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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,
Luis E. N.
Quadri,1,
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.
Present address: Cornell University, Department of Food Science and
Technology, New York State Agricultural Experiment Station, Geneva, NY
14456-0462.

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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