Genetic analysis of acidocin B, a novel bacteriocin produced by Lactobacillus acidophilus

1 TNO Nutrition and Food Research Institute, Department of Molecular Genetics and Gene-technology, PO Box 5815, 2280 HV Rijswijk, The Netherlands 2 Department of Microbiology, PO Box 360, 3700 AJ Zeist, The Netherlands 3 TNO Prevention and Health, PO Box 2215, 2301 CE Leiden, The Netherlands ABSTRACT Summary: The genes encoding the production of acidocin B, a bacteriocin produced by Lactobacillus acidophilus strain M46 which is active against Listeria monocytogenes, Clostridium sporogenes, Brochothrix thermosphacta, Lactobacillus fermentum and Lactobacillus delbrueckii subsp. bulgaricus , but inactive against most other Lactobacillus species, were previously localized on a 4 kb Xba l- Hin dIII fragment of plasmid pCV461. In the present work, DNA sequence analysis revealed the presence of three consecutive ORFs, which potentially code for hydrophobic peptides composed of 60, 91 and 114 amino acids, respectively, and a fourth ORF of opposite polarity which could potentially encode a peptide of 59 amino acids. The middle ORF (ORF-2; acdB ) was identified as the gene encoding acidocin B by comparing the amino acid composition of highly purified acidocin B with the deduced amino acid sequence of ORF-2. Our results suggest that acidocin B is synthesized as a precursor which is processed at a site which conforms to the ' –3, –1' rules of von Heijne to yield active acidocin B (59 amino acids). The presence of an immunity-protein-encoding gene on the 4 kb Xba l- Bam Hl fragment was deduced from the capacity of a plasmid vector harbouring this fragment to confer immunity upon transformation of L. fermentum NCK127. One of the three non-assigned ORFs may encode this immunity protein. 4 Author for correspondence: Rob J. Leer. Tel: +31 15 843171. Fax: +31 15 843989. Keywords: L actobacillus acidophilus , bacteriocin, acidocin B The EMBL accession number for the sequence reported in this paper is 234920.