Molecular characterisation of aureocin A70, a multipeptide bacteriocin isolated from Staphylococcus aureus

Staphylococcus aureus A70 produces a heat-stable bacteriocin designated aur eocin A70. Aureocin A70 is encoded within a mobilisable 8 kb plasmid, pRJ6, and is active against Listeria monocytogenes. Experiments of transposition mutagenesis and gene cloning had shown that aureocin A70 production and im munity were associated with the HindIII-A and B fragments of pRJ6. Therefor e, a 6332 bp region of the plasmid, encompassing both these fragments, was sequenced using a concatenation DNA sequencing procedure. DNA sequence and genetic analyses revealed the presence of three transcriptional units that appear to be involved in bacteriocin activity. The first transcriptional un it contains a single gene, aurT, which encodes a protein that resembles an ATP-dependent transporter, similar to those involved in lantibiotic export. AurT is required for aureocin A70 production and it appears to be essentia l for mobilisation of pRJ6. The second putative operon contains two open re ading frames (ORFs); the first gene, orfA, is predicted to encode a protein similar to small repressor proteins found in some Archaea, whose function remains to be elucidated. The second gene, orfB, codes for an 138 amino aci d residue protein which shares a number of characteristics (high pI and hyd rophobicity profile) with proteins associated with immunity, needed for sel f-protection against bacteriocin. Four other genes are present in the third operon, aurABCD. aurABCD encode four related peptides that are small (30-3 1 amino acid residues), strongly cationic (pI of 9.85 to 10.04) and highly hydrophobic. Theses peptides also have a high content of small amino acid r esidues like glycine and alanine, and no cysteine residue. Tn917-lac insert ional mutations, which affected aureocin A70 activity, reside within operon aurABCD. Analysis of purified bacteriocin preparations by mass spectrometr y demonstrated that all four peptides encoded by aurABCD operon are produce d, expressed and excreted without post-translational modifications. Thus, a ureocin A70 is a multi-peptide non-lantibiotic bacteriocin, which is transp orted without processing. (C) 2001 Academic Press.