inducer gene. This degenerate gene was restored and shown to encode a double-glycine leader-containing peptide. Furthermore, expression of the restored gene triggered high bacteriocin production in P. pentosaceus ATCC 25745, thus confirming its role as an inducer in the pen regulon.
Diep
Dzung B
DB
Laboratory of Microbial Gene Technology, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N1432 Aas, Norway.
Godager
Linda
L
Laboratory of Microbial Gene Technology, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N1432 Aas, Norway.
Brede
Dag
D
Laboratory of Microbial Gene Technology, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N1432 Aas, Norway.
Nes
Ingolf F
IF
Laboratory of Microbial Gene Technology, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N1432 Aas, Norway.
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001430:Bacteriocins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D001483:Base Sequence; D003013:Clostridium / Q000187:drug effects; D015964:Gene Expression Regulation, Bacterial; D016680:Genome, Bacterial; D007778:Lactobacillus / Q000235:genetics / Q000378:metabolism; D008087:Listeria / Q000187:drug effects; D008969:Molecular Sequence Data; D005810:Multigene Family; D010376:Pediococcus / Q000235:genetics* / Q000378:metabolism; D015533:Transcriptional Activation; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D001430:Bacteriocins