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Applied and Environmental Microbiology, May 1999, p. 2170-2178, Vol. 65, No. 5
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Atypical Genetic Locus Associated with Constitutive Production of Enterocin B by Enterococcus faecium BFE 900

Charles M. A. P. Franz,1 Randy W. Worobo,1,dagger Luis E. N. Quadri,1,Dagger Ulrich Schillinger,2 Wilhelm H. Holzapfel,2 John C. Vederas,3 and Michael E. Stiles1,*

Departments of Agricultural, Food and Nutritional Science1 and Chemistry,3 University of Alberta, Edmonton, Alberta T6G 2P5, Canada, and Federal Research Center for Nutrition, Institute of Hygiene and Toxicology, D-76131 Karlsruhe, Germany2

Received 5 November 1998/Accepted 9 March 1999

A purified bacteriocin produced by Enterococcus faecium BFE 900 isolated from black olives was shown by Edman degradation and mass spectrometric analyses to be identical to enterocin B produced by E. faecium T136 from meat (P. Casaus, T. Nilsen, L. M. Cintas, I. F. Nes, P. E. Hernández, and H. Holo, Microbiology 143:2287-2294, 1997). The structural gene was located on a 2.2-kb HindIII fragment and a 12.0-kb EcoRI chromosomal fragment. The genetic characteristics and production of EntB by E. faecium BFE 900 differed from that described so far by the presence of a conserved sequence like a regulatory box upstream of the EntB gene, and its production was constitutive and not regulated. The 2.2-kb chromosomal fragment contained the hitherto undetected immunity gene for EntB in an atypical orientation that is the reverse of that of the structural gene. Typical transport and other genes associated with bacteriocin production were not detected on the 12.0-kb chromosomal fragment containing the EntB structural gene. This makes the EntB genetic system different from most other bacteriocin systems, where transport and possible regulatory genes are clustered. EntB was subcloned and expressed by the dedicated secretion machinery of Carnobacterium piscicola LV17A. The structural gene was amplified by PCR, fused to the divergicin A signal peptide, and expressed by the general secretory pathway in Enterococcus faecalis ATCC 19433.

* Corresponding author. Mailing address: Department of Agricultural, Food and Nutritional Science, 4-10 Agriculture Forestry Centre, University of Alberta, Edmonton, Alberta T6G 2P5, Canada. Phone: (780) 492-2386. Fax: (780) 492-8914. E-mail: mstiles{at}afns.ualberta.ca.

dagger Present address: Cornell University, Department of Food Science and Technology, New York State Agricultural Experiment Station, Geneva, NY 14456-0462.

Dagger Present address: Harvard Medical School, Department of Biological Chemistry and Molecular Pharmacology, Boston, MA 02146.

Applied and Environmental Microbiology, May 1999, p. 2170-2178, Vol. 65, No. 5
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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